WO2022165092A1 - Méthodes de traitement de maladies fibrotiques - Google Patents

Méthodes de traitement de maladies fibrotiques Download PDF

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Publication number
WO2022165092A1
WO2022165092A1 PCT/US2022/014185 US2022014185W WO2022165092A1 WO 2022165092 A1 WO2022165092 A1 WO 2022165092A1 US 2022014185 W US2022014185 W US 2022014185W WO 2022165092 A1 WO2022165092 A1 WO 2022165092A1
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fold
compound
biomarkers
patient
animals
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PCT/US2022/014185
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Prakash Narayan
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Angion Biomedica Corp.
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    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P9/00—Drugs for disorders of the cardiovascular system
    • 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
    • A61K31/496—Non-condensed piperazines containing further heterocyclic rings, e.g. rifampin, thiothixene or sparfloxacin
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K9/00—Medicinal preparations characterised by special physical form
    • A61K9/0012—Galenical forms characterised by the site of application
    • A61K9/0053—Mouth and digestive tract, i.e. intraoral and peroral administration
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K9/00—Medicinal preparations characterised by special physical form
    • A61K9/48—Preparations in capsules, e.g. of gelatin, of chocolate
    • A61K9/4841—Filling excipients; Inactive ingredients
    • A61K9/4866—Organic macromolecular compounds
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P11/00—Drugs for disorders of the respiratory system
    • 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
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
    • G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
    • G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
    • G01N33/68—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving proteins, peptides or amino acids
    • G01N33/6887—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving proteins, peptides or amino acids from muscle, cartilage or connective tissue
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N2333/00—Assays involving biological materials from specific organisms or of a specific nature
    • G01N2333/435—Assays involving biological materials from specific organisms or of a specific nature from animals; from humans
    • G01N2333/78—Connective tissue peptides, e.g. collagen, elastin, laminin, fibronectin, vitronectin, cold insoluble globulin [CIG]
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N2800/00—Detection or diagnosis of diseases
    • G01N2800/04—Endocrine or metabolic disorders
    • G01N2800/044—Hyperlipemia or hypolipemia, e.g. dyslipidaemia, obesity
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N2800/00—Detection or diagnosis of diseases
    • G01N2800/08—Hepato-biliairy disorders other than hepatitis
    • G01N2800/085—Liver diseases, e.g. portal hypertension, fibrosis, cirrhosis, bilirubin
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N2800/00—Detection or diagnosis of diseases
    • G01N2800/12—Pulmonary diseases
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N2800/00—Detection or diagnosis of diseases
    • G01N2800/34—Genitourinary disorders
    • G01N2800/347—Renal failures; Glomerular diseases; Tubulointerstitial diseases, e.g. nephritic syndrome, glomerulonephritis; Renovascular diseases, e.g. renal artery occlusion, nephropathy

Definitions

  • Fibrotic diseases contribute to as much as 45% of deaths in the industrialized world (Wynn, T. A. J. Pathol. 2008;214: 199-210). Fibrosis, i.e., excessive tissue scarring, is a common feature of many chronic diseases, as well as acute injuries, where accumulation of scar tissue can lead to loss of organ function and, ultimately, organ failure (Friedman, S. L., et al. Sci. Transl. Med. 2019 Jan 9;5(167): 167srl).
  • fibrosis of the kidney can result in progressive loss of renal function, which can lead to end stage renal failure. Renal failure is fatal without regular dialysis or a kidney transplant. There exists a continuing need for improved treatments for fibrotic diseases of the kidney.
  • the present disclosure provides methods related to treatment of fibrotic disease(s) (e.g., as described herein, such as kidney fibrotic disease(s) or lung fibrotic disease(s)) and selecting, identifying, and/or characterizing patients likely to benefit from treatment with Compound 1 :
  • the present disclosure is based in part on the recognition that certain biomarkers can distinguish patients who are likely to respond to therapy, for example because the drivers of their fibrotic disease correspond with the mechanism of action of Compound 1.
  • the drivers of their kidney disease correspond with the mechanism of action of Compound 1.
  • a patient to be treated with a method of the present disclosure has an altered level of one or more gene products and/or one or more proteins (or fragments thereof) that are associated with the mechanism of action of Compound 1.
  • the present disclosure also encompasses the recognition that certain biomarkers can distinguish patients (e.g., patients with certain symptoms related to a fibrotic disease described herein, e.g., a kidney disease) who are likely to respond to therapy, regardless of disease pathology.
  • patients who present with nephrotic syndrome and/or proteinuria and/or hypoalbuminemia and/or hyperlipidemia and/or edema may be suffering from or susceptible to a variety of different kidney diseases or conditions.
  • the present disclosure provides methods of identifying and/or selecting among patients with such symptoms those patients who are likely to respond to Compound 1 therapy.
  • the present disclosure provides insights that a “signature” comprising levels of a plurality of biomarkers can be useful for methods provided herein (e.g., selecting, identifying and/or characterizing patients who are likely to respond to Compound 1 therapy). For example, in some embodiments, when a patient’s signature comprises levels of certain biomarkers that are different from corresponding threshold levels, that patient is likely to respond to Compound 1. In some embodiments, when a patient’s signature comprises levels of at least about 10%, about 20%, about 30%, about 40%, about 50%, about 60%, about 70%, about 80%, or about 90% of biomarkers that are altered relative to corresponding threshold levels, that patient is likely to respond to Compound 1.
  • the plurality of biomarkers that comprise a patient’s signature are selected from those described herein, including classes and subclasses described herein, both singly and in combination. In some embodiments, such methods further comprise administering an effective amount of Compound 1.
  • the present disclosure encompasses the recognition that one or more collagen biomarkers may be useful for methods provided herein (e.g., selecting, identifying and/or characterizing patients who are likely to respond to Compound 1 therapy). For example, in some embodiments, when a patient has been determined to have an elevated level of one or more collagen biomarkers (including, e.g., a collagen 6 biomarker), that patient is likely to respond to Compound 1.
  • one or more collagen biomarkers may be useful for methods provided herein (e.g., selecting, identifying and/or characterizing patients who are likely to respond to Compound 1 therapy). For example, in some embodiments, when a patient has been determined to have an elevated level of one or more collagen biomarkers (including, e.g., a collagen 6 biomarker), that patient is likely to respond to Compound 1.
  • such methods further comprise administering an effective amount of Compound 1.
  • the present disclosure provides methods of treating patients diagnosed with, suspected of having, or at risk of a fibrotic disease (e.g., of the kidney), comprising (i) obtaining or determining a level of one or more biomarkers (e.g., biomarkers described herein) in a biological sample obtained from the patients; and (ii) comparing the level with that of a corresponding threshold level.
  • methods further comprise administering Compound 1 therapy if the level of one or more biomarkers is different from the corresponding threshold level.
  • the present disclosure provides methods of monitoring therapy with Compound 1.
  • methods comprising (i) administering an effective amount of Compound 1; and (ii) monitoring the levels of one or more biomarkers (e.g., biomarkers described herein).
  • biomarkers e.g., biomarkers described herein.
  • such methods further comprise adjusting Compound 1 therapy based on the results (e.g., discontinuing Compound 1 therapy, changing dose and/or dosing frequency of Compound 1, etc.).
  • FIG. 1 is a graph showing results of Compound 1 treatment on urine protein in a PAN and uninephrectomy rat model of kidney disease.
  • FIG. 2A and FIG. 2B are graphs showing change from baseline in urine protein per animal on Study Day 17 (Treatment Day 14).
  • FIG. 3A is a graph showing lung-to-body weight ratio of mice treated with Compound 1 in a bleomycin-induced injury model of idiopathic pulmonary fibrosis (IPF).
  • FIG. 3B is a graph showing hydroxyproline levels in mice treated with Compound 1 in a bleomycin- induced injury model of IPF.
  • FIG. 3C is a graph showing histopathological damage in lung tissue sections using the Ashcroft scale from mice treated with Compound 1 in a bleomycin- induced injury model of IPF.
  • FIG. 3D is a graph showing extent of picrosirius red staining in lung tissue sections from mice treated with Compound 1 in a bleomycin-induced injury model of IPF.
  • FIG. 3E is a graph showing extent of immunohistological staining for TGF ⁇ 1 in lung tissue sections from mice treated with Compound 1 in a bleomycin-induced injury model of IPF.
  • FIG. 4A is a graph showing lung fibrosis score using Ashcroft scale based on H&E staining in lung tissue sections from mice treated with Compound 1 in an inducible TGF ⁇ 1 mouse model of lung fibrosis.
  • FIG. 4B is a graph hydroxyproline levels in mice treated with Compound 1 in an inducible TGF ⁇ 1 mouse model of lung fibrosis.
  • FIG. 4C is a graph showing extent of picrosirius red staining in lung tissue sections from mice treated with Compound 1 in an inducible TGF ⁇ 1 mouse model of lung fibrosis.
  • FIG. 4D is a graph showing extent of aSMA staining in lung tissue sections from mice treated with Compound 1 in an inducible TGF ⁇ 1 mouse model of lung fibrosis.
  • FIG. 5A is a graph showing dermal thickness, measured as the distance between epidermal-dermal junction and dermal-subcutaneous fat junction in H&E-stained slides prepared from a skin biopsy from mice treated with Compound 1 in a bleomycin systemic sclerosis mouse model.
  • FIG. 5B is a graph showing skin fibrotic score, judged based on H&E-stained slides prepared from a skin biopsy, from mice treated with Compound 1 in a bleomycin systemic sclerosis mouse model.
  • FIG. 5C is a graph showing lung hydroxyproline levels from mice treated with Compound 1 in a bleomycin systemic sclerosis mouse model.
  • FIG. 5D is a graph showing lung fibrosis score using Ashcroft scale based on H&E staining in lung tissue sections from mice treated with Compound 1 in a bleomycin systemic sclerosis mouse model.
  • FIG. 5E is a graph showing kidney hydroxyproline levels from mice treated with Compound 1 in a bleomycin systemic sclerosis mouse model.
  • FIG. 5F is a graph showing extent of picrosirius red staining in kidney tissue sections from mice treated with Compound 1 in a bleomycin systemic sclerosis mouse model.
  • FIG. 6A is a graph showing urine protein levels in rats treated with Compound 1 in a FSGS-relevant model of PAN-induced proteinuria.
  • FIG. 6B is a graph showing intraperitoneal fluid volume in rats treated with Compound 1 in a FSGS-relevant model of PAN-induced proteinuria.
  • FIG. 6C is a graph showing GFR, measured using FITC-sinistrin decay kinetics, in rats treated with Compound 1 in a FSGS-relevant model of PAN-induced proteinuria.
  • FIG. 6D is a graph showing glomerular diameter, measured using histopathological analysis of PAS- stained renal coronal sections from rats treated with Compound 1 in a FSGS-relevant model of PAN-induced proteinuria.
  • FIG. 6E is a graph showing collagen-3 expression, determined in glomeruli using quantitative immunohistochemistry, in rats treated with Compound 1 in a FSGS- relevant model of PAN-induced proteinuria.
  • FIG. 7A is a graph showing urine protein levels in rats treated with Compound 1 in a DOCA/salt model of renal fibrosis and injury.
  • FIG. 7B is a graph showing urine albumin levels in rats treated with Compound 1 in a DOCA/salt model of renal fibrosis and injury.
  • FIG. 7C is a graph showing urine albumin to creatinine ratio in rats treated with Compound 1 in a DOCA/salt model of renal fibrosis and injury.
  • FIG. 7D is a graph showing urine kidney injury molecule- 1 (KIMI) levels in rats treated with Compound 1 in a DOCA/salt model of renal fibrosis and injury.
  • FIG. KIMI urine kidney injury molecule- 1
  • FIG. 7E is a graph showing renal hydroxyproline levels in rats treated with Compound 1 in a DOCA/salt model of renal fibrosis and injury.
  • FIG. 7F is a graph showing renal damage scores, based on H&E stained slides of kidney tissue sections from rats treated with Compound 1 in a DOCA/salt model of renal fibrosis and injury.
  • FIG. 7G is a graph showing extent of picrosirius red staining from slides of kidney tissue sections from rats treated with Compound 1 in a DOCA/salt model of renal fibrosis and injury.
  • FIG. 7H is a graph showing extent of aSMA staining in kidney tissue sections from rats treated with Compound 1 in a DOCA/salt model of renal fibrosis and injury.
  • UUO unilateral ureteral obstruction
  • FIG. 9A is a graph showing serum creatinine (SCr) levels in rats treated with Compound 1 in a PCK rat model of polycystic kidney disease.
  • FIG. 9B is a graph showing BUN levels in rats treated with Compound 1 in a PCK rat model of polycystic kidney disease.
  • FIG. 9C is a graph showing kidney weight at sacrifice in rats treated with Compound 1 in a PCK rat model of polycystic kidney disease.
  • FIG. 9D is a graph showing kidney weight as a percentage of body weight at sacrifice of rats treated with Compound 1 in a PCK rat model of polycystic kidney disease.
  • FIG. 9A is a graph showing serum creatinine (SCr) levels in rats treated with Compound 1 in a PCK rat model of polycystic kidney disease.
  • FIG. 9B is a graph showing BUN levels in rats treated with Compound 1 in a PCK rat model of polycystic kidney disease.
  • FIG. 9C is
  • FIG. 9E is a graph showing kidney hydroxyproline levels in rats treated with Compound 1 in a PCK rat model of polycystic kidney disease.
  • FIG. 9F is a graph showing cystic index, quantified in H&E stained kidney tissue sections using digital planimetry, in rats treated with Compound 1 in a PCK rat model of polycystic kidney disease.
  • FIG. 9G is a graph showing 24-hour urine volume in rats treated with Compound 1 in a PCK rat model of polycystic kidney disease.
  • FIG. 9H is a graph showing urine protein levels in rats treated with Compound 1 in a PCK rat model of polycystic kidney disease.
  • FIG. 91 is a graph showing urine albumin levels in rats treated with Compound 1 in a PCK rat model of polycystic kidney disease.
  • FIG. 9J is a graph showing urine neutrophil gelatinase-associated lipocalin (NGAL) levels in rats treated with Compound 1 in a PCK rat model of polycystic kidney disease.
  • FIG. 9K is a graph showing urine KIMI levels in rats treated with Compound 1 in a PCK rat model of polycystic kidney disease.
  • FIG. 9L is a graph showing urine interleukin 18 (IL 18) levels in rats treated with Compound 1 in a PCK rat model of polycystic kidney disease.
  • FIG. 9M is a graph showing urine cystatin C levels in in rats treated with Compound 1 in a PCK rat model of polycystic kidney disease.
  • FIG. 10A is a graph showing body weight after sacrifice in mice treated with Compound 1 in a TNBS model of inflammatory bowel disease/acute colitis.
  • FIG. 10B is a graph showing colon weight after sacrifice in mice treated with Compound 1 in a TNBS model of inflammatory bowel disease/acute colitis.
  • FIG. 10C is a graph showing colon length after sacrifice in mice treated with Compound 1 in a TNBS model of inflammatory bowel disease/acute colitis.
  • FIG. 10D is a graph showing colon damage score in mice treated with Compound 1 in a TNBS model of inflammatory bowel disease/acute colitis.
  • FIG. 10A is a graph showing body weight after sacrifice in mice treated with Compound 1 in a TNBS model of inflammatory bowel disease/acute colitis.
  • FIG. 10B is a graph showing colon weight after sacrifice in mice treated with Compound 1 in a TNBS model of inflammatory bowel disease/acute colitis.
  • FIG. 10C is a graph showing
  • FIG. 10E is a graph showing colon histopathological scores of H&E-stained colon tissue sections from mice treated with Compound 1 in a TNBS model of inflammatory bowel disease/acute colitis. Each of (a) colon architecture, (b) degree of inflammatory cell infiltration, (c) muscle thickening, and (d) crypt damage and goblet cells loss was assessed.
  • FIG. 10F is a graph showing composite colon histopathological scores of H&E-stained colon tissue sections from mice treated with Compound 1 in a TNBS model of inflammatory bowel disease/acute colitis.
  • FIG. 10E is a graph showing colon histopathological scores of H&E-stained colon tissue sections from mice treated with Compound 1 in a TNBS model of inflammatory bowel disease/acute colitis.
  • FIG. 10G is a graph showing extent of Alcian blue staining in slides of colon tissue sections from mice treated with Compound 1 in a TNBS model of inflammatory bowel disease/acute colitis.
  • FIG. 10H is a graph showing extent of myeloperoxidase staining in slides of colon tissue sections from mice treated with Compound 1 in a TNBS model of inflammatory bowel disease/acute colitis.
  • FIG. 101 is a graph showing extent of F4/80 staining in slides of colon tissue sections from mice treated with Compound 1 in a TNBS model of inflammatory bowel disease/acute colitis.
  • FIG. 11A is a graph showing body weight in mice prior to treatment with Compound 1 in an acetic acid induced colitis model.
  • FIG. 11B is a graph showing colon weight in mice prior to treatment with Compound 1 in an acetic acid induced colitis model.
  • FIG. 11C is a graph showing colon length in mice prior to treatment with Compound 1 in an acetic acid induced colitis model.
  • FIG. 11D is a graph showing gross morphological colon damage score in mice prior to treatment with Compound 1 in an acetic acid induced colitis model.
  • FIG. HE is a graph showing colon length in mice treated with Compound 1 in an acetic acid induced colitis model.
  • FIG. 11F is a graph showing gross morphological colon damage score in mice treated with Compound 1 in an acetic acid induced colitis model.
  • FIG. 11G is a graph showing histopathological colon damage score in mice treated with Compound 1 in an acetic acid induced colitis model.
  • FIG. 11H is a graph showing extent of Alcian blue staining in slides of colon tissue sections from mice treated with Compound 1 in an acetic acid induced colitis model.
  • FIG. 12A is a graph showing colon length in mice treated with Compound 1 in a dextran sulfate sodium model of chronic colitis in inflammatory bowel disease.
  • FIG. 12A is a graph showing colon length in mice treated with Compound 1 in a dextran sulfate sodium model of chronic colitis in inflammatory bowel disease.
  • FIG. 12B is a graph showing macroscopic colon damage score in mice treated with Compound 1 in a dextran sulfate sodium model of chronic colitis in inflammatory bowel disease.
  • FIG. 12C is a graph showing colon hydroxyproline levels in mice treated with Compound 1 in a dextran sulfate sodium model of chronic colitis in inflammatory bowel disease.
  • FIG. 12D is a graph showing histopathological injury score from H&E-stained slides of colon tissue section from mice treated with Compound 1 in a dextran sulfate sodium model of chronic colitis in inflammatory bowel disease.
  • FIG. 13A is a graph showing serum pro-collagen 6 levels in animals treated with Compound 1 compared to sham and control animals.
  • FIG. 13B is a graph showing a correlation between serum pro-collagen 6 and TGF- ⁇ levels in animals evaluated in this Example 17.
  • FIG. 13C is a graph showing serum pro-collagen 6 levels in animals treated with Compound 1 compared to sham and control animals in the PANX model.
  • FIG. 13D is graph showing serum pro-collagen 6 levels in animals treated with Compound 1 compared to sham and control animals in the DOC A model.
  • administering typically refers to administration of a composition to a subject to achieve delivery of an active agent to a site of interest (e.g., a target site which may, in some embodiments, be a site of disease or damage, and/or a site of responsive processes, cells, tissues, etc.)
  • a site of interest e.g., a target site which may, in some embodiments, be a site of disease or damage, and/or a site of responsive processes, cells, tissues, etc.
  • a site of interest e.g., a target site which may, in some embodiments, be a site of disease or damage, and/or a site of responsive processes, cells, tissues, etc.
  • a site of interest e.g., a target site which may, in some embodiments, be a site of disease or damage, and/or a site of responsive processes, cells, tissues, etc.
  • one or more particular routes of administration may be feasible and/or useful in the practice of the present invention.
  • administration may be parenteral
  • administration may involve dosing that is intermittent (e.g., a plurality of doses separated in time) and/or periodic (e.g., individual doses separated by a common period of time) dosing.
  • intermittent e.g., a plurality of doses separated in time
  • periodic e.g., individual doses separated by a common period of time
  • dosing that is intermittent (e.g., a plurality of doses separated in time) and/or periodic (e.g., individual doses separated by a common period of time) dosing.
  • the term “comparable” refers to two or more agents, entities, situations, sets of conditions, circumstances, individuals, or populations, etc., that may not be identical to one another but that are sufficiently similar to permit comparison there between so that one skilled in the art will appreciate that conclusions may reasonably be drawn based on differences or similarities observed.
  • comparable agents, entities, situations, sets of conditions, circumstances, individuals, or populations are characterized by a plurality of substantially identical features and one or a
  • dosage form may be used to refer to a physically discrete unit of an active agent (e.g., a therapeutic agent) for administration to a subject.
  • an active agent e.g., a therapeutic agent
  • each such unit contains a predetermined quantity of active agent.
  • such quantity is a unit dosage amount (or a whole fraction thereof) appropriate for administration in accordance with a dosing regimen that has been determined to correlate with a desired or beneficial outcome when administered to a relevant population (i.e., with a therapeutic dosing regimen).
  • the total amount of a therapeutic composition or agent administered to a particular subject is determined by one or more attending physicians and may involve administration of multiple dosage forms.
  • salt form refers to a form of a relevant compound as a salt appropriate for use in pharmaceutical contexts, i.e., salts which are, within the scope of sound medical judgment, suitable for use in contact with the tissues of humans and/or lower animals without undue toxicity, irritation, allergic response and the like, and are commensurate with a reasonable benefit/risk ratio.
  • Pharmaceutically acceptable salts are well known in the art. For example, S. M. Berge, et al. describes pharmaceutically acceptable salts in detail in J. Pharmaceutical Sciences, 66: 1-19 (1977).
  • the term “reference” describes a standard or control relative to which a comparison is performed. For example, in some embodiments, an agent, individual, population, sample, sequence or value of interest is compared with a reference or control agent, individual, population, sample, sequence or value. In some embodiments, a reference or control is tested and/or determined substantially simultaneously with the testing or determination of interest. In some embodiments, a reference or control is a historical reference or control, optionally embodied in a tangible medium. Typically, as would be understood by those skilled in the art, a reference or control is determined or characterized under comparable conditions or circumstances to those under assessment. Those skilled in the art will appreciate when sufficient similarities are present to justify reliance on and/or comparison to a particular possible reference or control.
  • risk of a disease, disorder, and/or condition refers to a likelihood that a particular individual will develop the disease, disorder, and/or condition. In some embodiments, risk is expressed as a percentage. In some embodiments, risk is from 0, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 20, 30, 40, 50, 60, 70, 80, 90 up to 100%. In some embodiments risk is expressed as a risk relative to a risk associated with a reference sample or group of reference samples. In some embodiments, a reference sample or group of reference samples have a known risk of a disease, disorder, condition and/or event. In some embodiments a reference sample or group of reference samples are from individuals comparable to a particular individual. In some embodiments, relative risk is 0, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, or more.
  • the term “subject” refers an organism, typically a mammal (e.g., a human).
  • a subject is suffering from a relevant disease, disorder or condition.
  • a human subject is an adult, adolescent, or pediatric subject.
  • a subject is at risk of (e.g., susceptible to), e.g., at elevated risk of relative to an appropriate control individual or population thereof, a disease, disorder, or condition.
  • a subject displays one or more symptoms or characteristics of a disease, disorder or condition.
  • a subject does not display any symptom or characteristic of a disease, disorder, or condition.
  • a subject is someone with one or more features characteristic of susceptibility to or risk of a disease, disorder, or condition.
  • a subject is an individual to whom diagnosis and/or therapy and/or prophylaxis is and/or has been administered.
  • the terms “subject” and “patient” are used interchangeably herein.
  • the term “treat” refers to any administration of a therapy that partially or completely alleviates, ameliorates, relieves, inhibits, delays onset of, reduces severity of, and/or reduces incidence of one or more symptoms, features, and/or causes of a particular disease, disorder, and/or condition.
  • such treatment may be of a subject who does not exhibit signs of the relevant disease, disorder and/or condition and/or of a subject who exhibits only early signs of the disease, disorder, and/or condition.
  • such treatment may be of a subject who exhibits one or more established signs of the relevant disease, disorder and/or condition.
  • treatment may be of a subject who has been diagnosed as suffering from the relevant disease, disorder, and/or condition.
  • Compound 1 i.e., methyl (Z)-3-(((4-(N-methyl-2-(4-methylpiperazin-l-yl)acetamido)phenyl) amino)(phenyl)methylene)-2-oxo-2,3-dihydro-lH-pyrrolo[2,3-b]pyridine-6-carboxylate, is in a pharmacological class of tyrosine kinase inhibitors (TKI).
  • TKI tyrosine kinase inhibitors
  • Compound 1 is an orally bioavailable small molecule dual kinase inhibitor of platelet-derived growth factor receptors (PDGFR) and vascular endothelial growth factor receptors (VEGFR2). Compound 1 is useful in methods provided herein.
  • Compound 1 is provided and/or utilized (e.g., for inclusion in a composition and/or for delivery to a subject) in accordance with the present disclosure in a form such as a pharmaceutically acceptable salt form.
  • a pharmaceutically acceptable salt form As already noted herein, pharmaceutically acceptable salt forms are well known in the art.
  • Compound 1 is provided and/or utilized in a hydrochloride salt form, a maleate salt form, a mesylate salt form, or a tosylate salt form.
  • Compound 1 is provided and/or utilized (e.g., for inclusion in, e.g., during one or more steps of manufacturing of, a composition and/or for delivery to a subject) in accordance with the present disclosure in a form such as a solid form.
  • Compound 1 is provided and/or utilized in accordance with the present disclosure in an amorphous solid form, in a crystalline solid form, or in a mixture thereof.
  • a crystalline solid form may be or comprise a solvate, hydrate, or an unsolvated form. The use of any and all such forms are contemplated by the present disclosure.
  • Compound 1 is provided and/or utilized (e.g., for inclusion in, e.g., during one or more steps of manufacturing of, a composition and/or for delivery to a subject) as a hydrochloride salt form (e.g., a channel hydrate comprising up to about 4 equivalents of water, up to about 3 equivalents of water, up to about 2 equivalents of water, or up to about 1 equivalent of water).
  • a hydrochloride salt form of Compound 1 is characterized by one or more peaks in its XRPD selected from those at about 5.28, 10.63, 11.54, 17.05, and 20.98 degrees 2-theta.
  • such a hydrochloride salt form of Compound 1 is characterized by two or more peaks in its XRPD selected from those at about 5.28, 10.63, 11.54, 17.05, and 20.98 degrees 2-theta. In some embodiments, such a hydrochloride salt form of Compound 1 is characterized by three or more peaks in its XRPD selected from those at about 5.28, 10.63, 11.54, 17.05, and 20.98 degrees 2-theta. In some embodiments, such a hydrochloride salt form of Compound 1 is characterized by peaks in its XRPD pattern at about 5.28, 10.63, 11.54, 17.05, and 20.98 degrees 2-theta.
  • such a hydrochloride salt form of Compound 1 is characterized by peaks in its XRPD pattern at about 5.28, 10.63, 11.54, 17.05, and 20.98 degrees 2-theta, corresponding to d- spacing of about 16.74, 8.33, 7.67, 5.20, and 4.23 angstroms, respectively.
  • such a hydrochloride salt form of Compound 1 is characterized by substantially all of the peaks in its XRPD pattern selected from 5.28, 5.76, 10.63, 11.54, 12.73, 13.13, 14.08, 15.34, 15.64, 16.00, 16.55, 17.05, 17.78, 18.86, 19.08, 20.16, 20.68, 20.98, 21.62, 22.05, 22.82, 23.97, 24.94, 25.23, 25.61, 26.58, 27.01, 27.78, 29.89, 30.51, 30.91, 31.72, and 33.39 degrees 2- theta.
  • Compound 1 refers to Compound 1 (i.e., methyl (Z)-3-(((4-(N-methyl-2-(4-methylpiperazin-l- yl)acetamido)phenyl)amino)(phenyl)methylene)-2-oxo-2,3-dihydro-lH-pyrrolo[2,3-b]pyridine- 6-carboxylate) in any available form, such as, e.g., a salt form and/or solid form. It will be understood, therefore, that reference to an amount (e.g., in mg) of Compound 1 means the amount of Compound 1 in free base form.
  • Compound 1 may be provided and/or utilized as, e.g., a salt form of Compound 1 such that the amount of the salt (or other form) is an amount that corresponds to the “free base equivalent” of Compound 1.
  • “50 mg Compound 1” means, e.g., approx. 53.4 mg of Compound 1 Hydrochloride anhydrate, approx. 58.4 mg of Compound 1 Hydrochloride trihydrate, and approx. 58.9 mg of Compound 1 Mesylate anhydrate, etc.
  • the present disclosure provides certain biomarkers that can distinguish subjects (e.g., subjects suffering from or at risk of fibrosis, such as renal fibrosis, or associated diseases, disorders, and conditions) who are more likely than others to respond to therapy with Compound 1.
  • the present disclosure provides the insight that certain biomarkers can distinguish patients who are likely to respond to therapy, for example, because the drivers of their fibrotic disease correspond with the mechanism of action of Compound 1.
  • the drivers of their kidney disease correspond with the mechanism of action of Compound 1.
  • an altered level e.g., an expression level
  • a patient with an altered level of one or more biomarkers may have an improved response to treatment with Compound 1 relative to a patient that does not have a level of the biomarker that meets the threshold criteria.
  • a biomarker is a component of a biological sample that may be detected and/or quantified when present in the biological sample.
  • a biomarker may include one or more of a peptide, protein, nucleic acid (e.g., polynucleotide, DNA, RNA, etc.), polysaccharide (e.g., lectins or sugars), lipid, enzyme, small molecule, ligand, receptor, antigen, or antibody.
  • a biomarker comprises a protein.
  • a biomarker comprises a nucleic acid (e.g., mRNA).
  • a level of a biomarker corresponds to a level of gene expression (e.g., RNA expression, e.g., mRNA expression). In some certain embodiments, a level of a biomarker corresponds to a level of protein expression, including any fragment or degradation product thereof.
  • a biological fluid may be or comprise an intracellular fluid, an extracellular fluid, an intravascular fluid (blood plasma), an interstitial fluid, a lymphatic fluid, and/or a transcellular fluid.
  • a biological tissue or sample may be obtained, for example, by aspirate, biopsy (e.g., fine needle or tissue biopsy), swab (e.g., oral, nasal, skin, or vaginal swab), scraping, surgery, washing or lavage (e.g., bronchoalveolar, ductal, nasal, ocular, oral, uterine, vaginal, or other washing or lavage).
  • a biomarker is detected and/or quantified in a tissue sample (e.g., from a biopsy, such as a liver or kidney or lung biopsy) and/or in a biological fluid (e.g., blood, urine, BALF, etc.).
  • a biomarker e.g., a level of mRNA
  • a tissue sample e.g., obtained from a biopsy, e.g., a kidney biopsy.
  • a biomarker e.g., a level of mRNA
  • a kidney tissue sample e.g., obtained from a kidney biopsy.
  • a biomarker e.g., a level of mRNA
  • a liver tissue sample e.g., obtained from a liver biopsy.
  • a biomarker e.g., a level of mRNA
  • a lung tissue sample e.g., obtained from a lung biopsy.
  • a biomarker e.g., a level of a protein or protein fragment
  • a urine sample e.g., a level of a protein or protein fragment
  • a biomarker e.g., a level of a protein or protein fragment
  • a blood sample e.g., a level of a protein or protein fragment
  • a BALF sample e.g., a level of a protein or protein fragment
  • one biomarker is used to characterize subjects; in some embodiments, more than one biomarker (e.g., two, three, etc.) is used to characterize subjects. In some embodiments, a “signature” comprising levels of a plurality of biomarkers is used to characterize subjects.
  • biomarkers e.g., genes and/or proteins
  • non-human animal models can be predictive of biomarkers relevant to treatment of human subjects (e.g., according to methods described herein).
  • a corresponding human analog of a biomarker e.g., genes and/or proteins identified using a non-human animal model can be determined; in some embodiments, such corresponding human analogs are useful in the treatment of human subjects as described herein.
  • a rodent (e.g., rat or mouse) model is used to identify biomarkers expected to be relevant to treatment of human subjects (e.g., according to methods described herein).
  • Non-limiting examples of human analogs that correspond with certain rat biomarkers described herein are provided in Table 1 A.
  • a biomarker described herein is selected from the human analogs in Table 1 A.
  • a human analog described herein is the corresponding human analog listed in Table 1 A.
  • biomarkers e.g., genes and/or proteins
  • a disease model for fibrosis of one organ may be predictive of biomarkers relevant to treatment of fibrotic diseases of other organs.
  • biomarkers identified using a model of a kidney fibrotic disease may be predictive of biomarkers relevant to treatment of fibrotic diseases of, for example, the lung, liver, and/or skin.
  • one or more biomarkers are differentially present in a sample taken from a subject of one status as compared with a subject of another status (e.g., more responsive to Compound 1 therapy vs less responsive to Compound 1 therapy). In some embodiments, one or more biomarkers are differentially present in a sample taken from the same subject at two or more different time points, i.e., when the status of the subject has changed from one time point to another.
  • detection of levels of one or more biomarkers are used to select and/or characterize patients who may be responsive to Compound 1 therapy.
  • levels of one or more biomarkers in a sample obtained from a subject are compared to a threshold level.
  • a biomarker is considered altered if the level is altered relative to a threshold level (e.g., altered by at least about 1.5-fold, about 2-fold, about 3-fold, about 4-fold, about 5-fold, about 10-fold, or about 20-fold, or more).
  • an altered biomarker is elevated relative to a threshold level (e.g., elevated by at least about 1.5-fold, about 2-fold, about 3-fold, about 4-fold, about 5-fold, about 10-fold, or about 20-fold, or more). In some embodiments, an altered biomarker is reduced relative to a threshold level (e.g., reduced by at least about 1.5-fold, about 2-fold, about 3-fold, about 4-fold, about 5-fold, about 10-fold, or about 20-fold, or more).
  • a threshold level e.g., elevated by at least about 1.5-fold, about 2-fold, about 3-fold, about 4-fold, about 5-fold, about 10-fold, or about 20-fold, or more.
  • a biomarker is considered altered if the level is altered relative to a threshold level (e.g., altered by at least 0.5, about 1.0, about 1.5, about 2.0, about 2.5, about 3.0, about 3.5, about 4.0, about 4.5, or about 5.0, or more standard deviations).
  • a threshold level e.g., altered by at least 0.5, about 1.0, about 1.5, about 2.0, about 2.5, about 3.0, about 3.5, about 4.0, about 4.5, or about 5.0, or more standard deviations.
  • an altered biomarker is reduced relative to a threshold level (e.g., reduced by at least about 0.5, about 1.0, about 1.5, about 2.0, about 2.5, about 3.0, about 3.5, about 4.0, about 4.5, or about 5.0, or more standard deviations).
  • a threshold level is determined from a population of healthy volunteers (e.g., a mean or median level from a population of healthy volunteers).
  • a method includes an in vitro method for determining a level of a biomarker.
  • in vitro methods for determining a level of a biomarker include, but are not limited to, a chemiluminescence assay, enzymatic assay, enzyme immunoassay, multiplex immunoassay, ELISA, chromatographic immunoassay, electrophoresis assay, radioimmunoassay, colorimetric assay, chromatography/mass spectrometry (e.g., GC/MS, LC/MS, LC/MS/MS, etc.), High Performance Liquid Chromatography (“HPLC”), and/or PCR (e.g., real-time PCR).
  • HPLC High Performance Liquid Chromatography
  • a method for detecting a level of a biomarker includes chromatographic and/or MS methods.
  • Exemplary methods include, but are not limited to, gas chromatography (GC), liquid chromatography/mass spectroscopy (LC-MS), gas chromatography/mass spectroscopy (GC-MS), nuclear magnetic resonance (NMR), magnetic resonance imaging (MRI), Fourier Transform InfraRed (FT-IR), and inductively coupled plasma mass spectrometry (ICP-MS).
  • a level of a biomarker corresponds to a level of gene expression (e.g., RNA, e.g., mRNA) and is quantified using methods known in the art.
  • a method of determining a level of expression of a biomarker gene can be or include a chemiluminescence assay, UV spectroscopy, hybridization assay (e.g., Fluorescent in Situ Hybridization (FISH), e.g., RNA-FISH), enzymatic assay, enzyme immunoassay (e.g., ELISA), multiplex assay, electrophoresis assay, radioassay, colorimetric assay, chromatography/mass spectrometry (e.g., GC/MS, LC/MS, LC/MS/MS, etc.), High Performance Liquid Chromatography (“HPLC”), and/or PCR
  • a level of a biomarker corresponds to a level of protein, including any fragment or degradation product thereof, and is quantified using methods known in the art.
  • a method of determining a level of expression of a biomarker protein can be or include a chemiluminescence assay, enzymatic assay, enzyme immunoassay, multiplex immunoassay, ELISA, chromatographic immunoassay, electrophoresis assay, radioimmunoassay, colorimetric assay, UV spectroscopy, chromatography/mass spectrometry (e.g., GC/MS, LC/MS, LC/MS/MS, etc.), or High Performance Liquid Chromatography (“HPLC”).
  • HPLC High Performance Liquid Chromatography
  • a biomarker useful in methods provided herein is a biomarker selected from Table 1 (e.g., gene expression of and/or level of protein produced by a gene selected from Table 1), or a human analog thereof (e.g., a human analog selected from Table 1 A).
  • a level of a biomarker selected from Table l is a level of gene expression of a gene selected from Table 1 or a human analog thereof (e.g., a human analog selected from Table 1 A). Table 1.
  • patients are selected and/or characterized based on an elevated level of one or more biomarkers selected from Table 2 or a human analog thereof (e.g., a human analog selected from Table 1 A) and/or a reduced level of one or more biomarkers selected from Table 3 or a human analog thereof (e.g., a human analog selected from Table 1 A).
  • Table 2 a human analog thereof
  • Table 3 a human analog thereof
  • a biomarker useful in methods provided herein is selected from Table 5 or a human analog thereof (e.g., a human analog selected from Table 1 A) (see Example 2 below).
  • a biomarker useful in methods provided herein is a biomarker selected from Table 5, or a human analog thereof, with a change in mean expression for sham animals relative to PANX animals of at least about 1.5-fold, about 2-fold, about 3-fold, about 4-fold, about 5-fold, about 10-fold, or about 20-fold.
  • a biomarker useful in methods provided herein is a biomarker selected from Table 5, or a human analog thereof, with a change in mean expression for sham animals relative to PANX animals of at least about 0.5, about 1.0, about 1.5, about 2.0, about 2.5, about 3.0, about 3.5, about 4.0, about 4.5, or about 5.0 standard deviations.
  • a biomarker useful in methods provided herein is a biomarker selected from Table 5, or a human analog thereof, with a change in mean expression for PANX animals relative to PANX+Compound 1 animals of at least about 1.5-fold, about 2-fold, about 3-fold, about 4-fold, about 5-fold, about 10-fold, or about 20-fold.
  • a biomarker useful in methods provided herein is a biomarker selected from Table 5, or a human analog thereof, with a change in mean expression for PANX animals relative to PANX+Compound 1 animals of at least about 0.5, about 1.0, about 1.5, about 2.0, about 2.5, about 3.0, about 3.5, about 4.0, about 4.5, or about 5.0 standard deviations.
  • a biomarker useful in methods provided herein is a biomarker selected from Table 5, or a human analog thereof, with a change in mean expression for sham animals relative to PANX+Compound 1 animals of less than about 100%, about 90%, about 80%, about 70%, about 60%, about 50%, about 40%, about 30%, about 20%, or about 10%.
  • a biomarker useful in methods provided herein is a biomarker selected from Table 5, or a human analog thereof, with a change in mean expression for sham animals relative to PANX+Compound 1 animals of less than about 0.5, about 1.0, about 1.5, about 2.0, about 2.5, about 3.0, about 3.5, about 4.0, about 4.5, or about 5.0 standard deviations.
  • a biomarker useful in methods provided herein is a biomarker selected from Table 5, or a human analog thereof, with: a change in mean expression for sham animals relative to PANX animals of at least about 1.5- fold, about 2-fold, about 3-fold, about 4-fold, about 5-fold, about 10-fold, or about 20-fold; and/or a change in mean expression for PANX animals relative to PANX+Compound 1 animals of at least about 1.5-fold, about 2-fold, about 3-fold, about 4-fold, about 5-fold, about 10-fold, or about 20-fold; and/or a change in mean expression for sham animals relative to PANX+Compound 1 animals of less than about 100%, about 90%, about 80%, about 70%, about 60%, about 50%, about 40%, about 30%, about 20%, or about 10%.
  • a biomarker useful in methods provided herein is a biomarker selected from Table 5, or a human analog thereof, with: a change in mean expression for sham animals relative to PANX animals of at least about 0.5, about 1.0, about 1.5, about 2.0, about 2.5, about 3.0, about 3.5, about 4.0, about 4.5, or about 5.0 standard deviations; and/or a change in mean expression for PANX animals relative to PANX+Compound 1 animals of at least about 0.5, about 1.0, about 1.5, about 2.0, about 2.5, about 3.0, about 3.5, about 4.0, about 4.5, or about 5.0 standard deviations; and/or a change in mean expression for sham animals relative to PANX+Compound 1 animals of less than about 0.5, about 1.0, about 1.5, about 2.0, about 2.5, about 3.0, about 3.5, about 4.0, about 4.5, or about 5.0 standard deviations.
  • the present disclosure provides insights that altered levels of one or more biomarkers selected from Table 1 and/or Table 2 and/or Table 3 and/or Table 5, or a human analog thereof, may be useful in selecting and/or characterizing patients for Compound 1 therapy.
  • patients are selected and/or characterized based on the percentage of biomarkers with altered levels observed in a biological sample obtained from the patient. For example, in some embodiments, a patient has been determined to have an altered level of at least about 10%, about 20%, about 30%, about 40%, about 50%, about 60%, about 70%, about 80%, or about 90% of the biomarkers in Table 1, or a human analog thereof. In some embodiments, a patient has been determined to have an elevated level of at least about 10%, about 20%, about 30%, about 40%, about 50%, about 60%, about 70%, about 80%, or about 90% of the biomarkers in Table 2, or a human analog thereof.
  • a patient has been determined to have a reduced level of at least about 10%, about 20%, about 30%, about 40%, about 50%, about 60%, about 70%, about 80%, or about 90% of the biomarkers in Table 3, or a human analog thereof. In some embodiments, a patient has been determined to have an elevated level of at least one biomarker in Table 2, or a human analog thereof, and a reduced level of at least one biomarker in Table 3, or a human analog thereof.
  • a patient has been determined to have an elevated level of at least about 10%, about 20%, about 30%, about 40%, about 50%, about 60%, about 70%, about 80%, or about 90% of the biomarkers in Table 2, or a human analog thereof, and a reduced level of at least about 10%, about 20%, about 30%, about 40%, about 50%, about 60%, about 70%, about 80%, or about 90% of the biomarkers in Table 3, or a human analog thereof.
  • a patient has been determined to have an altered level of at least about 10%, about 20%, about 30%, about 40%, about 50%, about 60%, about 70%, about 80%, or about 90% of the biomarkers in Table 5 or a human analog thereof. In some embodiments, a patient has been determined to have an altered level of at least about 10%, about 20%, about 30%, about 40%, about 50%, about 60%, about 70%, about 80%, or about 90% of the biomarkers in Table 5, or a human analog thereof, with a change in mean expression for sham animals relative to PANX animals of at least about 1.5-fold, about 2-fold, about 3-fold, about 4-fold, about 5-fold, about 10-fold, or about 20-fold.
  • a patient has been determined to have an altered level of at least about 10%, about 20%, about 30%, about 40%, about 50%, about 60%, about 70%, about 80%, or about 90% of the biomarkers in Table 5, or a human analog thereof, with a change in mean expression for sham animals relative to PANX animals of at least about 0.5, about 1.0, about 1.5, about 2.0, about 2.5, about 3.0, about 3.5, about 4.0, about 4.5, or about 5.0 standard deviations.
  • a patient has been determined to have an altered level of at least about 10%, about 20%, about 30%, about 40%, about 50%, about 60%, about 70%, about 80%, or about 90% of the biomarkers in Table 5, or a human analog thereof, with a change in mean expression for PANX animals relative to PANX+Compound 1 animals of at least about 1.5-fold, about 2-fold, about 3-fold, about 4-fold, about 5-fold, about 10-fold, or about 20-fold.
  • a patient has been determined to have an altered level of at least about 10%, about 20%, about 30%, about 40%, about 50%, about 60%, about 70%, about 80%, or about 90% of the biomarkers in Table 5, or a human analog thereof, with a change in mean expression for PANX animals relative to PANX+Compound 1 animals of at least about 0.5, about 1.0, about 1.5, about 2.0, about 2.5, about 3.0, about 3.5, about 4.0, about 4.5, or about 5.0 standard deviations.
  • a patient has been determined to have an altered level of at least about 10%, about 20%, about 30%, about 40%, about 50%, about 60%, about 70%, about 80%, or about 90% of the biomarkers in Table 5, or a human analog thereof, with a change in mean expression for sham animals relative to PANX+Compound 1 animals of less than about 100%, about 90%, about 80%, about 70%, about 60%, about 50%, about 40%, about 30%, about 20%, or about 10%.
  • a patient has been determined to have an altered level of at least about 10%, about 20%, about 30%, about 40%, about 50%, about 60%, about 70%, about 80%, or about 90% of the biomarkers in Table 5, or a human analog thereof, with a change in mean expression for sham animals relative to PANX+Compound 1 animals of less than about 0.5, about 1.0, about 1.5, about 2.0, about 2.5, about 3.0, about 3.5, about 4.0, about 4.5, or about 5.0 standard deviations.
  • a patient has been determined to have an altered level of at least about 10%, about 20%, about 30%, about 40%, about 50%, about 60%, about 70%, about 80%, or about 90% of the biomarkers in Table 5, or a human analog thereof, with: a change in mean expression for sham animals relative to PANX animals of at least about 1.5- fold, about 2-fold, about 3-fold, about 4-fold, about 5-fold, about 10-fold, or about 20-fold; and/or a change in mean expression for PANX animals relative to PANX+Compound 1 animals of at least about 1.5-fold, about 2-fold, about 3-fold, about 4-fold, about 5-fold, about 10-fold, or about 20-fold; and/or a change in mean expression for sham animals relative to PANX+Compound 1 animals of less than about 100%, about 90%, about 80%, about 70%, about 60%, about 50%, about 40%, about 30%, about 20%, or about 10%.
  • a patient has been determined to have an altered level of at least about 10%, about 20%, about 30%, about 40%, about 50%, about 60%, about 70%, about 80%, or about 90% of the biomarkers in Table 5, or a human analog thereof, with: a change in mean expression for sham animals relative to PANX animals of at least about 0.5, about 1.0, about 1.5, about 2.0, about 2.5, about 3.0, about 3.5, about 4.0, about 4.5, or about 5.0 standard deviations; and/or a change in mean expression for PANX animals relative to PANX+Compound 1 animals of at least about 0.5, about 1.0, about 1.5, about 2.0, about 2.5, about 3.0, about 3.5, about 4.0, about 4.5, or about 5.0 standard deviations; and/or a change in mean expression for sham animals relative to PANX+Compound 1 animals of less than about 0.5, about 1.0, about 1.5, about 2.0, about 2.5, about 3.0, about 3.5, about 4.0, about
  • a patient has been determined to have an altered level of at least one biomarker selected from Table 5, or a human analog thereof, with a change in mean expression for sham animals relative to PANX animals of at least about 1.5-fold, about 2-fold, or about 3-fold and at least one biomarker selected from Table 5, or a human analog thereof, with a change in mean expression for sham animals relative to PANX animals of at least about 4-fold, about 5-fold, about 10-fold, or about 20-fold.
  • a patient has been determined to have an altered level of at least one biomarker selected from Table 5, or a human analog thereof, with a change in mean expression for sham animals relative to PANX animals of at least about 0.5, about 1.0, about 1.5, about 2.0, or about 2.5 standard deviations and at least one biomarker selected from Table 5, or a human analog thereof, with a change in mean expression for sham animals relative to PANX animals of at least about 3.0, about 3.5, about 4.0, about 4.5, or about 5.0 standard deviations.
  • a patient has been determined to have an altered level of at least one biomarker selected from Table 5, or a human analog thereof, with a change in mean expression for PANX animals relative to PANX+Compound 1 animals of at least about 1.5-fold, about 2-fold, or about 3-fold and at least one biomarker in Table 5, or a human analog thereof, with a change in mean expression for PANX animals relative to PANX+Compound 1 animals of at least about 4-fold, about 5-fold, about 10-fold, or about 20-fold.
  • a patient has been determined to have an altered level of at least one biomarker selected from Table 5, or a human analog thereof, with a change in mean expression for PANX animals relative to PANX+Compound 1 animals of at least about 0.5, about 1.0, about 1.5, about 2.0, or about 2.5 standard deviations and at least one biomarker selected from Table 5, or a human analog thereof, with a change in mean expression for PANX animals relative to PANX+Compound 1 animals of at least about 3.0, about 3.5, about 4.0, about 4.5, or about 5.0 standard deviations.
  • a patient has been determined to have an altered level of at least one biomarker selected from Table 5, or a human analog thereof, with a change in mean expression for sham animals relative to PANX+Compound 1 animals of less than about 100%, about 90%, about 80%, about 70%, or about 60% and at least one biomarker selected from Table 5, or a human analog thereof, with a change in mean expression for sham animals relative to PANX+Compound 1 animals of less than about 50%, about 40%, about 30%, about 20%, or about 10%.
  • a patient has been determined to have an altered level of at least one biomarker selected from Table 5, or a human analog thereof, a change in mean expression for sham animals relative to PANX+Compound 1 animals of less than about 0.5, about 1.0, about 1.5, about 2.0, or about 2.5 standard deviations and at least one biomarker selected from Table 5, or a human analog thereof, with a change in mean expression for sham animals relative to PANX+Compound 1 animals of less than about 3.0, about 3.5, about 4.0, about 4.5, or about 5.0 standard deviations.
  • a “change in mean expression” between, e.g., sham animals relative to PANX animals or PANX animals relative to PANX+Compound 1 animals or sham animals relative to PANX+Compound 1 animals, in Table 5 refers to a comparison of a mean expression value in Table 5 with another mean expression value in Table 5.
  • a biomarker with a change in mean expression for sham animals relative to PANX animals of at least about 2-fold refers to a biomarker with mean expression for PANX animals that is at least about 2-fold higher or lower than mean expression for sham animals.
  • a biomarker with a change in mean expression for sham animals relative to PANX+Compound 1 animals of less than about 50% refers to a biomarker with a mean expression for PANX+Compound 1 animals that is less than 50% higher or lower than (i.e., within 50% of) mean expression for sham animals.
  • a patient is determined to have an altered level of at least two biomarkers (e.g., those selected from Table 1 and/or Table 2 and/or Table 3 and/or Table 5) which are part of the same biochemical pathway.
  • biochemical pathways common to at least two biomarkers listed in Table 1 include: glycosphingolipid metabolic process, cytokine biosynthetic process, lysosome organization, regulation of transcription, protein localization, negative regulation of cell migration, JAK/STAT cascade, regulation of protein secretion, keratinocyte differentiation, embryonic morphogenesis, cell development, actin filament polymerization, positive regulation of lymphocyte activation, organ development, intrinsic apoptotic signaling, carboxylic acid metabolic process, growth, chromosome segregation, catabolic process, hemostasis, regulation of cytoskeleton organization, mitotic spindle organization, regulated secretory pathway, regulation of synapse structure and activity, viral infectious cycle, positive regulation of sequence specific DNA binding
  • the present disclosure encompasses the recognition that levels of one or more urinary and/or circulating biomarkers may be indicative of and/or correlated with levels of one or more biomarkers described herein, e.g., those selected from Table 1 and/or Table 2 and/or Table 3 and/or Table 5.
  • such urinary and/or circulating biomarkers may be used in provided methods, e.g., to select and/or characterize patients for Compound 1 therapy.
  • the present disclosure encompasses the recognition that a collagen 6 biomarker is useful in methods provided herein.
  • Collagen 6 is a member of the collagen family, primarily present in the extracellular matrix and reported to perform both mechanical and cytoprotective roles. See Cescon, M., et al., J. Cell Sci. (2015) 128 (19):3525- 3531.
  • a collagen 6 biomarker can be a level of gene expression (e.g., RNA, e.g., mRNA) or a level of protein expression (e.g., a level of collagen 6 protein and/or a corresponding procollagen polypeptide, collagen protein, preprocollagen polypeptide, amino-terminal procollagen polypeptide, collagen fibril, collagen fiber, and/or any fragment or degradation product of any thereof).
  • a collagen 6 biomarker can be a level of collagen 6, pro-collagen 6, and/or endotrophin, as well as fragments and/or degradation products thereof.
  • the pro-peptide of the alpha-3 chain of collagen 6 has a sequence overlap with endotrophin (see Rasmussen, D. G. K., et al., Scientific Reports, 7, 2017, 17328); as such, in some embodiments, a collagen 6 biomarker may be or comprise an endotrophin level.
  • the present disclosure encompasses the recognition that a collagen 1 and/or collagen 3 biomarker is useful in methods provided herein, e.g., in combination with one or more other biomarkers described herein.
  • a collagen 1 biomarker can include a level of gene expression (e.g., COL1 Al gene expression) or a level of protein expression (e.g., a level of collagen 1 protein and/or a corresponding procollagen polypeptide, collagen protein, preprocollagen polypeptide, amino-terminal procollagen polypeptide, collagen fibril, collagen fiber, and/or any fragment or degradation product of any thereof).
  • a collagen 3 biomarker can include a level of gene expression (e.g., COL3A1 gene expression) or a level of protein expression (e.g., a level of collagen 3 protein and/or a corresponding procollagen polypeptide, collagen protein, preprocollagen polypeptide, amino-terminal procollagen polypeptide, collagen fibril, collagen fiber, and/or any fragment or degradation product of any thereof).
  • a level of gene expression e.g., COL3A1 gene expression
  • a level of protein expression e.g., a level of collagen 3 protein and/or a corresponding procollagen polypeptide, collagen protein, preprocollagen polypeptide, amino-terminal procollagen polypeptide, collagen fibril, collagen fiber, and/or any fragment or degradation product of any thereof.
  • the present disclosure encompasses the recognition that a combination of a collagen 6 biomarker with a collagen 1 and/or collagen 3 biomarker may be useful in selecting and/or characterizing patients for Compound 1 therapy.
  • a combination of a collagen 6 biomarker with one or more biomarkers selected from Table 1, Table 2, Table 3, and/or Table 5 may be useful in selecting and/or characterizing patients for Compound 1 therapy.
  • a patient is determined to have an altered level of a biomarker when the level of the biomarker is above or below a threshold level (e.g., a predetermined median or mean level).
  • a patient is determined to have an altered level of a biomarker when the level of the biomarker is different from a threshold level by at least about 1.5-fold, about 2-fold, about 3-fold, about 4-fold, about 5-fold, about 10-fold, or about 20-fold, or more.
  • a patient is determined to have an altered level of a biomarker when the level of the biomarker is different from a threshold level by at least about 0.5, about 1.0, about 1.5, about 2.0, about 2.5, about 3.0, about 3.5, about 4.0, about 4.5, or about 5.0, or more standard deviations.
  • a patient is determined to have an elevated level of a biomarker when the level of the biomarker is above a threshold level (e.g., a predetermined median or mean level).
  • a patient is determined to have an elevated level of a biomarker when the level of the biomarker is above a threshold level by at least about 1.5- fold, about 2-fold, about 3-fold, about 4-fold, about 5-fold, about 10-fold, or about 20-fold, or more. In some embodiments, a patient is determined to have an elevated level of a biomarker when the level of the biomarker is above a threshold level by at least about 0.5, about 1.0, about 1.5, about 2.0, about 2.5, about 3.0, about 3.5, about 4.0, about 4.5, or about 5.0, or more standard deviations.
  • a patient is determined to have a reduced level of a biomarker when the level of the biomarker is below a threshold level (e.g., a predetermined median or mean level). In some embodiments, a patient is determined to have a reduced level of a biomarker when the level of the biomarker is below a threshold level by at least about 1.5-fold, about 2- fold, about 3-fold, about 4-fold, about 5-fold, about 10-fold, or about 20-fold, or more.
  • a threshold level e.g., a predetermined median or mean level.
  • a patient is determined to have a reduced level of a biomarker when the level of the biomarker is below a threshold level by at least about 0.5, about 1.0, about 1.5, about 2.0, about 2.5, about 3.0, about 3.5, about 4.0, about 4.5, or about 5.0, or more standard deviations.
  • the present disclosure provides methods of identifying biomarkers useful for selecting, identifying, and/or characterizing patients likely to benefit from a treatment with Compound 1.
  • biomarkers are identified based on a mean change across a population of subjects administered Compound 1 relative to a comparable reference population (e.g., as described in Example 2 or 17).
  • biomarkers useful in methods provided herein are biomarkers that have been established to have a mean increase or decrease of at least about 1.5-fold, about 2-fold, about 3-fold, about 4-fold, about 5-fold, about 10-fold, or about 20-fold, or more in a population of subjects administered Compound 1 relative to a comparable reference population.
  • biomarkers useful in methods provided herein are biomarkers that have been established to have a mean increase or decrease of at least about 0.5, about 1.0, about 1.5, about 2.0, about 2.5, about 3.0, about 3.5, about 4.0, about 4.5, or about 5.0, or more standard deviations in a population of subjects administered Compound 1 relative to a comparable reference population.
  • a population of subjects is a population of human subjects. In some embodiments, a population of subjects is a population of non-human animal subjects (e.g., rodent subjects). In some embodiments, a reference population has not received Compound 1. In some embodiments, a reference population has received an otherwise comparable composition that does not provide Compound 1 (e.g., a placebo).
  • biomarkers are identified based on a mean change across a population of subjects with confirmed fibrotic disease (e.g., confirmed fibrotic disease of the kidney) relative to a population of healthy volunteers (e.g., as described in Example 2 or 17).
  • biomarkers useful in methods provided herein are biomarkers that have been established to have a mean increase or decrease of at least about 1.5-fold, about 2-fold, about 3-fold, about 4-fold, about 5-fold, about 10-fold, or about 20- fold, or more in a population of subjects with confirmed fibrotic disease (e.g., confirmed fibrotic disease of the kidney) relative to a population of healthy volunteers.
  • biomarkers useful in methods provided herein are biomarkers that have been established to have a mean increase or decrease of at least about 0.5, about 1.0, about 1.5, about 2.0, about 2.5, about 3.0, about 3.5, about 4.0, about 4.5, or about 5.0, or more standard deviations in a population of subjects with confirmed fibrotic disease (e.g., confirmed fibrotic disease of the kidney) relative to a population of healthy volunteers.
  • a population of subjects is a population of human subjects.
  • a population of subjects is a population of non-human animal subjects (e.g., rodent subjects).
  • compositions that comprises and/or delivers Compound 1 as described herein are provided herein.
  • such administering is achieved by administering a composition that delivers Compound 1 (e.g., in some embodiments, a composition that is or comprises Compound 1, or a composition that otherwise delivers Compound 1 - e.g., that is or comprises a prodrug of Compound 1, a complex or other entity that releases Compound 1 upon administration, etc.).
  • a composition that delivers Compound 1 e.g., in some embodiments, a composition that is or comprises Compound 1, or a composition that otherwise delivers Compound 1 - e.g., that is or comprises a prodrug of Compound 1, a complex or other entity that releases Compound 1 upon administration, etc.
  • provided methods relate to treatment of fibrotic disease(s) (e.g., fibrotic diseases of the kidney) and selecting, identifying, and/or characterizing patients likely to benefit from a treatment with Compound 1.
  • the present disclosure is based in part on the recognition that certain biomarkers can distinguish patients who are likely to respond to therapy, for example because the drivers of their fibrotic disease correspond with the mechanism of action of Compound 1.
  • drivers of their kidney disease correspond with the mechanism of action of Compound 1.
  • a patient to be treated with a method of the present disclosure has an altered level of one or more gene products or proteins that are part of the mechanism of action of Compound 1.
  • the present disclosure provides a method of treating a patient diagnosed with, suspected of having, or at risk of a fibrotic disease (e.g., a fibrotic disease described herein, such as a fibrotic disease of the kidney), comprising administering an effective amount of Compound 1 to a patient that has been determined to have an altered level of one or more biomarkers described herein.
  • a fibrotic disease e.g., a fibrotic disease described herein, such as a fibrotic disease of the kidney
  • the present disclosure provides a method of treating a fibrotic disease (e.g., a fibrotic disease described herein, such as a fibrotic disease of the kidney) in a patient characterized by an altered level of one or more biomarkers described herein, comprising administering an effective amount of Compound 1 to the patient.
  • a fibrotic disease e.g., a fibrotic disease described herein, such as a fibrotic disease of the kidney
  • the present disclosure provides a method comprising administering an effective amount of Compound 1 to a patient that has been determined to have (i) at least one symptom selected from proteinuria and/or hypoalbuminemia and/or hyperlipidemia and/or edema; and (ii) an altered level of one or more biomarkers described herein.
  • the present disclosure provides a method comprising administering an effective amount of Compound 1 to a patient that has been determined to have (i) nephrotic syndrome; and (ii) an altered level of one or more biomarkers described herein.
  • the present disclosure provides a method comprising administering an effective amount of Compound 1 to a patient in need thereof, wherein the patient has been determined to have an altered level of one or more biomarkers described herein.
  • one or more biomarkers are selected from Table 1 and/or Table 2 and/or Table 3 and/or Table 5, or a human analog thereof (e.g., a human analog selected from Table 1 A), including any classes and subclasses thereof as described herein, both singly and in combination.
  • one or more biomarkers comprise a collagen 6 biomarker.
  • one or more biomarkers comprise a collagen 1 biomarker or a collagen 3 biomarker.
  • one or more biomarkers are selected from a collagen 6 biomarker, a collagen 1 biomarker, a collagen 3 biomarker, and a biomarker in Table 1 and/or Table 2 and/or Table 3 and/or Table 5, or a human analog thereof (e.g., a human analog selected from Table 1 A), including any classes and subclasses thereof as described herein, both singly and in combination.
  • a human analog thereof e.g., a human analog selected from Table 1 A
  • a patient has been determined to have an altered level of at least about 10%, about 20%, about 30%, about 40%, about 50%, about 60%, about 70%, about 80%, or about 90% of the biomarkers in Table 1 and/or Table 2 and/or Table 3 and/or Table 5, or a human analog thereof (e.g., a human analog selected from Table 1 A), including any classes and subclasses thereof as described herein, both singly and in combination.
  • a patient has been determined to have an altered level of a collagen 6 biomarker.
  • a patient has been determined to have an altered level of a collagen 1 and/or collagen 3 biomarker.
  • an altered level of a biomarker is a level that is different from (e.g., at least about 1.5-fold, about 2-fold, about 3-fold, about 4-fold, about 5-fold, about 10-fold, or about 20-fold, or more, above or below) a corresponding threshold level.
  • one or more biomarkers are selected from biomarkers identified using a method described herein. In some embodiments, one or more biomarkers are selected from biomarkers whose levels have been established to have a mean increase or decrease of at least about 1.5-fold, about 2-fold, about 3-fold, about 4-fold, about 5-fold, about 10-fold, or about 20-fold, or more in a population of subjects administered Compound 1 relative to a comparable reference population.
  • one or more biomarkers are selected from biomarkers whose levels have been established to have a mean increase or decrease of at least about 1.5-fold, about 2-fold, about 3-fold, about 4-fold, about 5-fold, about 10-fold, or about 20-fold, or more in a population of subjects with confirmed fibrotic disease (e.g., fibrotic disease of the kidney) relative to a population of healthy volunteers.
  • fibrotic disease e.g., fibrotic disease of the kidney
  • the present disclosure provides a method of treating a patient diagnosed with, suspected of having, or at risk for a fibrotic disease (e.g., a fibrotic disease described herein, such as a fibrotic disease of the kidney), comprising (i) obtaining or determining a level of one or more biomarkers described herein in a biological sample obtained from the patient; and (ii) comparing the determined level(s) to a corresponding threshold level.
  • a method further comprises performing an assay on a biological sample obtained from the patient to determine level(s) of one or more biomarkers.
  • Compound 1 is administered to the patient.
  • Compound 1 is not administered to the subject.
  • the present disclosure provides methods of administering Compound 1 to a subject or population of subjects described herein, according to a regimen established to achieve one or more desirable outcomes.
  • the fibrotic disease is stabilized (i.e., does not worsen) and/or is ameliorated (i.e., one or more symptoms improve) in a patient treated with Compound 1.
  • treatment of a patient with Compound 1 increases or decreases a level of one or more biomarkers (i.e., such that the level of the one or more biomarkers is less different from a threshold level than prior to treatment with Compound 1).
  • treatment of a patient with Compound 1 decreases a level of one or more biomarkers that was elevated prior to treatment with Compound 1 (e.g., one or more biomarkers selected from Table 2 or a human analog thereof, and/or a collagen 6, collagen 1, or collagen 3 biomarker). In some embodiments, treatment of a patient with Compound 1 increases a level of one or more biomarkers that was reduced prior to treatment with Compound 1 (e.g., one or more biomarkers selected from Table 3 or a human analog thereof).
  • a regimen has been established to achieve one or more desirable outcomes, relative to that observed for a comparable reference population that has not received Compound 1 (e.g., that has received a placebo).
  • a placebo as used herein is a dosage form that matches that of an active study compound, but does not deliver the active study compound (e.g., Compound 1).
  • a placebo can be a capsule that visually matches an active study drug and is composed of the same capsule shell but is filled with the pharmaceutical excipient (and lacking the active study drug), e.g., silicified microcrystalline cellulose.
  • a reference composition may be or may have been administered at the same intervals and/or in the same amounts as a composition providing Compound 1.
  • provided methods are useful for monitoring subjects (e.g., monitoring status of subjects over time and/or monitoring therapy).
  • the present disclosure provides methods comprising (i) administering an effective amount of Compound 1 to a patient characterized by an altered level of one or more biomarkers described herein; and (ii) monitoring levels of the one or more biomarkers, e.g., over a period of time.
  • therapy with Compound 1 is discontinued.
  • the dose and/or dosing frequency of Compound 1 is increased.
  • the present disclosure provides a method comprising determining levels of one or more biomarkers described herein in each of a plurality of biological samples obtained at different time points from a single patient; and comparing the determined levels from a first time point with those from at least one later time point.
  • the present disclosure provides a method comprising determining levels of one or more biomarkers described herein from a biological sample obtained from a subject for whom levels of the one or more biomarkers have previously been obtained at least once; and comparing the determined levels with the previously obtained levels.
  • a first time point and one or more later time points are separated from one another by a reasonably consistent interval.
  • such methods further comprise administering Compound 1 therapy to a subject (e.g., a subject who has been determined to have moved from a non- responsive to a responsive state).
  • a meaningful change in a determined level over time indicates a change in the subject’s status.
  • a meaningful change in a determined level over time is a change (e.g., an increase or a decrease) of at least about 1.5-fold, about 2- fold, about 3-fold, about 4-fold, about 5-fold, about 10-fold, or about 20-fold, or more compared to a threshold level.
  • a meaningful change in a determined level over time is a change of more than about 0.5, about 1.0, about 1.5, or about 2.0, or more standard deviations away from a threshold level.
  • provided methods are useful for monitoring therapy (e.g., efficacy and/or other indicators of response).
  • a sample from a first time point is or was obtained from a subject prior to administration of Compound 1
  • a sample from a second time point is or was obtained from the subject after administration of Compound 1.
  • the levels of one or more biomarkers are no longer altered and/or are altered to a lesser degree in a later sample compared to a first sample, then Compound 1 therapy is continued.
  • the present disclosure provides a method for treatment with Compound 1 that includes: (i) receiving a report listing the level of one or more biomarkers (e.g., one or more biomarkers described herein) for a patient with a fibrotic disease (e.g., a fibrotic kidney disease) and/or a symptom of a fibrotic disease (e.g., proteinuria); (ii) receiving a request for reimbursement of the screening and/or of a particular therapeutic regimen; and (iii) approving payment and/or reimbursement for treatment with Compound 1 therapy if the report indicates the level of one or more biomarkers is above a threshold level.
  • a fibrotic disease e.g., a fibrotic kidney disease
  • a symptom of a fibrotic disease e.g., proteinuria
  • the present disclosure provides methods of treating diseases, disorders, and conditions (e.g., according to methods provided herein). In some embodiments, provided methods are useful for reducing fibrosis in a subject in need thereof. In some embodiments, provided methods are useful for treating a disease, disorder, or condition characterized by or otherwise associated with fibrosis.
  • the present disclosure encompasses the recognition that treating fibrosis (e.g., using provided methods) instead of the underlying etiology may allow for broadly applicable antifibrotic therapies.
  • provided methods may be suitable for reducing fibrosis in a variety of tissues and/or organs; the present disclosure contemplates use of Compound 1 for treating diseases, disorders, and conditions characterized by or otherwise associated with fibrosis in any suitable tissue and/or organ.
  • provided methods are suitable for treating diseases, disorders and conditions that are or comprise fibrosis of gastrointestinal tract, heart, kidney, lung, liver, muscle, pancreas, and/or skin.
  • provided methods are suitable for treating diseases, disorders, and conditions characterized by or otherwise associated with cysts (e.g., in the kidney, liver, pancreas, ovaries, spermatic duct, etc.).
  • fibrosis is the sole or a predominant component, as well as those in which fibrosis is a secondary component (e.g., a symptom and/or result of an underlying disease, disorder, or condition). It will also be appreciated that there are a variety of sources or causes of fibrosis.
  • certain injuries can progress to development of fibrosis.
  • provided methods are useful for treating acute injuries (e.g., acute organ injuries, such as acute lung injury, acute liver injury, or acute kidney injury), as well as for treating chronic injuries (e.g., chronic organ injuries, such as chronic lung injury, chronic liver injury, or chronic kidney injury).
  • provided methods are useful for treating fibrosis associated with an acute injury, such as that incurred from trauma and/or surgery and/or infection (e.g., a viral infection).
  • provided methods are useful for treating damaged and/or ischemic organs, transplants, or grafts, as well as ischemia/reperfusion injury or post-surgical scarring.
  • provided methods are useful for treating pulmonary diseases, disorders, and conditions. In some embodiments, provided methods are useful for treating pulmonary fibrosis. In some embodiments, provided methods are useful for treating pulmonary fibrosis secondary to, or otherwise associated with, an underlying indication. In some embodiments, provided methods are useful for treating interstitial lung diseases (e.g., fibrosing interstitial lung diseases). In some embodiments, provided methods are useful for treating pneumonias (e.g., idiopathic interstitial pneumonias). In some embodiments, provided methods are useful for treating idiopathic pulmonary fibrosis (IPF).
  • IPF idiopathic pulmonary fibrosis
  • provided methods are useful for treating pulmonary fibrosis associated with an infection (e.g., a bacterial, viral, or fungal infection).
  • an infection e.g., a bacterial, viral, or fungal infection.
  • provided methods are useful for treating pulmonary fibrosis associated with a viral infection (e.g., an influenza or coronavirus infection, such as COVID-19).
  • a fibrotic disease to be treated by methods of the present disclosure is pulmonary fibrosis.
  • Pulmonary fibrosis is a chronic, progressive and ultimately a fatal interstitial lung disease resulting from epithelial cell injury due to many factors.
  • activation of inflammatory cells and fibroblasts/myofibroblasts involves a cascade of cytokines/chemokines, growth factor network and deposit extracellular matrix, including collagen), which leads to pulmonary fibrosis and respiratory failure.
  • Pulmonary fibrosis causes high morbidity and mortality. At least five million people worldwide and -200,000 people in the United States suffer from pulmonary fibrosis. There is an unmet critical need for effective and affordable treatments for acute and chronic lung injuries.
  • Pulmonary fibrosis is associated with pronounced morbidity with high impact on economic burden.
  • the prevalence of pneumoconiosis (a disease caused by inhalation of dust and silica that causes inflammation and lung fibrosis) caused direct and indirect economic losses of around 28 billion yuan in China (4.3 billion US dollars) for 1 year.
  • a fibrotic disease to be treated by methods of the present disclosure is idiopathic pulmonary fibrosis.
  • Idiopathic pulmonary fibrosis is a chronic, irreversible, and progressive fibrotic disorder of the lower respiratory tract that typically affects adults over the age of 40.
  • Idiopathic interstitial pneumonias are diffuse parenchymal lung diseases, of which IPF is the most common and severe type of fibrotic lung disease. It is anatomically characterized by scarring of the lungs with a pattern of usual interstitial pneumonia (UIP) on high resolution computed tomography or histologic appearance on lung biopsy.
  • IPF Intra-pulmonary disease characterized by exertional dyspnea and cough.
  • Median survival following diagnosis of IPF ranges between 2 and 5 years, lower than that for many common cancers (Ley, B., et al. Am. J. Respir. Crit. Care Med. 2011;183:431-440; Seigel, R. L., et al. CA. Cancer J. Clin. 2016;66:7- 30).
  • Fibrotic process in IPF is progressive and, regardless of the nature of the initial injury, may follow a common pathway characterized by alveolar epithelial cell (AEC) dysfunction.
  • AEC alveolar epithelial cell
  • AECsl type I epithelial cells
  • AECs2 type II epithelial cells
  • TGF- ⁇ 1 transforming growth factor beta 1
  • PDGF platelet-derived growth factor
  • VEGF vascular endothelial growth factor
  • FGF fibroblast growth factor
  • the progression of fibrosis in IPF follows a common and complex path in which the AECs, fibroblasts, and endothelial cells produce an array of cytokines and growth factors that stimulate fibroblast proliferation and matrix synthesis.
  • provided methods are useful for treating hepatic diseases, disorders, or conditions. In some embodiments, provided methods are useful for treating hepatic fibrosis (e.g., fibrotic liver disease). In some embodiments, provided methods are useful for treating cirrhosis. In some embodiments, provided methods are useful for treating hepatic fibrosis and/or cirrhosis secondary to, or otherwise associated with, an underlying indication.
  • hepatic fibrosis e.g., fibrotic liver disease
  • provided methods are useful for treating cirrhosis.
  • provided methods are useful for treating hepatic fibrosis and/or cirrhosis secondary to, or otherwise associated with, an underlying indication.
  • provided methods are useful for treating hepatic fibrosis associated with hepatitis C, hepatitis B, delta hepatitis, chronic alcoholism, nonalcoholic steatohepatitis (NASH), extrahepatic obstructions (e.g., stones in bile duct), cholangiopathies (e.g., primary biliary cirrhosis or sclerosing cholangitis), autoimmune liver disease, or inherited metabolic disorders (e.g., Wilson’s disease, hemochromatosis, or alpha-1 antitrypsin deficiency).
  • NASH nonalcoholic steatohepatitis
  • a fibrotic disease to be treated by methods of the present disclosure is liver fibrosis.
  • Liver fibrosis is a scarring response of the liver to chronic liver injury; when fibrosis progresses to cirrhosis, morbid complications can develop.
  • endstage liver fibrosis or cirrhosis is the seventh leading cause of death in the United States, and afflicts hundreds of millions of people worldwide; deaths from end-stage liver disease in the United States are expected to increase, mainly due to the hepatitis C epidemic.
  • liver disease In addition to the hepatitis C virus, many other forms of chronic liver injury also lead to end-stage liver disease and cirrhosis, including other viruses such as hepatitis B and delta hepatitis, chronic alcoholism, non-alcoholic steatohepatitis, extrahepatic obstructions (e.g., stones in the bile duct), cholangiopathies (e.g., primary biliary cirrhosis and sclerosing cholangitis), autoimmune liver disease, and inherited metabolic disorders (e.g., Wilson's disease, hemochromatosis, and alpha- 1 anti trypsin deficiency).
  • viruses such as hepatitis B and delta hepatitis, chronic alcoholism, non-alcoholic steatohepatitis, extrahepatic obstructions (e.g., stones in the bile duct), cholangiopathies (e.g., primary biliary cirrhosis and sclerosing
  • liver fibrosis has traditionally focused on eliminating a primary injury. For extrahepatic obstructions, biliary decompression is the recommended mode of treatment whereas patients with Wilson's disease are treated with zinc acetate. Treatments for other chronic liver diseases such as hepatitis B, autoimmune hepatitis and Wilson's disease are also associated with many side effects, while primary biliary cirrhosis, primary sclerosing cholangitis and nonalcoholic fatty liver disease have no effective treatment other than liver transplantation.
  • provided methods are useful for treating renal diseases, disorders, and conditions. In some embodiments, provided methods are useful for reducing fibrosis of the kidney in a subject in need thereof. In some embodiments, provided methods are useful for treating a kidney disease, disorder, or condition characterized by or otherwise associated with fibrosis.
  • the present disclosure encompasses the recognition that treating fibrosis (e.g., using provided methods) instead of the underlying etiology may allow for broadly applicable antifibrotic kidney therapies.
  • kidney diseases, disorders, and conditions in which fibrosis is the sole or a predominant component may be suitable for treating kidney diseases, disorders, and conditions in which fibrosis is the sole or a predominant component, as well as those in which fibrosis is a secondary component (e.g., a symptom and/or result of an underlying disease, disorder, or condition).
  • provided methods are useful for treating acute injuries (e.g., acute organ injuries, such as acute kidney injury), as well as for treating chronic injuries (e.g., chronic kidney injury).
  • provided methods are useful for treating fibrosis associated with an acute injury, such as that incurred from trauma and/or surgery.
  • provided methods are useful for treating damaged and/or ischemic organs, transplants, or grafts, as well as ischemia/reperfusion injury or post-surgical scarring.
  • provided methods are useful for treating renal fibrosis.
  • provided methods are useful for treating renal fibrosis secondary to, or otherwise associated with, an underlying indication.
  • provided methods are useful for treating renal fibrosis associated with renal failure, renal obstruction, renal trauma, renal transplantation, chronic kidney disease, diabetes, hypertension, radiocontrast nephropathy, immune-mediated glomerulonephritides (e.g., lupus nephritis, ANCA-associated glomerulonephritides (e.g., Wegener’s granulomatosis, microscopic polyangiitis, or renal limited vasculitis), anti-GBM nephropathy, IgA nephropathy, membranous glomerulonephritis, or focal and segmental glomerulosclerosis), non-immune-mediated glomerulonephritides (e.g., autosomal dominant polycystic kidney disease, collagen type
  • provided methods are useful for treating nephrotic syndrome and/or diseases, disorders, or conditions associated with nephrotic syndrome (e.g., focal and segmental glomerulosclerosis, minimal change disease, and membranous nephropathy).
  • nephrotic syndrome e.g., focal and segmental glomerulosclerosis, minimal change disease, and membranous nephropathy.
  • provided methods are useful for treating a fibrotic disease of the kidney that is or comprises: focal segmental glomerulosclerosis (FSGS), steroid resistant nephrotic syndrome (SRNS), proteinuria, lupus nephritis, minimal change disease, an anti- neutrophil cytoplasmic antibody (ANCA)-associated glomerulonephritis, Alport syndrome, anti- globular basement membrane (anti-GBM) nephropathy, IgA nephropathy, membranous glomerulonephritis (MG), autosomal dominant polycystic kidney disease (ADPKD), or chronic kidney disease.
  • FSGS focal segmental glomerulosclerosis
  • SRNS steroid resistant nephrotic syndrome
  • proteinuria lupus nephritis
  • minimal change disease an anti- neutrophil cytoplasmic antibody (ANCA)-associated glomerulonephritis
  • ANCA anti- neutrophil cytoplasmic antibody
  • Alport syndrome anti
  • provided methods are useful for treating a fibrotic disease of the kidney that is or comprises an anti-neutrophil cytoplasmic antibody (ANCA)- associated glomerulonephritis.
  • ANCA-associated glomerulonephritis is selected from Wegener’s granulomatosis, microscopic polyangiitis (MPA), or renal limited vasculitis.
  • provided methods are useful for treating focal and segmental glomerulosclerosis.
  • provided methods are useful for treating Alport syndrome.
  • provided methods are useful for treating polycystic kidney disease (e.g., autosomal dominant polycystic kidney disease or autosomal recessive polycystic kidney disease).
  • provided methods are useful for treating primary proteinuric kidney disease (PPKD).
  • PPKD primary proteinuric kidney disease
  • PGDs primary glomerular diseases
  • PGDs are among the leading causes of chronic kidney disease and end-stage kidney disease in the world. PGDs predominantly affect younger patients, significantly reducing their quality of life, productivity, and longevity.
  • FSGS, membranous nephropathy (MN), and IgA nephropathy are among the three most common primary glomerular diseases in adults. Accordingly, in some embodiments, provided methods are useful for treating FSGS. In some embodiments, provided methods are useful for treating MN. In some embodiments, provided methods are useful for treating IgA nephropathy.
  • CNIs calcineurin inhibitors
  • Some of the drugs have a narrow therapeutic index necessitating close monitoring of the drug levels.
  • Long-term use of CNIs is associated with hypertension, nephrotoxicity, and metabolic abnormalities such as diabetes and dyslipidemia.
  • cessation of calcineurin inhibitors results in the relapse of proteinuria (Meyrier, A. et al., Kidney International. 1994;45(5): 1446-56).
  • a significant number of patients eventually become either resistant or dependent on these toxic agents.
  • Some of these glomerular diseases also recur after renal transplantation posing unique management problems (Choy, B.Y., et al., Am. J. Transplant. 2006;6(l l):2535-42).
  • provided methods are useful for treating patients with proteinuria (e.g., persistent proteinuria). It is well established that higher rates of urinary protein excretion are associated with worse prognosis, and therapies that reduce proteinuria are desirable for improving renal outcomes.
  • Patients with persistent proteinuria e.g., who continue to have > 1 gram of proteinuria per day
  • EKD end-stage kidney disease
  • eGFR estimated glomerular filtration rate
  • Patients with persistent proteinuria also develop further complications of chronic kidney disease (CKD) such as dyslipidemia, cardiovascular disease, abnormalities in mineral-bone metabolism, and hypertension, resulting in significant increases in morbidity and mortality and utilization of health care resources.
  • CKD chronic kidney disease
  • RAAS renin-angiotensin-aldosterone system
  • ARB angiotensin-receptor blockers
  • RAAS blockers reduce proteinuria and improve clinical outcomes in proteinuric renal diseases regardless of the etiology.
  • Other standard of care recommendations include aggressive blood pressure control ( ⁇ 130/80 mmHg), and HMG-CoA reductase inhibitors (e.g., statins) in patients with hyperlipidemia.
  • statins HMG-CoA reductase inhibitors
  • the inhibitors of the mineralocorticoid receptor and sodium glucose cotransporter-2 (SGLT-2) are increasingly being used in these patients as well.
  • provided methods are useful for treating primary glomerular diseases (e.g., FSGS, membranous nephropathy, or IgA nephropathy) and persistent proteinuria.
  • primary glomerular diseases e.g., FSGS, membranous nephropathy, or IgA nephropathy
  • persistent proteinuria e.g., FSGS, membranous nephropathy, or IgA nephropathy
  • PDGFRP Platelet- derived growth factor receptor beta
  • a kidney disease to be treated by methods of the present disclosure is nephrotic syndrome (NS).
  • NS is a group of rare renal diseases, including focal and segmental glomerulosclerosis (FSGS), minimal change disease (MCD), and membranous nephropathy.
  • FSGS focal and segmental glomerulosclerosis
  • MCD minimal change disease
  • membranous nephropathy is a rare disease that attacks the kidney’s filtering units (glomeruli) causing serious scarring which leads to permanent kidney damage and even failure (Fogo, A.B. Nat. Rev. Nephrol. 2015 Feb;l l(2):76-87, PMCID:PMC4772430). It will be appreciated that there are at least three types of FSGS.
  • Primary FSGS is FSGS that has no known cause (also referred to as idiopathic FSGS). Secondary FSGS is caused by one or more factors such as infection, drug toxicity, diseases such as diabetes or sickle cell disease, obesity, or other kidney diseases. Genetic FSGS (also called familial FSGS) is caused by one or more genetic mutations. Primary FSGS is idiopathic in nature. Manifestations of this disease include hypoalbuminemia and edema, lipid abnormalities and nephrotic range proteinuria. More than 5400 patients are diagnosed with FSGS every year (O’Shaughnessy, M.M., et al. Nephrol. Dial. Transplant 2018 Apr l;33(4):661-9).
  • a kidney disease to be treated by methods of the present disclosure is minimal change disease (MCD).
  • MCD is a kidney disease in which large amounts of protein are lost in the urine. It is one of the most common causes of the nephrotic syndrome worldwide. In children, MCD is usually primary (or idiopathic), but in adults, the disease is usually secondary. Secondary causes for MCD include allergic reactions, use of certain painkillers such as non-steroidal anti-inflammatory drugs (NSAIDs), tumors, or viral infections.
  • a kidney disease to be treated by methods of the present disclosure is membranous glomerulonephritis (MG or MGN). MG is a slowly progressive renal disease caused by immune complex formation in the glomerulus. Immune complexes are formed by binding of antibodies to antigens in the glomerular basement membrane. The antigens may be part of the basement membrane, or deposited from elsewhere by the systemic circulation.
  • a kidney disease to be treated by methods of the present disclosure is anti -neutrophil cytoplasmic antibody (ANCA)-associated glomerulonephritis.
  • ANCA-associated glomerulonephritis is a rapidly progressive renal disease and includes, e.g., Wegener's granulomatosis, microscopic polyangiitis, and renal limited vasculitis.
  • Wegener's granulomatosis is an organ- and/or life-threatening autoimmune disease of unknown etiology.
  • the classical clinical triad consists of necrotizing granulomatous inflammation of the upper and/or lower respiratory tract, necrotizing glomerulonephritis, and an autoimmune necrotizing systemic vasculitis affecting predominantly small vessels.
  • the detection of anti-neutrophil cytoplasmic antibodies directed against proteinase 3 (PR3-ANCA) is a highly specific indicator for Wegener's granulomatosis.
  • Microscopic polyangiitis is a disorder that causes blood vessel inflammation (vasculitis), which can lead to organ damage.
  • the kidneys, lungs, nerves, skin, and joints are the most commonly affected areas of the body.
  • MPA is diagnosed in people of all ages, all ethnicities, and both genders. The cause of this disorder is unknown.
  • Renal limited vasculitis is a type of anti-neutrophil cytoplasmic antibody (ANCA)-associated vasculitis that presents with only a renal manifestation; no other organs, including lungs, are involved.
  • ANCA anti
  • a kidney disease to be treated by methods of the present disclosure is lupus nephritis.
  • Lupus nephritis is inflammation of the kidney that is caused by an autoimmune disease, systemic lupus erythematous (SLE). With lupus, the body's immune system targets its own body tissues; lupus nephritis occurs when lupus involves the kidneys.
  • a kidney disease to be treated by methods of the present disclosure is anti-globular basement membrane (anti-GBM) nephropathy.
  • Anti-GBM nephropathy is a disease that occurs as a result of injury to small blood vessels (capillaries) in the kidneys and/or lungs.
  • capillaries small blood vessels
  • autoantibodies are targeted to the basement membrane in capillary blood vessels of the kidneys and lung, where they target and damage GBM.
  • a kidney disease to be treated by methods of the present disclosure is IgA nephropathy, also known as Berger’s disease.
  • IgA nephropathy is a kidney disease that occurs when IgA deposits build up in the kidneys, causing inflammation that damages kidney tissues.
  • IgA nephropathy affects the kidneys by attacking the glomeruli. The buildup of IgA deposits inflames and damages the glomeruli, causing the kidneys to leak blood and protein into the urine. The damage may lead to scarring of the nephrons that progresses slowly over many years. Eventually, IgA nephropathy can lead to end-stage kidney disease.
  • a kidney disease to be treated by methods of the present disclosure is Alport syndrome (AS).
  • AS is a genetic condition characterized by kidney disease, hearing loss, and eye abnormalities. Most affected individuals experience progressive loss of kidney function, usually resulting in end-stage kidney disease.
  • Alport syndrome is inherited in an X-linked manner and is caused by mutation(s) in the COL4A5 gene. In other cases, it can be inherited in either an autosomal recessive, or rarely in an autosomal dominant manner, and is caused by mutation(s) in the COL4A3 and/or COL4A4 genes.
  • Current therapies include hearing aid, hemodialysis, peritoneal dialysis and kidney transplantation.
  • a kidney disease to be treated by methods of the present disclosure is polycystic kidney disease (e.g., autosomal recessive polycystic kidney disease (ARPKD) - congenital hepatic fibrosis (CHF)).
  • ARPKD-CHF is a highly aggressive fibropolycystic disease that is characterized by the formation and expansion of fluid-filled cysts in the kidneys, enlargement of the kidneys and progressive fibrosis of both the kidney and the liver (Hartung, E.A., and Guay -Woodford, L.M. Pediatrics 2014 Sep;134(3):e833-e845; Gunay- Aygun, M., et al. J.
  • ARPKD-CHF congenital hepatic fibrosis
  • Caroli’s disease manifests as cystic dilatation of the intrahepatic ducts, often accompanies ARPKD-CHF (Sung, J.M., et al. Clin. Nephrol. 1992 Dec;38(6):324-8).
  • a subject is suffering from, susceptible to, or at risk of Caroli’s disease.
  • the need for transplantation is often driven by both progressive organ dysfunction and by significant enlargement of the diseased organ(s), and is accompanied by severe pain (www.arpkdchf.org).
  • a kidney disease to be treated by methods of the present disclosure is or comprises renal cysts.
  • Aberrant signaling by tyrosine kinases including platelet- derived growth factor (PDGF) and vascular endothelial growth factor (VEGF) and their receptors (R), PDGFR and VEGFR/KDR, respectively, has been implicated in the formation and expansion of renal cysts.
  • PDGF platelet- derived growth factor
  • VEGF vascular endothelial growth factor
  • R vascular endothelial growth factor
  • a PDGF-driven ciliopathy and/or overexpression of PDGF in the cyst lining and adjacent tubules are thought to, in part, drive renal cystic disease (Torres, V.E., et al. Lancet 2007 Apr 14;369(9569): 1287-301; Park. J.H. et al.
  • a kidney disease to be treated by methods of the present disclosure is collagen type III glomerulopathy.
  • Collagen type III glomerulopathy also known as collagenic or collagenofibrotic glomerulopathy, is characterized by pathological accumulation of collagen type III in glomeruli.
  • Collagen type III glomerulopathy presents either in childhood, often with a family history suggesting autosomal recessive inheritance, or in adults as a sporadic occurrence.
  • Proteinuria is a typical manifestation, with progression to end stage renal disease (ESRD) in approximately 10 years.
  • ESRD end stage renal disease
  • a kidney disease to be treated by methods of the present disclosure is nail-patella syndrome.
  • Nail-patella syndrome is a multi-organ disorder caused by mutations in the LMX1B gene. Nail-patella syndrome manifests with orthopedic and cutaneous deformities, as well as kidney complications due to development of structural lesions of collagen type III within glomerular basement membranes. Although the structural lesions may be asymptomatic, they are usually accompanied by proteinuria.
  • VEGF-driven angiogenesis is also thought to contribute to the growth of renal cysts, and inhibition of VEGFR/KDR signaling is associated with decreased tubule cell proliferation, decreased cystogenesis, and blunted renal enlargement (Bello-Ruess, E., et al. Kidney Int. 2001 Jul;60(l):37-45; Schrijvers, B.F., et al. Kidney Int. 2004 Jun;65(6):2003-17). Nevertheless, the role of VEGF in fibropoly cystic disease is more controversial, with at least some reports suggesting that this growth factor might be associated with disease mitigation (Spirli, C., et al. Gastroenterology 2010 Jan;138(l):360-71).
  • provided methods are useful for treating dermal diseases, disorders, or conditions. In some embodiments, provided methods are useful for treating dermal fibrosis. In some embodiments, provided methods are useful for treating dermal fibrosis secondary to, or otherwise associated with, an underlying indication. In some embodiments, provided methods are useful for treating scleroderma and/or systemic sclerosis (e.g., diffuse systemic sclerosis or limited systemic sclerosis).
  • systemic sclerosis e.g., diffuse systemic sclerosis or limited systemic sclerosis.
  • a fibrotic disease to be treated by methods of the present disclosure is scleroderma and/or systemic sclerosis (SSc).
  • Scleroderma which literally means hard skin, is a chronic fibrotic disorder of unknown etiology that affects the skin and other internal organs (SSc) (www.scleroderma.org).
  • SSc systemic sclerosis
  • Many patients who suffer from scleroderma/SSc also have loss of lung function.
  • Scleroderma/SSc and related diseases afflict approximately 400,000 to 990,000 people in the USA every year. Mortality and morbidity in scleroderma/SSc are very high and directly related to the extent of fibrosis of the involved organs (Hinchcliff, M.
  • Scleroderma/SSc can be classified in terms of the degree and location of the skin involvement and has been categorized into two major groups - diffuse and limited.
  • the diffuse form of scleroderma/SSc involves symmetric thickening of skin of the extremities, face and trunk. Organs affected include the esophagus, intestines, lungs, heart, and kidneys (Mayes, M. D. Semin. Cutan. Med. Surg. 1998 Mar;17(l):22-6; Jacobsen, L. et al. J. Am. Acad. Dermatol. 2003 Aug;49(2):323-5).
  • the limited form of scleroderma/SSc tends to be confined to the skin of fingers and face.
  • scleroderma/SSc The limited form of scleroderma/SSc is the CREST variant of scleroderma/SSc based on the clinical pattern of calcinosis with tiny deposits of calcium in the skin, Raynaud's phenomenon in the fingers, toes, nose, tongue, or ears, poor functioning of muscle of esophagus, sclerodactyly of the skin of the fingers or toes, and telangiectasias on the face, hands and mouth (Winterbauer, R.H. Bull. Johns Hopkins Hospital 1964;114:361-83; Wollheim, F.A. Classification of systemic sclerosis. Visions and reality. Rheumatology (Oxford) 2005).
  • fibrotic pathways are activated in scleroderma/SSc for reasons that are not completely understood.
  • the pathogenesis of fibrosis in scleroderma/SSc involves a complex set of interactions involving immune activation, microvascular damage and the activation of fibroblasts.
  • Scleroderma/SSc is characterized by excessive deposition of collagen in the skin and other involved organs and abnormalities of blood vessels (Jimenez, S. A., et al. Rheum. Dis. Clin. North Am. 1996 Nov;22(4):647-74; Sakkas, L. I. Autoimmunity 2005 Mar;38(2): 113-6).
  • TGF ⁇ 1 a multifunctional cytokine, is an indirect mitogen for human fibroblasts, which through upregulating PDGF, is capable of inducing normal fibroblasts into a pathogenic myofibroblast phenotype that mediates ECM (collagen) accumulation (Mauch, C., et al. J. Invest. Dermatol. 1993 Jan;100(l):92S-96S; Hummers, L. K., et al. J. Rheumatol. 2009 Mar;36(3):576-82).
  • the ubiquitous growth factors TGFP and PDGF are the most potent proteins involved in fibroblast proliferation, collagen gene expression and connective tissue (collagen) accumulation (Antoniades, H.N. Baillieres Clin.
  • the pathogenic cascade at different stages of scleroderma/SSc may have autoimmune, inflammatory, fibrotic and vascular components with systemic fibrosis and vasculopathy.
  • provided methods are useful for treating gastrointestinal diseases, disorders, or conditions.
  • provided methods are useful for treating gastrointestinal fibrosis (e.g., fibrosis of esophagus, stomach, intestines, and/or colon).
  • provided methods are useful for treating gastrointestinal fibrosis secondary to, or otherwise associated with, an underlying indication.
  • provided methods are useful for treating inflammatory bowel disease (e.g., ulcerative colitis or Crohn’s disease), e.g., treating gastrointestinal fibrosis associated with inflammatory bowel disease.
  • a disease to be treated by methods of the present disclosure is inflammatory bowel disease (IBD).
  • IBD is an inflammatory condition that comprises both ulcerative colitis (UC) and Crohn's disease (CD). While UC affects the entire colon, CD typically affects the ileum but can occur to any part of GI tract. IBD can manifest as acute or chronic colitis, characterized by recurrent intestinal inflammation accompanied by diarrhea and abdominal pain (Arivarasu, N., et al. Tissue Barriers 2018;6(2):el463897; Ponder, A. and Long, M.D. Clin. Epidemiol. 2013;5:237-47).
  • IBD Incidence of IBD is increasing worldwide and is an expanding global health problem (Amosy, E., et al. Clin. Med. Insights Gastroenterol. 2013;6:33-47). An estimated 2.5-3 million people in Europe are affected by IBD (Burisch, J., et al. J. Crohns Colitis 2013 May;7(4):322- 37). According to the Centers for Disease Control and Prevention (CDC), 3.1 million adults in this country were diagnosed with IBD in 2015, a substantial increase from the ⁇ 1.4 million adults diagnosed per 2008 reports (www.cdc.gov/IBD; www.cdc.gov/ibd/pdf/inflammatory- bowel-disease-an-expensive-disease.pdf).
  • IBD accounts for -1,300,000 physician visits and -92,000 hospitalizations each year in the United States. Of these, 75% patients diagnosed with CD and 25% patients diagnosed with UC and require surgery. Risk factors associated with IBD include environmental, genetic and immunologic factors (Abegunde, A.T., et al. World J. Gastroenterol. 2016 Jul 21;22(27):6296-6317; Frolkis, A., et al. Can. J. Gastroenterol. 2013 Mar;27(3):e28-24). [0138] IBD is a major cause of morbidity in patients and is a major consumer of the health care budget. A European study estimated that direct healthcare costs for IBD in Europe are ⁇ 5 billion Euros/year (Bursich 2013).
  • IBD is an autoimmune disease with excessive activation of the adaptive immune response.
  • Various factors including genetic factors alter the intestinal flora and trigger an inflammatory reaction, activate T cells, B cells, mast cells, macrophages and microglia, smooth muscle cells and fibroblasts in the colon, inducing mucosal disruption (Hildner, K., et al. Dig. Dis. 2016;34Suppl 1 :40-7; Curciarello, R., et al. Front Med. (Lausanne) 2017 Aug 7;4: 126).
  • Activated fibroblasts i.e. myofibroblasts, secrete growth factors including platelet derived growth factor (PDGF) and vascular endothelial growth factor (VEGF) (Scaldaferri, et al. Gastroenterology 2009 Feb;136(2):585-95.e5).
  • PDGF platelet derived growth factor
  • VEGF vascular endothelial growth factor
  • angiogenesis is also an important part of IBD pathogenesis in the colon of IBD patients.
  • Alkim, et al. demonstrated enhanced microvessel density in the intestinal tissue of both UC and CD patients, which correlated both the level of local VEGF expression and disease activity (Int. J. Inflam. 2015;2015:970890).
  • Anti-inflammatory drugs including 5-aminosalicylic acid (5-ASA)-based preparations, are often the first line of therapy in IBD (Segars, L.W., et al. Clin. Pharm. 1992 Jun; 11(6):514-28).
  • Anti-TNFa antibodies such as infliximab and adalimumab are also being used. Nevertheless, patients treated with adalimumab are at increased risk for serious infections and lymphoma (Dulai, P.S., et al. Clin. Gastroenterol. Hepatol. 2014 Sep; 12(9): 1443-51).
  • PDGF activates fibroblasts and IBD-fibroblasts proliferate more rapidly than normal fibroblasts; collagen secretion from IBD patients’ fibroblasts was increased compared to collagen secretion by normal fibroblasts. IBD is also associated with increased circulating PDGF and the level of this growth factor has been reported to correspond with disease severity (Andrae, J., et al. Genes Dev. 2008 May 15;22(10): 1276-1312).
  • angiogenesis as a novel component of IBD pathogenesis and angiogenic activity is increased in IBD patients.
  • Serum VEGF levels were significantly higher in IBD patients compared to controls in several studies.
  • Griga et al. demonstrated that sources of increased serum VEGF were from inflamed intestinal tissue of IBD patients (Scand. J. Gastroenterol. 1998 May;33(5):504-8; Hepatogastroenterology 2002 Jan-Feb;49(43): 116-23; Hepatogastroenterology 1999 Mar-Apr;46(26):920-3; Eur. J. Gastroenterol. Hepatol. 1999 Feb; 11(2): 175-9).
  • VEGF expression was markedly increased in the inflamed mucosa of both CD and UC patients, when compared with normal mucosa of the same patient. Studies also showed that VEGF expression was increased in colon and was higher across all IBD groups (both CD and UC) when compared with healthy controls. Scaldaferri, et al. (2009) reported that VEGF receptor (VEGFR/KDR) levels were increased in intestinal samples of IBD patients, and in mice with experimental colitis.
  • provided methods are useful for treating certain other diseases, disorders, or conditions.
  • provided methods are useful for treating cardiac fibrosis and/or fibrosis associated with cardiovascular system.
  • provided methods are useful for treating cardiac fibrosis secondary to, or otherwise associated with, an underlying indication.
  • provide methods are useful for treating cardiac and/or cardiovascular fibrosis associated with ischemic heart disease, myocardial ischemia, athereosclerosis, myocardial perfusion (e.g., as a consequence of chronic cardiac ischemia or myocardial infarction), vascular occlusion, or restenosis.
  • a disease to be treated by methods of the present disclosure is ischemic heart disease.
  • Ischemic heart disease is a leading cause of morbidity and mortality in the US, afflicting millions of Americans each year at a cost expected to exceed $300 billion/year.
  • Numerous pharmacological and interventional approaches are being developed to improve treatment of ischemic heart disease including reduction of modifiable risk factors, improved revascularization procedures, and therapies to halt progression and/or induce regression of atherosclerosis.
  • atherosclerosis comprises a fibrotic component.
  • provided methods are useful for treating fibrosis associated with central nervous system (CNS) and/or one or more CNS-related diseases, disorders, or conditions.
  • CNS central nervous system
  • provided methods are useful for treating CNS-associated fibrosis secondary to, or otherwise associated with, an underlying indication.
  • provided methods are useful for treating fibrosis associated with cerebral infarction, stroke, or amyotrophic lateral sclerosis.
  • provided methods are useful for treating fibrosis associated with musculoskeletal system and/or one or more musculoskeletal diseases, disorders, or conditions. In some embodiments, provided methods are useful for treating musculoskeletal- associated fibrosis secondary to, or otherwise associated with, an underlying indication. In some embodiments, provided methods are useful for treating fibrosis associated with muscular dystrophy.
  • provided methods are useful for treating pancreatic fibrosis. In some embodiments, provided methods are useful for treating pancreatic fibrosis secondary to, or otherwise associated with, an underlying indication. In some embodiments, provided methods are useful for treating fibrosis associated with pancreatitis.
  • one or more subjects or populations are selected to receive Compound 1 as described herein based on one or more markers and/or characteristics such as, for example, one or more risk factors of fibrosis or an associated disease, disorder or condition and/or an altered level of one or more biomarkers, etc.
  • a subject or population thereof is selected to receive Compound 1 using technologies provided herein (e.g., based on assessment of one or more markers and/or characteristics, such as an assessment of one or more biomarkers described herein).
  • technologies are used to inform or determine one or more features of a therapeutic regimen (e.g., selection of subject(s) to receive a particular therapy (e.g., Compound 1 therapy) and/or dose thereof and/or timing of administration of such therapy).
  • assessment of one or more markers and/or characteristics is performed with respect to the same subject at a plurality of different time points.
  • assessment of one or more markers and/or characteristics is performed with respect to a particular patient prior to initiation of a particular therapeutic regimen (e.g., a Compound 1 therapeutic regimen) and/or prior to administration of a particular dose of therapy (e.g., Compound 1 therapy) in accordance with such therapeutic regimen.
  • a particular therapeutic regimen e.g., a Compound 1 therapeutic regimen
  • a particular dose of therapy e.g., Compound 1 therapy
  • a subject or population thereof is suffering from or is susceptible to fibrosis. In some embodiments, a subject or population thereof is suffering from or is susceptible to a disease, disorder, or condition characterized by or otherwise associated with fibrosis. In some embodiments, a subject or population thereof is suffering from or is susceptible to fibrosis of gastrointestinal tract, heart, kidney, lung, liver, muscle, pancreas, and/or skin.
  • a subject or population thereof is suffering from or is susceptible to an acute injury (e.g., an acute organ injury, such as acute lung injury, acute liver injury, or acute kidney injury).
  • a subject or population thereof is suffering from or is susceptible to a chronic injury (e.g., a chronic organ injury, such as chronic lung injury, chronic liver injury, or chronic kidney injury).
  • a subject or population thereof is suffering from a traumatic injury.
  • a subject or population thereof has undergone, is undergoing, or will undergo an organ transplantation.
  • a subject or population thereof is suffering from or susceptible to a damaged and/or ischemic organ, transplant, or graft.
  • a subject or population thereof is suffering from or susceptible to ischemia/reperfusion injury.
  • a subject or population thereof is suffering from or susceptible to post-surgical scarring.
  • a subject or population thereof is suffering from or is susceptible to a pulmonary disease as described herein. In some embodiments, a subject or population thereof is suffering from or is susceptible to pulmonary fibrosis. In some embodiments, a subject or population thereof is suffering from or is susceptible to pulmonary fibrosis secondary to, or otherwise associated with, an underlying indication. In some embodiments, a subject or population thereof is suffering from or is susceptible to interstitial lung disease (e.g., fibrosing interstitial lung disease). In some embodiments, a subject or population thereof is suffering from or is susceptible to idiopathic interstitial pneumonia. In some embodiments, a subject or population thereof is suffering from or is susceptible to idiopathic pulmonary fibrosis.
  • interstitial lung disease e.g., fibrosing interstitial lung disease
  • a subject or population thereof is suffering from or is susceptible to idiopathic interstitial pneumonia. In some embodiments, a subject or population thereof is suffering from or is susceptible to idiopathic pulmonary
  • a subject or population thereof is suffering from or is susceptible to a hepatic disease as described herein.
  • a subject or population thereof is suffering from or is susceptible to hepatic fibrosis (e.g., fibrotic liver disease).
  • a subject or population thereof is suffering from or is susceptible to cirrhosis.
  • a subject or population thereof is suffering from or is susceptible to hepatic fibrosis secondary to, or otherwise associated with, an underlying indication.
  • a subject or population thereof is suffering from or is susceptible to hepatitis C, hepatitis B, delta hepatitis, chronic alcoholism, nonalcoholic steatohepatitis (NASH), extrahepatic obstructions (e.g., stones in bile duct), cholangiopathies (e.g., primary biliary cirrhosis or sclerosing cholangitis), autoimmune liver disease, or inherited metabolic disorders (e.g., Wilson’s disease, hemochromatosis, or alpha-1 antitrypsin deficiency).
  • a subject or population thereof is suffering from or is susceptible to a kidney disease as described herein.
  • a subject or population thereof is suffering from or is susceptible to fibrotic disease of the kidney as described herein. In some embodiments, a subject or population thereof is suffering from or is susceptible to a disease, disorder, or condition characterized by or otherwise associated with fibrosis disease of the kidney as described herein.
  • a subject or population thereof is suffering from or is susceptible to an acute kidney injury. In some embodiments, a subject or population thereof is suffering from or is susceptible to a chronic kidney injury. In some embodiments, a subject or population thereof is suffering from a traumatic injury. In some embodiments, a subject or population thereof has undergone, is undergoing, or will undergo an organ transplantation. In some embodiments, a subject or population thereof is suffering from or susceptible to a damaged and/or ischemic organ, transplant, or graft. In some embodiments, a subject or population thereof is suffering from or susceptible to ischemia/reperfusion injury. In some embodiments, a subject or population thereof is suffering from or susceptible to post-surgical scarring.
  • a subject or population thereof is suffering from or is susceptible to renal fibrosis. In some embodiments, a subject or population thereof is suffering from or is susceptible to renal fibrosis secondary to, or otherwise associated with, an underlying indication.
  • a subject or population thereof is suffering from or is susceptible to renal failure, renal obstruction, renal trauma, renal transplantation, chronic kidney disease, diabetes, hypertension, radiocontrast nephropathy, immune-mediated glomerulonephritides (e.g., lupus nephritis, ANCA-associated glomerulonephritides (e.g., Wegener’s granulomatosis, microscopic polyangiitis, or renal limited vasculitis), anti-GBM nephropathy, IgA nephropathy, membranous glomerulonephritis, or focal and segmental glomerulosclerosis), non-immune-mediated glomerulonephritides (e.g., polycystic kidney disease, collagen type III glomerulopathy, nail-patella syndrome, or Alport syndrome), minimal change disease, or nephrotic syndrome (e.g., steroid-resistant nephrotic syndrome).
  • a subject or population thereof is suffering from or is susceptible to a fibrotic disease of the kidney that is or comprises: focal segmental glomerulosclerosis (FSGS), steroid resistant nephrotic syndrome (SRNS), proteinuria, lupus nephritis, minimal change disease, an anti-neutrophil cytoplasmic antibody (ANCA)-associated glomerulonephritis, Alport syndrome, anti-globular basement membrane (anti-GBM) nephropathy, IgA nephropathy, membranous glomerulonephritis (MG), autosomal dominant polycystic kidney disease (ADPKD), or chronic kidney disease.
  • FSGS focal segmental glomerulosclerosis
  • SRNS steroid resistant nephrotic syndrome
  • proteinuria proteinuria
  • lupus nephritis minimal change disease
  • an anti-neutrophil cytoplasmic antibody (ANCA)-associated glomerulonephritis Alport syndrome
  • a subject or population thereof is suffering from or is susceptible to a fibrotic disease of the kidney that is or comprises an anti-neutrophil cytoplasmic antibody (ANCA)-associated glomerulonephritis.
  • ANCA-associated glomerulonephritis is selected from Wegener’s granulomatosis, microscopic polyangiitis (MPA), or renal limited vasculitis.
  • a subject or population thereof is suffering from or is susceptible to collagen type III glomerulopathy or nail-patella syndrome.
  • a subject or population thereof is suffering from or is susceptible to nephrotic syndrome and/or diseases, disorders, or conditions associated with nephrotic syndrome (e.g., focal and segmental glomerulosclerosis, minimal change disease, and membranous nephropathy).
  • a subject or population thereof is suffering from or is susceptible to focal and segmental glomerulosclerosis (FSGS).
  • FSGS focal and segmental glomerulosclerosis
  • a subject or population thereof is suffering from or is susceptible to collagen type III glomerulopathy.
  • a subject or population thereof is suffering from or is susceptible to nailpatella syndrome.
  • a subject or population thereof is suffering from or is susceptible to Alport syndrome.
  • a subject or population thereof is suffering from or is susceptible to polycystic kidney disease (e.g., autosomal dominant polycystic kidney disease or autosomal recessive polycystic kidney disease).
  • a subject or population thereof is suffering from or is susceptible to renal fibrosis and has an altered level of one or more biomarkers (e.g., an altered level of one or more biomarkers described herein).
  • a subject or population thereof is suffering from or is susceptible to primary proteinuric kidney disease (e.g., as confirmed from a renal biopsy).
  • a subject or population thereof is suffering from or is susceptible to primary glomerular diseases (e.g., as confirmed from a renal biopsy).
  • a subject or population thereof is suffering from or susceptible to persistent proteinuria.
  • a subject or population thereof is suffering from or susceptible to primary glomerular disease (e.g., as confirmed from a renal biopsy) and persistent proteinuria.
  • a subject or population thereof is suffering from or susceptible to proteinuric chronic kidney disease.
  • a subject or population thereof is suffering from or is susceptible to focal and segmental glomerulosclerosis. In some embodiments, a subject or population thereof is suffering from or is susceptible to membranous nephropathy. In some embodiments, a subject or population thereof is suffering from or susceptible to IgA nephropathy.
  • a subject or population thereof has one or more symptoms selected from proteinuria, hypoalbuminemia, hyperlipidemia, and edema.
  • a subject or population thereof has proteinuria.
  • a subject or population thereof has hypoalbuminemia.
  • a subject or population thereof has hyperlipidemia.
  • a subject or population thereof has edema.
  • a subject or population thereof is suffering from or is susceptible to a dermal disease as described herein. In some embodiments, a subject or population thereof is suffering from or is susceptible to dermal fibrosis. In some embodiments, a subject or population thereof is suffering from or is susceptible to dermal fibrosis secondary to, or otherwise associated with, an underlying indication. In some embodiments, a subject or population thereof is suffering from or is susceptible to scleroderma and/or systemic sclerosis (e.g., diffuse systemic sclerosis or limited systemic sclerosis). [0161] In some embodiments, a subject or population thereof is suffering from or is susceptible to a gastrointestinal disease as described herein.
  • a subject or population thereof is suffering from or is susceptible to gastrointestinal fibrosis (e.g., fibrosis of esophagus, stomach, intestines, and/or colon).
  • gastrointestinal fibrosis e.g., fibrosis of esophagus, stomach, intestines, and/or colon.
  • a subject or population thereof is suffering from or is susceptible to gastrointestinal fibrosis secondary to, or otherwise associated with, an underlying indication.
  • a subject or population thereof is suffering from or is susceptible to inflammatory bowel disease (e.g., ulcerative colitis or Crohn’s disease).
  • a subject or population thereof is suffering from or is susceptible to cardiac fibrosis and/or fibrosis associated with cardiovascular system. In some embodiments, a subject or population thereof is suffering from or is susceptible to cardiac fibrosis secondary to, or otherwise associated with, an underlying indication. In some embodiments, a subject or population thereof is suffering from or is susceptible to ischemic heart disease, myocardial ischemia, atherosclerosis, myocardial perfusion (e.g., as a consequence of chronic cardiac ischemia or myocardial infarction), vascular occlusion, or restenosis.
  • a subject or population thereof is suffering from or is susceptible to fibrosis associated with central nervous system (CNS) and/or one or more CNS- related diseases, disorders, or conditions.
  • CNS central nervous system
  • a subject or population thereof is suffering from or is susceptible to CNS-associated fibrosis secondary to, or otherwise associated with, an underlying indication.
  • a subject or population thereof is suffering from or is susceptible to cerebral infarction, stroke, or amyotrophic lateral sclerosis.
  • a subject or population thereof is suffering from or is susceptible to fibrosis associated with musculoskeletal system and/or one or more musculoskeletal diseases, disorders, or conditions.
  • a subject or population thereof is suffering from or is susceptible to musculoskeletal-associated fibrosis secondary to, or otherwise associated with, an underlying indication. In some embodiments, a subject or population thereof is suffering from or is susceptible to muscular dystrophy.
  • a subject or population thereof is suffering from or is susceptible to pancreatic fibrosis. In some embodiments, a subject or population thereof is suffering from or is susceptible to pancreatic fibrosis secondary to, or otherwise associated with, an underlying indication. In some embodiments, a subject or population thereof is suffering from or is susceptible to pancreatitis.
  • composition providing Compound 1, as described herein can be administered to subjects in accordance with methods provided herein.
  • a composition providing Compound 1 is a composition comprising Compound 1 (in a pharmaceutically acceptable form as described herein), formulated together with one or more pharmaceutically acceptable carriers.
  • a composition providing Compound 1 is or comprises Compound 1 present in a unit dose amount appropriate for administration in a therapeutic regimen that shows a statistically significant probability of achieving a predetermined therapeutic effect when administered to a relevant population.
  • compositions providing Compound 1 may be specially formulated for administration in solid or liquid form, including those adapted for the following: oral administration, for example, drenches (aqueous or non-aqueous solutions or suspensions), capsules, tablets, e.g., those targeted for buccal, sublingual, and systemic absorption, boluses, powders, granules, pastes for application to the tongue; parenteral administration, for example, by subcutaneous, intramuscular, intravenous or epidural injection as, for example, a sterile solution or suspension, or sustained-release formulation; topical application, for example, as a cream, ointment, or a controlled-release patch or spray applied to the skin, lungs, or oral cavity; intravaginally or intrarectally, for example, as a pessary, cream, or foam; sublingually; ocularly; transdermally; or nasally, pulmonary, and to other mucosal surfaces.
  • oral administration for example, drenches (aqueous or non
  • composition providing Compound 1 is formulated for oral administration (e.g., in a capsule form). In some embodiments, a composition providing Compound 1 is administered orally.
  • a composition providing Compound 1 is administered as one or more unit dosage forms.
  • a composition providing Compound 1 is administered as one or more solid unit dosage forms (e.g., one or more capsules or tablets).
  • Compound 1 is administered as one or more oral unit dosage forms.
  • a composition providing Compound 1 is an immediate release solid unit dosage form.
  • a composition providing Compound 1 is a capsule.
  • a composition providing Compound l is a tablet.
  • Compound 1 is administered as a capsule comprising 10 mg, 50 mg, or 250 mg of Compound 1. In some embodiments, Compound 1 is administered as a capsule comprising 10 mg, 50 mg, or 250 mg of Compound 1 with no excipients. In some embodiments, Compound 1 is administered as a capsule comprising 50 mg of Compound 1 (e.g., with no excipients). In some embodiments, Compound 1 is administered as a capsule comprising 250 mg of Compound 1 (e.g., with no excipients).
  • Compound 1 is administered as a capsule comprising 100 mg of Compound 1 (e.g., a Form A Compound 1 Hydrochloride Trihydrate). In some embodiments, Compound 1 is administered as a capsule comprising 200 mg of Compound 1 (e.g., a Form A Compound 1 Hydrochloride Trihydrate). In some such embodiments, a capsule comprising Compound 1 has no excipients.
  • unit dosage forms e.g., tablets or capsules
  • one unit dosage form e.g., tablet or capsule
  • a suitable dose e.g., a dose of about 50 mg, about 100 mg, about 200 mg, about 250 mg, about 400 mg, about 500 mg, or about 600 mg
  • more than one (e.g., 2, 3, 4, 5, etc.) unit dosage forms e.g., tablets or capsules
  • a suitable dose e.g., a dose of about 50 mg, about 100 mg, about 200 mg, about 250 mg, about 400 mg, about 500 mg, or about 600 mg.
  • each unit dosage form when multiple unit dosage forms are administered, each unit dosage form contains the same amount of Compound 1, in order to provide a suitable dose as described herein; in some embodiments, when multiple unit dosage forms are administered, each unit dosage form contains different amounts of Compound 1, in order to provide a suitable dose as described herein.
  • a suitable number of unit dosage forms are administered in order to provide a dose of about 50 mg, about 100 mg, about 150 mg, about 200 mg, about 250 mg, about 300 mg, about 350 mg, about 400 mg, about 450 mg, about 500 mg, about 550 mg, or about 600 mg.
  • one unit dosage form is administered to provide a dose of about 50 mg, about 100 mg, about 150 mg, about 200 mg, about 250 mg, about 300 mg, about 350 mg, about 400 mg, about 450 mg, about 500 mg, about 550 mg, or about 600 mg.
  • more than one (e.g., 2, 3, or 4) unit dosage forms are administered to provide a dose of about 50 mg, about 100 mg, about 150 mg, about 200 mg, about 250 mg, about 300 mg, about 350 mg, about 400 mg, about 450 mg, about 500 mg, about 550 mg, or about 600 mg.
  • a composition providing Compound 1 is administered as a single dose. In some embodiments, a composition providing Compound 1 is administered at regular intervals. Administration at an “interval,” as used herein, indicates that the therapeutically effective amount is administered periodically (as distinguished from a one-time dose). In some embodiments, a composition providing Compound 1 is administered bimonthly (Q2M), monthly (QM), twice monthly (BIM), triweekly (Q3W), biweekly (Q2W), weekly (QW), twice weekly (BIW), thrice weekly (TIW), daily (QD), twice daily (BID), thrice daily (TID), or four times a day (QID) in accordance with methods provided herein. In some embodiments, a composition providing Compound 1 is administered twice daily (BID). In some embodiments, a composition providing Compound 1 is administered once daily (QD).
  • a composition providing Compound 1 is administered in a suitable number of unit dosage forms (e.g., tablets or capsules) in order to provide a dose of about 100 mg, about 200 mg, about 300 mg, about 400 mg, about 500 mg, or about 600 mg once daily.
  • a composition providing Compound 1 is administered in a suitable number of unit dosage forms (e.g., tablets or capsules) in order to provide a dose of about 50 mg, about 100 mg, about 150 mg, about 200 mg, about 250 mg, about 300 mg, about 350 mg, or about 400 mg twice daily.
  • Compound 1 is administered in a daily dose of from about 50 mg to about 600 mg, from about 100 mg to about 600 mg, from about 200 mg to about 600 mg, from about 400 mg to about 600 mg, from about 50 mg to about 500 mg, from about 100 mg to about 500 mg, from about 250 mg to about 500 mg, from about 50 mg to about 250 mg, from about 100 mg to about 1000 mg, from about 200 mg to about 1000 mg, from about 500 mg to about 1000 mg, or from about 200 mg to about 500 mg. In some embodiments, Compound 1 is administered in a dose of about 50 mg, about 100 mg, about 200 mg, about 250 mg, about 400 mg, about 500 mg, or about 600 mg.
  • Compound 1 is administered in a dose of about 50 mg, about 100 mg, about 200 mg, about 250 mg, about 400 mg, about 500 mg, or about 600 mg. In some embodiments, Compound 1 is administered in a dose of about 50 mg, about 100 mg, about 150 mg, about 200 mg, about 250 mg, about 300 mg, about 350 mg, about 400 mg, about 450 mg, about 500 mg, about 550 mg, or about 600 mg. In some embodiments, Compound 1 is administered in a daily dose of about 50 mg, about 100 mg, about 200 mg, about 400 mg, about 500 mg, or about 600 mg. In some embodiments, Compound 1 is administered in a twice daily dose of about 50 mg, about 100 mg, about 250 mg, or about 500 mg.
  • Compound 1 is administered in a dose of about 100 mg once daily. In some embodiments, Compound 1 is administered in a dose of about 200 mg once daily. In some embodiments, Compound 1 is administered in a dose of about 300 mg once daily. In some embodiments, Compound 1 is administered in a dose of about 400 mg once daily. In some embodiments, Compound 1 is administered in a dose of about 500 mg once daily. In some embodiments, Compound 1 is administered in a dose of about 600 mg once daily.
  • Compound 1 is administered in a dose of about 50 mg twice daily. In some embodiments, Compound 1 is administered in a dose of about 100 mg twice daily. In some embodiments, Compound 1 is administered in a dose of about 150 mg twice daily. In some embodiments, Compound 1 is administered in a dose of about 200 mg twice daily. In some embodiments, Compound 1 is administered in a dose of about 250 mg twice daily. In some embodiments, Compound 1 is administered in a dose of about 300 mg twice daily. In some embodiments, Compound 1 is administered in a dose of about 350 mg twice daily. In some embodiments, Compound 1 is administered in a dose of about 400 mg twice daily.
  • a composition providing Compound 1 is administered at regular intervals indefinitely. In some embodiments, a composition providing Compound 1 is administered at regular intervals for a defined period of time.
  • a composition providing Compound 1 is administered to a subject in a fed state (e.g., after a meal, such as within 1 hour, 45 minutes, 30 minutes, or 15 minutes of a meal).
  • Compound 1 is administered to a subject in a fasted state (e.g., after a fast of at least 4 hours, at least 6 hours, at least 8 hours, at least 10 hours, at least 12 hours, or at least 16 hours).
  • a method comprising administering an effective amount of Compound 1 : to a patient that has been determined to have an altered level of one or more biomarkers selected from a collagen 6 biomarker and/or Table 1 or a human analog thereof.
  • a method of treating a fibrotic disease of the kidney comprising administering an effective amount of Compound 1 : to a patient that has been determined to have an altered level of one or more biomarkers selected from a collagen 6 biomarker and/or Table 1 or a human analog thereof.
  • a method comprising administering an effective amount of Compound 1 :
  • a method comprising administering an effective amount of Compound 1 : to a patient that has been determined to have (i) nephrotic syndrome; and (ii) an altered level of one or more biomarkers selected from a collagen 6 biomarker and/or Table 1 or a human analog thereof.
  • a method comprising administering an effective amount of Compound 1 : to a patient that has been determined to have an altered level of one or more biomarkers selected from Table 1 or a human analog thereof.
  • a method of treating a fibrotic disease comprising administering an effective amount of Compound 1 : to a patient that has been determined to have an altered level of one or more biomarkers selected from Table 1 or a human analog thereof.
  • a method comprising administering an effective amount of Compound 1 : to a patient that has been determined to have (i) proteinuria and/or hypoalbuminemia and/or hyperlipidemia and/or edema; and (ii) an altered level of one or more biomarkers selected from Table 1 or a human analog thereof.
  • a method comprising administering an effective amount of Compound 1 :
  • nephrotic syndrome a patient that has been determined to have (i) nephrotic syndrome; and (ii) an altered level of one or more biomarkers selected from Table 1 or a human analog thereof.
  • the threshold level corresponds to a predetermined mean or median level of the biomarker in a population of healthy subjects.
  • a method comprising:
  • a method comprising:
  • a method of treating a fibrotic disease comprising administering an effective amount of Compound 1 : to a patient that has been determined to have an altered level of one or more biomarkers, wherein the one or more biomarkers are selected from biomarkers or human analogs thereof whose levels have been established to have: a mean change in a population of subjects administered Compound 1 relative to a comparable reference population; and/or a mean change in a population of subjects with confirmed fibrotic disease relative to a population of healthy volunteers.
  • biomarkers are selected from biomarkers or human analogs thereof whose levels have been established to have: a mean change of at least about 1.5-fold, about 2-fold, about 3-fold, about 4-fold, about 5-fold, about 10-fold, or about 20-fold in a population of subjects administered Compound 1 relative to a comparable reference population; and/or a mean change of at least about 1.5-fold, about 2-fold, about 3-fold, about 4-fold, about 5-fold, about 10-fold, or about 20-fold in a population of subjects with confirmed fibrotic disease relative to a population of healthy volunteers.
  • the threshold level corresponds to a predetermined mean or median level of the biomarker in a population of healthy subjects.
  • FSGS focal segmental glomerulosclerosis
  • SRNS steroid resistant nephrotic syndrome
  • proteinuria proteinuria
  • lupus nephritis minimal change disease
  • ANCA antineutrophil cytoplasmic antibody
  • anti-GBM anti-globular basement membrane
  • IgA IgA nephropathy
  • MG membranous glomerulonephritis
  • ADPKD autosomal dominant polycystic kidney disease
  • collagen type III glomerulopathy nailpatella syndrome, Alport syndrome, or chronic kidney disease.
  • ANCA anti-neutrophil cytoplasmic antibody
  • MPA microscopic polyangiitis
  • a representative procedure for obtaining Compound 1 is as follows.
  • the quantities of materials used are approximate and may be increased or decreased in unison to obtain a larger or smaller lot size.
  • Conditions such as time or temperature are approximate and may be used as targets.
  • Step 1 Acetic anhydride (5.40 kg) and methyl 2-oxo-2,3-dihydro-lH-pyrrolo[2, 3- b]pyridine-6-carboxylate (1.0 kg) were added to a reactor at room temperature and stirred to combine. Trimethylorthobenzoate (1.90 kg) was added to the reaction mixture. The mixture was then heated to 105 °C and stirred for 1 hr. The reaction was cooled to 40 °C and isopropyl alcohol (3.14 kg) was added. The reaction was cooled further to 5 °C and stirred for 4 hr. The mixture was then filtered, and the product washed with isopropyl alcohol twice.
  • Step 3 Purified water (2.50 kg) and methyl (Z)-3-(((4-(N-methyl-2-(4- methylpiperazin-l-yl)acetamido)phenyl)amino)(phenyl)methylene)-2-oxo-2,3-dihydro-lH- pyrrolo[2,3-b]pyridine-6-carboxylate (1.0 kg) were added to a reactor at room temperature and stirred to combine. Acetone (1.975 kg) was then added, followed by iron-free HC1 (1.20 mol eq.). The mixture was stirred for 1 hr at 30 °C, and then filtered through a micron filter and washed with purified water (0.2 kg).
  • Compound 1 was formulated in a capsule for oral administration.
  • the capsule formulation included a Size 00 Swedish orange capsule containing Compound 1 (10 mg, 50 mg, or 250 mg) with no excipients.
  • Ingredients of the capsule shell were hypromellose (hydroxypropylmethyl cellulose), iron oxide as a coloring agent, and titanium dioxide as an opacifier.
  • Capsule formulations were prepared as follows. First, an optional sieving step was performed to deagglomerate the active agent if needed. Then, Compound 1 was filled into HPMC capsules, using either an automated Xelodose machine (e.g., for 10 mg and 50 mg capsules) or a semiautomated process (e.g., for 250 mg capsules). All capsules were polished or dedusted, either by an inline deduster (e.g., for 10 mg and 50 mg capsules) or a separate capsule polisher (e.g., for 250 mg capsules).
  • an automated Xelodose machine e.g., for 10 mg and 50 mg capsules
  • a semiautomated process e.g., for 250 mg capsules. All capsules were polished or dedusted, either by an inline deduster (e.g., for 10 mg and 50 mg capsules) or a separate capsule polisher (e.g., for 250 mg capsules).
  • FIG. l is a graph showing results of Compound 1 treatment on urine protein. Rats treated with Compound 1 displayed reduced urine protein on Study Day 17 (Treatment Day 14) compared to vehicle-treated animals.
  • FIG. 2 A and FIG. 2B are graphs showing change from Treatment Day 8 to Treatment
  • Rats treated with Compound 1 displayed reduced urine protein on Study Day 17 (Treatment Day 14) compared to vehicle-treated animals.
  • Kidney samples preserved with formalin were analyzed and scored blindly according to the following renal damage scale: 0 (no damage); 1 (>25% damage); 2 (26-50% damage); 3 (51-75% damage); or 4 (>75% damage).
  • the average renal damage score in the group treated with Compound 1 was significantly reduced compared to the vehicle-treated group.
  • RNA samples were quantified using Qubit 2.0 Fluorometer (Life Technologies, Carlsbad, CA, USA) and RNA integrity was checked using Agilent TapeStation 4200 (Agilent Technologies, Palo Alto, CA, USA).
  • RNA sequencing libraries were prepared using the NEBNext Ultra RNA Library Prep Kit for Illumina using manufacturer’s instructions (NEB, Ipswich, MA, USA). Briefly, mRNAs were initially enriched with Oligod(T) beads. Enriched mRNAs were fragmented for 15 minutes at 94 °C. First strand and second strand cDNA were subsequently synthesized. cDNA fragments were end repaired and adenylated at 3 ’ends, and universal adapters were ligated to cDNA fragments, followed by index addition and library enrichment by PCR with limited cycles.
  • the sequencing library was validated on the Agilent TapeStation (Agilent Technologies, Palo Alto, CA, USA), and quantified by using Qubit 2.0 Fluorometer (Invitrogen, Carlsbad, CA), as well as by quantitative PCR (KAPA Biosystems, Wilmington, MA, USA).
  • the sequencing libraries were clustered on a single lane of a flowcell. After clustering, the flowcell was loaded on the Illumina HiSeq instrument (4000 or equivalent) according to manufacturer’s instructions. The samples were sequenced using a 2xl50bp Paired End (PE) configuration. Image analysis and base calling were conducted by the HiSeq Control Software (HCS). Raw sequence data (.bcl files) generated from Illumina HiSeq were converted into fastq files and de-multiplexed using Illumina's bcl2fastq 2.17 software. One mismatch was allowed for index sequence identification.
  • HCS HiSeq Control Software
  • sequence reads were trimmed to remove possible adapter sequences and nucleotides with poor quality using Trimmomatic v.0.36.
  • the trimmed reads were mapped to the reference genome available on ENSEMBL using the STAR aligner v.2.5.2b.
  • the STAR aligner uses a splice aligner that detects splice junctions and incorporates them to help align the entire read sequences.
  • BAM files were generated as a result of this step.
  • Unique gene hit counts were calculated by using feature Counts from the Subread package v.1.5.2. Only unique reads that fell within exon regions were counted.
  • the gene hit counts table was used for downstream differential expression analysis.
  • DESeq2 a comparison of gene expression between the groups of samples was performed.
  • the Wald test was used to generate p-values and Log2 fold changes. Genes with adjusted p-values ⁇ 0.05 and absolute log2 fold changes > 1 were called as differentially expressed genes for each comparison.
  • a gene ontology analysis was performed on the statistically significant set of genes by implementing the software GeneSCF.
  • the mgi GO list was used to cluster the set of genes based on their biological process and determine their statistical significance.
  • a PCA analysis was performed using the "plotPCA" function within the DESeq2 R package.
  • a nephropathy patient with suspected or confirmed glomerular disease (e.g., suspected of having FSGS or MCD) is subjected to a renal biopsy. For example, mRNA is extracted from single glomeruli (see Menon et al., JCI Insight.
  • the subject has a level of one or more biomarkers that is altered compared to a threshold level, then the subject is to be treated with Compound 1. If the subject does not have a level of one or more biomarkers that is altered compared to a threshold level, then alternate treatment is determined. In some embodiments, in a patient to be treated with Compound 1, the kidney disease is stabilized and/or ameliorated.
  • the present disclosure encompasses a recognition that in different patients, diseasedriving pathways may have divergent outcomes or converge on a common disease related outcome. For example, in some instances, different individuals may have distinct genomic responses that all result in a common lung disease endpoint, while in other instances, different individuals may have a shared genomic response that leads to different lung disease endpoints. For example, distinct disease-driving pathways in different individuals may lead to a common outcome of a fibrotic lung disease, such as, e.g., IPF. As such, different patients may respond to different therapies and interventions for the same disease.
  • the present disclosure provides the insight that identifying and neutralizing the particular disease drivers of a given patient may provide an early and effective way to manage lung disease endpoints. Specifically, the present example determined the signalosome in various models of idiopathic pulmonary fibrosis (IPF) to identify biomarkers for treatment with Compound 1.
  • IPPF idiopathic pulmonary fibrosis
  • Two exemplary and etiologically distinct models of pulmonary fibrosis are used to characterize the lung transcriptome and proteome that are associated with therapeutic activity of Compound 1. Dosing of Compound 1 starts 2 weeks after disease induction with bleomycin or SiO 2 instillation, and dosing is continued via the oral route for 4 weeks. Lung hydroxyproline (HYP) and histopathology are determined. BAL inflammatory cell infiltrates, BAL protein, MPO, BAL cytokines, tissue cytokines, and lung fibrotic markers are evaluated by IHC staining and transcriptomic analysis by performing real-time PCR. A cytokine/chemokine/growth factor array on minimally invasive BALF samples from the animal models.
  • HEP Lung hydroxyproline
  • BAL inflammatory cell infiltrates, BAL protein, MPO, BAL cytokines, tissue cytokines, and lung fibrotic markers are evaluated by IHC staining and transcriptomic analysis by performing real-time PCR.
  • the present example describes treatment of fibrotic disease of the lung in a patient.
  • a sample of BALF is obtained from a patient with suspected or confirmed pulmonary disease (e.g., suspected of having idiopathic pulmonary fibrosis).
  • mRNA is extracted from the sample, and sequenced to quantify levels of certain biomarkers associated with Compound l’s mechanism of action.
  • expression of biomarkers identified using the method of Example 4 are quantified. If the subject has a level of a biomarker that is above a threshold level, then the subject is to be treated with Compound 1. If a subject does not have a level of biomarker above the threshold level, then alternate treatment is determined.
  • the lung disease is stabilized and/or ameliorated.
  • Example 6 A Phase 1, Randomized, Double-Blind, Placebo-controlled, Single and Multiple Ascending Dose Study to Determine the Safety, Tolerability, Pharmacokinetics, and Food Effect of Compound 1 in Healthy Adult Participants
  • Primary Endpoint The frequency and severity of treatment-emergent adverse events (TEAEs), including clinically significant abnormal vital signs, electrocardiograms (ECGs), laboratory test results, and physical examination findings.
  • TEAEs treatment-emergent adverse events
  • ECGs electrocardiograms
  • Plasma PK endpoints include:
  • Urine PK endpoints include:
  • Cohort A3 was a food effect cohort, and participants in Cohort A3 only returned to the CRU on Day 14 and, following a 14-day washout, received a second single dose of their assigned treatment on Day 15 following consumption of a high fat meal. For Cohort A3 only, a second period of confinement applied from Day 14 until completion of the 72-hour post-dose assessments on Day 18.
  • Part A participants returned to the CRU for a follow-up visit 7 days ( ⁇ 1 day) after their final dose of study drug.
  • SAD the decision to escalate a dose or modify a dose was determined by the SMC following review of the 7-day blinded safety and available PK data from the preceding cohort.
  • Compound 1 dose level tested in Part A did not exceed 1200 mg or the dose level that led to a mean whole blood C max > 600 ng/mL or mean whole blood AUC 0-last > 2000 ng*h/mL.
  • C max and AUC 0-last values were extrapolated from the mean plasma C max of 800 ng/mL and the mean plasma AUCiast of 4000 ng*h/mL observed in non-human primates at the no observed adverse effect level (NOAEL) of 75 mg/kg/day.
  • Part B Multiple Ascending Dose (MAD)
  • the Compound 1 dose level tested in Part B did not exceed 1000 mg per day or a dose level that led to a mean whole blood C max > 600 ng/mL or mean whole blood AUC 0-last > 2000 ng*h/mL.
  • C max and AUC 0-last values were extrapolated from the mean plasma C max of 800 ng/mL and the mean plasma AUCiast of 4000 ng*h/mL observed in non-human primates at the no observed adverse effect level (NOAEL) of 75 mg/kg/day.
  • Part D Single-Dose Food-Effect Cohort
  • Part D Single Dose Food Effect
  • Part B up to 32 healthy volunteers; Part C: up to 16 healthy volunteers; Part D: up to 8 healthy volunteers).
  • Participants must be in good health, with no significant medical history, have no clinically significant abnormalities on physical examination at screening and/or before administration of the initial dose of study drug. Participants must have a minimum body weight of 50 kg and a body mass index (BMI) between >18.0 and ⁇ 32.0 kg/m 2 at screening. Participants must have clinical laboratory values within normal range as specified by the testing laboratory, unless deemed not clinically significant by the investigator or delegate. Participants who smoke no more than 2 cigarettes or equivalent per week can be included in study but must be willing to abstain from smoking during the confinement period. Participants must have no relevant dietary restrictions, and be willing to consume standard meals provided during the confinement period.
  • BMI body mass index
  • Double contraception is defined as a condom and one other form of the following:
  • FSH follicle-stimulating hormone
  • Periodic abstinence e.g., calendar, ovulation, symptothermal, post-ovulationmethods
  • withdrawal are not considered highly effective methods of birth control. Participant complete abstinence for the duration of the study and for 1 months after last study treatment is acceptable. Female participants who are in same sex relationships are not required to use contraception.
  • WOCBP must have negative pregnancy test at screening and Day 1 and be willing to have additional pregnancy test as required.
  • Acceptable methods of contraception include the use of condoms and the use of an effective contraceptive for the female partner that includes: OCPs, long-acting implantable hormones, injectable hormones, a vaginal ring or an IUD. Participants with same sex partners (abstinence from penile-vaginal intercourse) are eligible when this is their preferred and usual lifestyle.
  • GI gastrointestinal
  • H. Helicobacter
  • Fever body temperature > 38 °C
  • symptomatic viral or bacterial infection within 2 weeks prior to screening. Any acute illness within 30 days prior to Day 1. History of severe allergic or anaphylactic reaction.
  • HCV hepatitis C antibody
  • HBV hepatitis B surface antigen
  • HAV human immunodeficiency virus
  • Participants with a positive toxicology screening panel urine test including qualitative identification of barbiturates, tetrahydrocannabinol (THC), amphetamines, benzodiazepines, opiates, and cocaine), or alcohol breath test.
  • a positive toxicology screening panel urine test including qualitative identification of barbiturates, tetrahydrocannabinol (THC), amphetamines, benzodiazepines, opiates, and cocaine
  • CYP3 A4 cytochrome P450 3 A4
  • any strong inhibitors or inducers of CYP3 A4 starting from 2 weeks prior to first dose of study drug and until end of study assessments.
  • Compound 1 was provided as a powder in capsule formulation for oral administration.
  • the formulation was a Size 00 Swedish orange capsule containing drug substance (50 mg or 250 mg) with no excipients.
  • Ingredients of the capsule shell were hypromellose (hydroxypropylmethyl cellulose (HPMC)), iron oxide as coloring agent, and titanium dioxide as an opacifier.
  • HPMC hydroxypropylmethyl cellulose
  • the drug product was stored at room temperature (15 °C - 25 °C).
  • Compound 1 is in a pharmacological class of tyrosine kinase inhibitors (TKI).
  • Compound 1 is an orally bioavailable small molecule dual kinase inhibitor of platelet-derived growth factor receptors (PDGFR) and vascular endothelial growth factor receptors (VEGFR2). Duration of Treatment
  • PDGFR platelet-derived growth factor receptors
  • VEGFR2 vascular endothelial growth factor receptors
  • Part A In Part A (SAD), participants randomized to active treatment in cohorts Al, A2, A4, A5, and A6 received a single oral dose of Compound 1 administered once on Day 1 only. Participants randomized to active treatment in Cohort A3 (Food Effect) received a single oral dose of Compound 1 on Days 1 and 15.
  • Part B In Part B (MAD), participants randomized to active treatment received oral Compound 1 administered twice daily for 7 consecutive days (Day 1 to Day 7) or twice daily for 14 consecutive days (Day 1 to Day 14), to be determined on completion of Part A (SAD) of the study.
  • Part C In Part C (MAD QD), participants randomized to active treatment received oral Compound 1 administered once daily for 14 consecutive days (Day 1 to Day 14).
  • the placebo capsules visually matched to active study drug and were composed of the same capsule shell but were filled with silicified microcrystalline cellulose.
  • Silicified microcrystalline cellulose is a pharmaceutical excipient composed of co-process microcrystalline cellulose and colloidal silicon dioxide.
  • Safety The safety and tolerability of single and repeat-doses of Compound 1 was investigated according to the following specific assessments: vital signs (systolic and diastolic blood pressure, pulse rate, body temperature, and respiratory rate), 12-lead ECG, clinical laboratory tests (hematology, biochemistry, coagulation, and urinalysis), physical examination, and assessment of TEAEs.
  • ITT Intent-to-Treat
  • Safety Population All participants who received any amount of study drug (Compound 1 or placebo) were included in the Safety population. The Safety population was used for the summaries of all safety assessments. Participants were analyzed according to treatment received.
  • Pharmacokinetic Population All participants who received any amount of active study drug (Compound 1) and have sufficiently evaluable concentration-time profile to allow determination of at least one PK parameter were included in the PK population. An evaluable PK profile was determined at the discretion of the pharmacokineticist following examination of subjects with dosing or protocol deviations that could potentially affect the PK profile. The PK population was used for the summaries of all PK data.
  • Concomitant medications were coded using the World Health Organization (WHO) drug dictionary Version B3 September 2018 Drug Global. Concomitant medications were listed by participant and summarized by anatomical therapeutic class and preferred name.
  • WHO World Health Organization
  • Plasma Compound 1 concentrations, actual blood sampling times, and PK parameters were listed by treatment and protocol specified time point and summarized using descriptive statistics for PK data as outlined above for each scheduled time point by treatment arm. Individual and mean Compound 1 concentration-time profiles were also presented graphically for each treatment. Pharmacokinetic parameters were computed from the individual plasma Compound 1 concentrations using a non-compartmental approach.
  • Urine collection time, volume collected, and Compound 1 concentration (Aeti-t2) duration of each sampling interval was listed for each participant and summarized by nominal sampling time point and treatment using descriptive statistics (Number of subjects [N], arithmetic mean, SD, CV%, geometric mean, median, minimum and maximum). Individual and mean Compound 1 cumulative urinary excretion-time profiles for each treatment were also presented graphically. Where urine was collected for PK analysis, the following parameters were calculated, as appropriate: Aeti-t2, CLR, CL/F, and Feti-t2.
  • Example 7 Compound 1 Ameliorates Fibrosis in Bleomycin-induced Injury Model of Idiopathic Pulmonary Fibrosis
  • mice Male C57BL/6 mice ( ⁇ 20 to 25 g) were anaesthetized for a short period of time using ketamine (100 mg/kg) and xylazine (10 mg/kg).
  • bleomycin mice Two weeks after bleomycin instillation, a subgroup of animals was sacrificed and the establishment of lung fibrosis was confirmed (initially as determined by increased lung weight).
  • H&E haematoxylin-eosin
  • FIG. 3 A is a graph showing lung to body weight ratio (mg/g).
  • Compound 1 also significantly reduced fibrosis score (Ashcroft) on histopathology. Lung tissue sections were stained with H&E, and slides were scored for histopathological damage using the Ashcroft scale. Two weeks after bleomycin exposure, animals treated with bleomycin had increased histopathological damage compared to sham-treated animals, indicative of development of pulmonary fibrosis. Animals treated with Compound 1 for three weeks had reduced histopathological damage compared to vehicle-treated animals (FIG. 3C).
  • TGF ⁇ 1 transforming growth factor beta
  • Example 8 Compound 1 Reduces Fibrosis in Inducible TGF ⁇ 1 Mouse Model of Lung Fibrosis
  • mice which expresses TGF ⁇ 1 in the lung via an externally regulatable, triple transgenic system using a doxycycline-inducible promoter.
  • a doxycycline-inducible promoter See Lee, C. G., et al. Proc. Am. Thorac. Soc. 2006 Jul;3(5):418-23; Lee C. G., et al. J. Exp. Med. 2004 Aug 2;200(3)377-89.
  • Eight-to-ten week old TGF ⁇ 1 positive female mice (approximately 25 g of body weight) were induced (fed) with doxycycline (dox) (0.5 mg/ml dox and 20 g/L sucrose in drinking water) for 4 weeks.
  • dox doxycycline
  • the TGF ⁇ 1 transgene was under the control of a dox-inducible promoter.
  • age and gender matched TGF ⁇ 1 negative mice from the transgenic mouse breeding colony not fed with dox were included as control mice (sham).
  • Marked loss of normal bronchial tree architecture due to TGF ⁇ 1 overexpression was evidenced after 4 weeks of dox compared to sham mice using micro-CT images.
  • mice not fed with dox were sacrificed to confirm that pathological changes had happened (increases in lung weight) in dox fed mice.
  • lung hydroxyproline (HYP) for collagen content, picrosirius red (PSR) staining for collagen signal, and the histopathological observations from (H&E) slides by Ashcroft scoring and IHC staining for alpha SMA were performed. All histological quantitations were performed using a Bioquant image analysis program.
  • Compound 1 reduced lung fibrosis score (Ashcroft) in dox-fed TGF ⁇ 1 transgenic mice.
  • Dox feeding of TGF ⁇ 1 transgenic mice for 4 weeks resulted in a significant increase in lung fibrotic score (Ashcroft score based on H&E staining), compared to normal mice (FIG. 4A).
  • Further dox feeding for an additional 4 weeks resulted in more increases in lung fibrotic score, as shown by the vehicle group.
  • a reduction in hydroxyproline, a component of collagen is indicative of a reduction in fibrosis.
  • aSMA is a marker of myofibroblast activation, an early step in fibrosis development.
  • Example 9 Compound 1 Has Anti-Fibrotic Effects Across Multiple Organ Systems in Bleomycin Systemic Sclerosis Mouse Model
  • SC subcutaneous
  • the right lung and right kidney were also fixed in formalin for histopathological evaluation.
  • Dermal thickness measurements were made from H&E-stained tissue sections by measuring the distance from the epidermis to the dermal junction using Bioquant planimetric software.
  • Lung and kidney hydroxyproline (HYP) assays were performed to determine tissue collagen content. Skin, lung and kidney histopathological fibrotic scores were determined by two independent observers, and the scores were averaged.
  • Picrosirius red (PSR) staining of kidney sections was also performed to determine renal collagen deposition. A subset of animals were randomly chosen from each group for histological staining. All histological quantitations were performed using Bioquant image analysis software. [0275] Scleroderma is characterized by thickening of the skin caused by accumulation of collagen.
  • dermal thickness was determined by measuring the distance between the epidermal-dermal junction and the dermal-subcutaneous fat junction. This was done in H&E-stained slides prepared from a skin biopsy. After 4 weeks of subcutaneous bleomycin injections (Bleo Pre-Rx Group), dermal thickness was significantly increased compared to sham-injected animals (Sham Group). This indicates significant skin thickening and scleroderma at the time of initiation of compound treatment.
  • Therapeutic treatment of Compound 1 for 5 weeks 25 mg/kg, PO, BID) (Bleo Compound 1 Group) significantly decreased dermal thickness compared to the bleomycin- exposed vehicle cohort (Bleo Vehicle Group).
  • FIG. 5A summarizes these results.
  • Hydroxyproline is a major component of collagen, and it is used to indicate tissue collagen deposition and the development of fibrosis.
  • pulmonary hydroxyproline levels were determined. After 4 weeks of subcutaneous bleomycin injections, lung hydroxyproline levels were significantly increased compared to sham treated animal (Sham vs Bleo PreRx Groups), indicating the development of pulmonary fibrosis.
  • Treatment of bleomycin exposed animals with Compound 1 for 5 weeks (25 mg/kg, PO, BID) (Bleo Compound 1 Group) significantly decreased lung hydroxyproline levels compared to bleomycin exposed vehicle treated animals (Bleo Vehicle), indicating anti-fibrotic activity of Compound 1.
  • FIG. 5C summarizes these results.
  • Example 10 Compound 1 Reduces Proteinuria and Sclerosis in FSGS-Relevant Rat Model of PAN-induced Proteinuria
  • Urine was again collected (for 24 hours) from day 10-11 after first PAN injection (referred to as Day 11 urines).
  • Day 11 urines On day 11 after first PAN administration GFR was measured by computing FITC-sinistrin decay (Medibeacon) and animals were sacrificed. The left kidney was cut coronally, immersed in formalin (4%) and submitted for sectioning and mounting on slides for subsequent analysis (PAS stain for determining glomerular diameter) and anti-COL 3 antibody H4C. All microscopic analyses (40X) were conducted by an observer blinded to the treatment groups. Glomerular diameter was determined from digital microscopic pictures using ImageJ image analysis software to perform planimetry. Proteinuria was determined from urine samples.
  • every endpoint was not measurable in each animal within a group due to logistical limitations. In such cases, randomly selected animals were queried for data. For instance, no urine was collected for certain animals due to the limited number of metabolic cages available, and only a few animals in each cohort were randomly selected for GFR measurement due to the limited number of probes available for this measurement.
  • the highly sensitive FITC-sinistrin clearance method was employed in conscious animals. This method is capable of measuring the elimination kinetics of FITC-sinistrin using an optical device (Medibeacon, Germany) that can monitor fluorescence trans-cutaneously. FITC-sinistrin clearance is a readout of glomerular filtration rate. The optical device measures fluorescence emitted by FITC-sinistrin through the skin and was temporarily secured on the back of the animal. A single dose of FITC-sinistrin (5 mg/100 g body weight FITC-sinistrin) was injected into the tail vein and the measurement with the optical device was performed for 1.5 hr.
  • Collagen 3 (COL-3) expression a measure of scar, was determined in glomeruli by quantitative immuno-histochemistry.
  • the COL-3 expression level was markedly elevated in glomeruli in the kidneys from PAN Vehicle animals compared to Sham treated animals (FIG. 6E).
  • the COL-3 level was significantly reduced with Compound 1 treatment (FIG. 6E) compared to PAN Vehicle treatment.
  • Example 11 Compound 1 Reduces Proteinuria and Fibrosis in DOCA/Salt Model of Renal Injury and Fibrosis
  • Induction of renal dysfunction was started on day 1, at which point experimental animals received a first subcutaneous dose of deoxycorticosterone acetate (DOCA) at 30 mg/kg and animals were switched from normal drinking water (tap water) to drinking water with 1% NaCl. Animals were kept on the 1% NaCl drinking water for the remainder of the experiment. Animals received weekly subcutaneous doses of DOCA; 30 mg/kg in weeks 1, 2, 3 and 4 and 15 mg/kg in weeks 5 and 6. Control groups were subcutaneously injected with PEG400 and received regular tap water ad libitum. After two weeks, overnight urine was collected in metabolic cages and a subset of animals was sacrificed to establish baseline values.
  • DOCA deoxycorticosterone acetate
  • the time of body weight measurement was at week 2 for Groups 1 and 2 and at week 6 for Groups 3, 4, and 5. All animals that were treated with DOCA + Salt had significantly reduced body weight compared to control animals that did not receive DOCA and were given regular tap water to drink.
  • the body weight in the DOCA + Salt + Compound 1 treated group (Group 5) was not significantly different from animals in the DOCA + Salt + Vehicle (Group 4), indicating that there was no obvious deleterious effect of compound treatment on body weight.
  • kidneys were collected and their weights recorded.
  • the kidney weights from animals of DOCA + Vehicle group (Group 4) were significantly more than the kidney weights in Sham animals (Group 3).
  • the kidney weight in DOCA + Compound 1 group (Group 5) was not significantly different from the DOCA + Vehicle group (Group 4).
  • the weight of the kidney was also expressed as a percentage of the total body weight of each individual animal.
  • the % kidney weight from animals of DOCA + Vehicle group (Group 4) was significantly higher than the % kidney weights in Sham animals (Group 3).
  • Compound 1 increased body weight and decreased kidney weight compared to vehicle, albeit not statistically significant.
  • the % kidney weight (taken both body weight and kidney weight into consideration) in DOCA + Compound 1 group (Group 5) was significantly reduced compared to the DOCA + Vehicle group (Group 4).
  • Urines were collected from animals pre-randomization and immediately prior to study end. Animals from the DOCA + 1% NaCl in the drinking water groups (Groups 2, 4 and 5) showed markedly increased urine volumes compared to Sham control animals (Groups 1 and 3). Animals from the DOCA + Compound 1 group (Group 5) showed statistically reduced urine output compared to DOCA + Vehicle group (Group 4).
  • Urines were analyzed for protein content and the total amount of protein produced over 24 hours was calculated from the protein concentration and the urine volume produced in 24 hours (FIG. 7A).
  • a statistically significant elevation in proteinuria was observed in DOCA + Salt animals (Group 2) compared to Sham animals (Group 1), indicating renal dysfunction in the DOCA + Salt animals at the start of compound or vehicle treatment.
  • a statistically significant elevation in proteinuria was again observed in DOCA + Vehicle animals (Group 4) compared to Sham animals (Group 3), indicative of continued renal dysfunction.
  • DOCA-exposed animals treated with Compound 1 (Group 5) were found to have statistically significant reduction in proteinuria compared to DOCA + Vehicle treated animals (Group 4). This is consistent with an improvement in renal function by Compound 1 treatment.
  • Urines were analyzed for albumin concentration and the total amount of albumin produced over 24 hours was calculated from the albumin concentration and the urine volume produced in 24 hours (FIG. 7B).
  • elevated albuminuria was observed in DOCA + Salt animals (Group 2) compared to Sham animals (Group 1), albeit that this effect was not statistically significant.
  • a statistically significant elevation in albuminuria was observed in DOCA + Vehicle animals (Group 4) compared to Sham animals (Group 3), indicative of renal dysfunction.
  • DOCA-exposed animals treated with Compound 1 (Group 5) were found to have statistically significant reduction in albuminuria compared to DOCA + Vehicle treated animals (Group 4). This is consistent with the proteinuria data and with an improvement in renal function by Compound 1 treatment.
  • Urines were also analyzed for creatinine levels and the albumin-to-creatinine-ratio (ACR) was calculated (FIG. 7C). Data for ACR were similar to the overall albumin production data. As before, DOCA-exposed animals treated with Compound 1 (Group 5) were found to have statistically significant reduction in ACR compared to DOCA + Vehicle treated animals (Group 4). This is consistent with the proteinuria data and the albuminuria data, and with an improvement in renal function by Compound 1 treatment.
  • Kidney Injury Molecule- 1 (also known as Hepatitis A virus cellular receptor 1 (HAVcr-1) or T-cell immunoglobulin and mucin domain 1 (TIM1)) is a protein that is highly upregulated in injured kidneys by various types of insults (Bonventre, J. V. Trans. Am. Clin. Climatol. Assoc. 2014;125:293-9). In patients with Minimal Change Disease or Focal Segmental Glomerulosclerosis, elevated KIMI levels are found in the urine (Zhang, Q., et al. Am. J. Med. Sci. 2018 Apr;355(4):314-21).
  • Urines were analyzed for KIMI and the total amount of KIMI produced over 24 hours was calculated (FIG. 7D).
  • elevated KIMI production was observed in DOCA + Salt animals (Group 2) compared to Sham animals (Group 1), indicating renal injury at the start of compound or vehicle treatment.
  • a statistically significant elevation in KIMI production was observed in DOCA + Vehicle animals (Group 4) compared to Sham animals (Group 3), indicative of continued renal injury.
  • DOCA-exposed animals treated with Compound 1 (Group 5) were found to have statistically significant reduction in urinary KIMI production compared to DOCA + Vehicle treated animals (Group 4). This is consistent with the notion that Compound 1 treatment reduces renal damage.
  • Hydroxyproline is a major component of collagen, and tissue hydroxyproline levels are considered a good indicator of tissue collagen content and therefore of tissue fibrosis.
  • the hydroxyproline content of renal samples was determined and expressed as the total hydroxyproline content per kidney (FIG. 7E).
  • kidneys from pre-randomization DOCA + Salt treated animals (Group 2) no elevation in renal hydroxyproline content was observed, compared to pre-randomization Sham animals (Group 1). This indicates absence of renal fibrosis at the start of compound or vehicle treatment.
  • a statistically significant elevation in renal hydroxyproline content was seen in DOCA + Vehicle animals (Group 4) compared to Sham animals (Group 3), consistent with marked renal fibrosis at study end.
  • DOCA-exposed animals treated with Compound 1 were found to have statistically significant reduction in renal hydroxyproline content compared to DOCA + Vehicle treated animals (Group 4). This indicates that Compound 1 treatment prevents the development of renal fibrosis in DOCA + Salt treated animals.
  • Picrosirius red-staining in the slides was quantified using Bioquant Image Analysis software. The data were expressed as arbitrary units of staining intensity and also expressed as a % of control in relation to the average staining intensity of the control group (Group 1). As shown in FIG. 7G, in pre-randomization animals, no increase in picrosirius red staining intensity was observed in DOCA + Salt animals (Group 2) compared to Sham control animals (Group 1). This indicates lack of renal fibrosis at the time of compound of vehicle treatment initiation. This lack of histologically observable fibrosis in the renal samples from Group 2 is consistent with the hydroxyproline data in this group.
  • DOCA + Vehicle treated animals (Group 4) had markedly increased insterstitial picrosirius red staining compared to Sham control animals (Group 3). This indicates marked collagen deposition and renal fibrosis (tubulointerstitial fibrosis) as a result of DOCA + Salt treatment.
  • animals that received DOCA + Salt that were treated with Compound 1 (Group 5) showed statistically significantly reduced picrosirius red staining intensity compared to DOCA + Vehicle animals (Group 4). This is consistent with the observed differences in renal hydroxyproline in the various experimental groups and shows anti-fibrotic effects of Compound 1 in this model.
  • Example 12 Unilateral Ureteral Obstruction (UUO) Model of Renal Fibrosis in Mice [0311] Male C57BL/6 mice were anesthetized and placed on a heated surgical pad to maintain body temperature between 37 °C and 37.5 °C throughout the surgical procedure and recovery. The left ureter was visualized via a flank incision and ligated with a 6-0 suture. Starting the day after UUO, animals were randomized to groups receiving vehicle or Compound 1.
  • Table 11 a 100 mg/kg Compound 1 Hydrochloride trihydrate.
  • the weight of the obstructed kidney was also expressed as a percentage of the body weight of each individual animal.
  • the percentage kidney weight for animals from the UUO + Vehicle group (Group 2) was elevated compared to Control animals (Group 1). There was no statistically significant difference between the UUO + Compound 1 group (Group 3) and the other experimental groups.
  • kidneys were fixed in 10% neutral buffered formalin for at least 48 hours before preparation for histology. Kidneys tissues were grossed, processed, dehydrated, embedded in paraffin and cut into 5 pm sections.
  • Compound 1 was evaluated in the Unilateral Ureteral Obstruction (UUO) model of renal fibrosis in the mouse.
  • Primary endpoints for evaluating compound effects were renal damage score, collagen deposition (as determined by picrosirius red staining) and formation of myofibroblasts (as determined by a-smooth muscle actin staining).
  • Compound 1 had statistically significant beneficial effects on renal damage, picrosirius red staining and alpha-SMA staining.
  • the PCK rat which has a mutation in PKHD1 (ARPKD gene) phenotypically resembles both human ARPKD-CHF and human PKD in that it exhibits large renal cysts, and renal and hepatic scarring.
  • the PCK rat model is a well-established and well-characterized model that resembles human polycystic kidney and liver disease (Lager, D. J., et al. Kidney Int. 2001 Jan;59(l): 126-36). Results described in this Example are also described in Paka, P., et al. World J. Nephrol. 2018 Sept 8;7(5):96-107).
  • WT wild-type SD rat
  • animals were anesthetized with isoflurane (2%), a midline incision was made and the left kidney was removed. The left kidney was removed to accelerate disease progression.
  • PCK rat kidney was enlarged and exhibited numerous cysts compared the kidney from the wild-type (WT) Sprague Dawley animal.
  • PCK animals were randomized to Compound 1 and vehicle treatment groups and animals were treated for 8 weeks, before collection of 24-hour urine and sacrifice of animals at 14 weeks of age.
  • PCK rats that were treated with either vehicle or with Compound 1 did not differ in their body weights from age and sex matched wild-type (WT) Sprague Dawley rats.
  • SCr Serum creatinine
  • BUN BUN levels were determined in sera of 14-week old animals (FIG. 9A and FIG. 9B, respectively). Both the SCr and BUN levels were elevated in PCK Vehicle rats compared to wild-type (WT) rats, indicating mild, but statistically significant level of renal dysfunction in the PCK animals.
  • Compound 1 treatment for 8 weeks reduced both SCr and BUN levels compared to PCK Vehicle animals, indicating improvement in renal function as a result of Compound 1 treatment.
  • Kidneys were weighed at sacrifice and kidney weights are given in absolute weight (g) and as a percentage of body weight (FIG. 9C and FIG. 9D, respectively). Kidneys from PCK Vehicle rats were markedly larger than kidneys from wild-type (WT) rats. Compound 1 treatment in PCK rats reduced kidney size and the % kidney weight compared to PCK Vehicle animals.
  • Hydroxyproline is a major component of collagen, and it is used to indicate tissue collagen deposition and fibrosis. Hydroxyproline levels were determined in kidney samples of 14-week old animals (FIG. 9E). Kidney hydroxyproline levels were markedly increased in PCK Vehicle rats compared to wild-type (WT) rats. Compound 1 treatment significantly reduced renal hydroxyproline levels compared to PCK Vehicle animals, indicating reduced renal collagen deposition as a result of Compound 1 treatment. [0329] The percentage of the renal parenchyma occupied by cysts (cystic index) was quantified in H&E-stained kidney sections using digital planimetry.
  • Increased urine protein excretion can be due to diseases of the glomerulus, an increased quantity of proteins in serum (overflow proteinuria) or low reabsorption at proximal tubule.
  • Urine protein excretion was determined in urines collected over 24 hours immediately prior to sacrifice (FIG. 9H). Protein excretion was found to be elevated in PCK Vehicle rats compared to wild-type (WT) rats, and Compound 1 treatment markedly reduced proteinuria in PCK rats compared to PCK Vehicle animals.
  • Albumin is a major plasma protein. Usually, only trace amounts of it are present in urine, whereas larger amounts occur in the urine of patients with kidney disease. Urine albumin concentrations were determined in urines collected over 24 hours immediately prior to sacrifice (FIG. 91). Urine albumin production was found to be strongly elevated in PCK Vehicle rats compared to wild-type (WT) rats, and Compound 1 treatment markedly reduced urine albumin production in PCK rats compared to Vehicle treated PCK animals.
  • Lipocalin-2 (LCN2) is also known as neutrophil gelatinase-associated lipocalin (NGAL) and is a protein that is expressed in neutrophils and at low levels in normal kidneys (Cowland, J. B., et al. Genomics 1997 Oct 1 ;45(1): 17-23).
  • NGAL neutrophil gelatinase-associated lipocalin
  • Urine NGAL excretion was determined for 14-week old animals (FIG. 9J).
  • NGAL excretion was found to be markedly elevated in PCK Vehicle rats compared to wild-type (WT) rats, albeit that the increase was not statistically significant by one way ANOVA followed by Tukey’s test. This is probably the result of high variability in NGAL excretion amongst the PCK Vehicle animals.
  • Compound 1 treatment reduced NGAL excretion in PCK rats compared to Vehicle treated animals, albeit that this decrease was also not statistically significant when tested by ANOVA followed by Tukey’s test.
  • HAVcr-1 also known as KIM-1 (KIMI, kidney injury molecule 1
  • Interleukin 18 (IL- 18 or IL 18) is also known as interferon-gamma inducing factor and is a pro-inflammatory cytokine.
  • Urine levels of IL- 18 are associated with urine protein excretion and have been used as a biomarker for the progression of diabetic nephropathy (Zhang, D., et al., PeerJ. 2019;7:e7079; Nakamura, A., et al. Diabetes Care 2005 Dec;28(12):2890-5). Urine IL-18 excretion was determined for 14-week old WT and PCK animals (FIG. 9L).
  • IL-18 excretion was found to be elevated in PCK Vehicle rats compared to wild-type (WT) rats, and Compound 1 treatment markedly reduced IL-18 excretion in PCK rats compared to Vehicle treated animals.
  • Cystatin C or cystatin 3 is used as a biomarker of kidney function (Dharnidharka, V. R., et al. Am. J. Kidney Dis. 2002 Aug;40(2):221-6; Roos, J. F., et al. Clin. Biochem. 2007 Mar;40(5-6):383-91).
  • Urine Cystatin C excretion was determined for 14-week old animals (FIG. 9M).
  • Cystatin C excretion was found to be elevated in PCK Vehicle rats compared to wild-type (WT) rats, and Compound 1 treatment markedly reduced Cystatin C excretion in PCK rats compared to Vehicle treated animals. These results are indicative of an improvement of renal function, consistent with the observation that Compound 1 reduced serum BUN and SCr levels in PCK rats compared to PCK Vehicle treated animals.
  • Example 14 Model of Inflammatory Bowel Disease (IBD)/Acute Colitis Induced by TNBS in Mice
  • TNBS (2,4,6-trinitrobenzenesulfonic acid)/ethanol induces a severe illness characterized by bloody diarrhea and a dramatic loss of body weight during the first week with increased colon damage score
  • TNBS-induced colitis characteristic of acute colitis and the early phase of IBD, is a commonly utilized animal model in rodents that shares significant properties with human Crohn's disease. Advantages of this model include reproducibility and technical simplicity (Filipescu, I. E., et al. PLoS One 2018 Aug 23;13(8):e0202929).
  • TNBS-induced colitis is a well-validated animal model to assess compounds with potential therapeutic effects such as anti-TNF ⁇ , corticosteroids, natural compounds and traditional medicine (Mateus 2018; Filipescu 2018; Wirtz, S. et al. Nat. Protoc. 2017 Jul; 12(7): 1295-1309). Compound 1 was therefore tested in the well-established model of TNBS-induced IBD/ Acute Colitis.
  • mice were randomized to vehicle and Compound 1 at 5, 15 and 45 mg/kg Compound 1 Hydrochloride trihydrate, PO, BID for 4 days. After 4 days of Compound 1 or vehicle treatment, mice were sacrificed. Body weight, colon length and colon weight were recorded. The middle and distal part of colon were fixed in 10% neutral buffered formalin.
  • Colon histopathology was determined from H&E-stained tissue sections by two independent observers using a previously published scoring system.
  • the score for each individual component (0-3) of the Histopathological Score is shown in FIG. 10E and the composite (total) Colon Histopathological Score (the sum of individual component scores; 0-12) is shown in FIG. 10F. Histopathological Scores showed that Compound 1 treatment at 15 and 45 mg/kg markedly improved colon histology compared to pre-treatment and vehicle treated animals.
  • Colon tissue sections were stained with Alcian blue to evaluate crypt architecture/goblet cells. The degree of Alcian blue staining was quantified to determine crypt damage and goblet cell loss (FIG. 10G, group averages shown). Compound 1 treatment at 15 and 45 mg/kg significantly reduced TNBS-induced goblet cell loss compared to pre-treatment and vehicle treated animals.
  • MPO Myeloperoxidase
  • IBD inflammatory bowel disease
  • Example 15 Compound 1 Decreases Acetic Acid (AA) Induced Colitis in Mice
  • Example 16 Compound 1 Decreases Chronic Colitis in IBD Mouse Model
  • Efficacy of Compound 1 in a dextran sulfate sodium (DSS) induced chronic colitis/IBD model was evaluated.
  • Adult male CD-I mice were fed with 3% DSS in drinking water for 5 days with alternate cycles of normal drinking water for 5 days and continued a total of 3 DSS cycles in 4 weeks. This model has been established to induce chronic colitis/IBD.
  • DSS-Vehicle cohort had the shortest colon length; DSS-Compound 1 cohort had significantly increased colon length compared to the DSS-vehicle cohort (FIG. 12A).
  • Compound 1 treatment decreased macroscopic colon damage score in terms of appearance, inflammation and rectal bleeding (FIG. 12B), decreased colonic hydroxyproline (FIG. 12C) and histopathological injury score based on crypt damage, epithelial erosion and inflammatory cell infiltration as seen in H&E stained colon sections (FIG. 12D). These results indicate that Compound 1 could prevent progression of inflammation-induced fibrosis.
  • Example 17 Identification of Col6 as a Serum Biomarker for Treatment of Fibrotic Kidney Diseases with Compound 1
  • Compound 1 was evaluated in three rodent models of renal dysfunction: PANX model, DOCA/salt model, and PHN model.
  • PHN model 65 CD® rats from Charles River Laboratory were obtained and acclimated. 60 of them were administered anti-FXl A serum (tail vein, IV, 600 pL), and 5 were administered saline (tail vein, IV, 600 pL, sham), on two consecutive days, when the average mass of the animals was 300 g. Twenty-four hour urine samples were collected 7 days later (on Day 9) and proteinuria was determined. Animals were randomized based on proteinuria levels, ensuring equivalent average protein to creatinine ratio (PCR) and standard deviation in each group. Experimental groups were (1) 15 mg/kg Compound 1, (2) Vehicle (water), and (3) Sham. Dosing of animals in the Compound 1 and Vehicle groups began on day 10 and continued 5 days a week for 12 weeks (PO, BID). At the end of the study (day 93), animals were sacrificed, and kidney tissue and blood samples were collected.
  • Serum samples obtained from the models were analyzed for pro-collagen 6 levels using quantitative competitive ELISA.
  • FIG. 13 A is a graph showing serum pro-collagen 6 levels in animals treated with Compound 1 compared to sham and control animals.
  • FIG. 13B is a graph showing a correlation between serum pro-collagen 6 and TGF-P levels in animals evaluated in this Example.
  • FIG. 13C is a graph showing serum pro-collagen 6 levels in animals treated with Compound 1 compared to sham and control animals in the PANX model.
  • FIG. 13D is graph showing serum pro-collagen 6 levels in animals treated with Compound 1 compared to sham and control animals in the DOC A model.

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Abstract

La présente invention concerne des biomarqueurs et des procédés qui sont utiles dans le traitement, le criblage et/ou l'évaluation du traitement de maladies fibrotiques avec le méthyl (Z)-3-(((4-(N-méthyl-2-(4-méthylpipérazin-1-yl)acétamido)phényl)amino)(phényl)méthylène)-2-oxo-2,3-dihydro-1H-pyrrolo[2,3-b]pyridine-6-carboxylate.
PCT/US2022/014185 2021-01-29 2022-01-28 Méthodes de traitement de maladies fibrotiques WO2022165092A1 (fr)

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Citations (5)

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Publication number Priority date Publication date Assignee Title
US20040038856A1 (en) * 2002-05-17 2004-02-26 Sarvajit Chakravarty Treatment of fibroproliferative disorders using TGF-beta inhibitors
US20120135882A1 (en) * 2009-05-14 2012-05-31 Mount Sinai School Of Medicine Methods for diagnosing chronic kidney disease and assessing the risk of disease progression
US20140371234A1 (en) * 2012-01-26 2014-12-18 Angion Biomedica Corp. Antifibrotic compounds and uses thereof
US9181547B2 (en) * 2011-04-25 2015-11-10 Regulus Therapeutics Inc. MicroRNA compounds and methods for modulating MIR-21 activity
US20200002424A1 (en) * 2015-11-23 2020-01-02 Merck Patent Gmbh Anti-alpha-v integrin antibody for the treatment of fibrosis and/or fibrotic disorders

Patent Citations (5)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US20040038856A1 (en) * 2002-05-17 2004-02-26 Sarvajit Chakravarty Treatment of fibroproliferative disorders using TGF-beta inhibitors
US20120135882A1 (en) * 2009-05-14 2012-05-31 Mount Sinai School Of Medicine Methods for diagnosing chronic kidney disease and assessing the risk of disease progression
US9181547B2 (en) * 2011-04-25 2015-11-10 Regulus Therapeutics Inc. MicroRNA compounds and methods for modulating MIR-21 activity
US20140371234A1 (en) * 2012-01-26 2014-12-18 Angion Biomedica Corp. Antifibrotic compounds and uses thereof
US20200002424A1 (en) * 2015-11-23 2020-01-02 Merck Patent Gmbh Anti-alpha-v integrin antibody for the treatment of fibrosis and/or fibrotic disorders

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