WO2021054978A1 - Cardiac cardenolides to reduce fibrosis and enhance epithelial differentiation - Google Patents
Cardiac cardenolides to reduce fibrosis and enhance epithelial differentiation Download PDFInfo
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K31/00—Medicinal preparations containing organic active ingredients
- A61K31/56—Compounds containing cyclopenta[a]hydrophenanthrene ring systems; Derivatives thereof, e.g. steroids
- A61K31/58—Compounds containing cyclopenta[a]hydrophenanthrene ring systems; Derivatives thereof, e.g. steroids containing heterocyclic rings, e.g. danazol, stanozolol, pancuronium or digitogenin
- A61K31/585—Compounds containing cyclopenta[a]hydrophenanthrene ring systems; Derivatives thereof, e.g. steroids containing heterocyclic rings, e.g. danazol, stanozolol, pancuronium or digitogenin containing lactone rings, e.g. oxandrolone, bufalin
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K31/00—Medicinal preparations containing organic active ingredients
- A61K31/70—Carbohydrates; Sugars; Derivatives thereof
- A61K31/7042—Compounds having saccharide radicals and heterocyclic rings
- A61K31/7048—Compounds having saccharide radicals and heterocyclic rings having oxygen as a ring hetero atom, e.g. leucoglucosan, hesperidin, erythromycin, nystatin, digitoxin or digoxin
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- the present invention relates to the use of cardiac cardenolide compounds for the purposes of reducing fibrosis and enhancing epithelial differentiation BACKGROUND OF THE INVENTION 1.0.
- Introduction Injury in many tissues is followed by self-limiting fibrosis, which is followed by epithelial differentiation to repair functional damage.
- An example would be a scar, However, many disease entities are characterized by pathogenic fibrosis in which the process continues and does not stop before creating more damage. Furthermore, epithelial differentiation for detailed repair does not follow. Drugs to arrest fibrosis are thus considered necessary as therapeutics.
- differentiated epithelial cells line the outside and inside cavities and lumina within the body. For example, these include the mouth, esophagus, small and large intestine, lungs, GI tract (including liver and pancreas), the reproductive and urinary tracts, and the exocrine and endocrine glands, sweat glands, mammary glands and salivary glands, and include all organs whose discharges exit to outside the body.
- drugs that promote epithelial differentiation following injury would be considered advantageous therapeutic entities.
- TGFbeta Transforming Growth Factor beta
- Tissues and disease entities so affected include, but are not limited to, the kidney [2], including diabetic nephropathy [3]; gastrointestinal tract [4], including colitis/Crohn's Disease [5]; radiation-induced fibrosis [6]; salivary gland fibrosis [7]; heart, including /myocarditis [8]; rheumatoid arthritis [9]; liver , including hepatic fibrosis [10]; skeletal muscle fibrosis [11]; pancreatic fibrosis [12]; systemic scleroderma/systemic sclerosis [13]; and lung diseases, including chronic obstructive pulmonary disease (COPD), severe forms of asthma, and idiopathic pulmonary fibrosis (IPF), and cystic fibrosis (CF) [14].
- COPD chronic obstructive pulmonary disease
- IPF idiopathic pulmonary fibrosis
- CF cystic fibrosis
- TGFbeta The cellular source of TGFbeta in the normal human lung is believed to be the epithelial cells lining the airways [15].
- Experimental and clinical suppression of TGFbeta signaling in some of these cases has apparently been accompanied by beneficial consequences.
- At least two small molecules, pirfenidone [16, 17] and losartan [18] have been approved to target this pathway.
- both drugs contribute to enhanced progression-free survival in some of these diseases, neither are able to support a durable suppression of the disease process.
- Side effects are also a problem, and thus development of anti-fibrotic drugs targeting TGFbeta remain a life-limiting, unmet need. 3.0. Reducing TGFbeta expression reduces fibrosis in model systems.
- TGFbeta Reducing TGFbeta has been shown to be protective in model systems [19, 20]. For example, attempts have been made to reduce circulating TGFbeta levels with a TGFBR2-based ligand trap [21]. In another example, knockout of Tgfbr2 in mice confers increased survival and resistance to bleomycin-induced pulmonary fibrosis [19]. So far, however, there has been no practical success [22]. Currently approved therapy for bronchiectasis includes inhaled glucocorticoids [23]. A classical example is beclomethasone propionate [24]. However, side effects from this therapeutic class have usually precluded chronic use.
- Elevated KRT8 is expressed in cells undergoing epithelial differentiation for repair and reconstitution of damaged epithelial tissues.
- Epithelial differentiation from basal stem cells known to be marked by Cytokeratin 8 (KRT8), is the mechanism for repair and reconstitution of normal epithelial morphology of damaged epithelial tissues.
- Organs expressing intrinsically high levels of KRT8 mRNA and protein include most endocrine tissues, bone marrow and immune system, lung, liver and gall bladder, pancreas, gastrointestinal tract, kidney and urinary bladder.
- KRT8 high levels of KRT8 are found in male genital tissues (for example, testes, prostate, epididymis, seminal vesicles), and in female genital tissues (for example, fallopian tubes, breast, vagina, uterine cervix, endometrium, ovary and placenta). Details for KRT8 protein distribution are found in The Human Protein Atlas (protein atlas.erg). In the lung, for example, repair and reconstitution follows the embryological pathway, emphasizing reduction in TGFbeta and elevation in signals such as KRT8, which mark the conversion of basal stem cells to pseudostratified epithelial cells [25].
- KRT8-marked precursor cells In the lung, Notch signaling drives basal stem cells to form KRT8-marked intermediate basal luminal precursor cells [26]. KRT8-marked precursor cells then differentiate into all other lung epithelial cells. In the proximal lung the latter include club, ciliated, neuroendocrine cells, and goblet cells. In the distal lung these KRT8- marked precursor cells differentiate into the alveolar Type I and Type II cells, which mediate oxygen/CO2 exchange [25]. Thus KRT8 denotes cells associated with highly differentiated states in multiple epithelial tissues. Therefore drugs that elevate KRT8, and thus represent the process of epithelial differentiation, might have beneficial therapeutic value.
- the present invention relates to the use of cardiac cardenolide compounds for the purposes of reducing fibrosis and enhancing epithelial differentiation.
- BRIEF DESCRIPTION OF THE DRAWINGS Figure 1. Examples of cardiac cardenolide drugs
- Figure 3. Digitoxin can be used as an adjuvant with other drugs DETAILED DESCRIPTION OF THE INVENTION The following detailed descriptions are presented to enable any person skilled in the art to make and use the invention. For purposes of explanation, specific nomenclature is set forth to provide a thorough understanding of the present invention. However, it will be apparent to one skilled in the art that these specific details are not required to practice the invention.
- the cardiac cardenolide compounds of Figure 1 are administered to a mammal for the treatment of a fibrotic disease process.
- these compounds are oleandrin, digitoxin, digoxin, ouabain, digoxigenin, digitoxigenin, acetylstrophanthidin, and digitoxigenin di-acetyl.
- the cardiac cardenolide compounds of Figure 1 are administered to a mammal for the treatment of a fibrotic disease process, using one or more of the cardiac cardenolide compounds of Table 1.
- Another aspect of the invention provides a method for treating a pro- fibrotic disease, in a mammal by administering to a mammal a therapeutically effective amount of one or more cardiac cardenolide compounds typified by those shown in Figure 1 of the present invention.
- Administration of the cardiac cardenolide compounds(s) may occur prior to the manifestation of symptoms characteristic of fibrosis, such that the symptoms of fibrosis are prevented, or alternatively, delayed in its progression.
- Another aspect of the invention provides a method for being included as an adjuvant to existing anti-fibrotic drugs, or fibrotic drugs which may be developed in the future.
- the term "therapeutically effective amount,” as used herein, is that amount that achieves at least partially a desired therapeutic or prophylactic effect in the symptoms of a fibrotic disease process.
- the amount of cardiac cardenolide compound necessary to bring about prevention and/or therapeutic treatment of pro-fibrotoc disease is not fixed per se.
- An effective amount is necessarily dependent upon the identity and the form of the cardiac cardenolide employed, the extent of the protection needed, or the severity of the pro-fibrotic condition.
- Cardiac cardenolide drugs include, but are not limited to, digitoxin, oleandrin, digoxin, and ouabain.
- compositions comprising one or more cardiac cardenolide compound(s) of the present invention and a pharmaceutically acceptable carrier.
- pharmaceutically acceptable carrier is intended to include any and all solvents, solubilizers, fillers, stabilizers, binders, absorbents, bases, buffering agents, lubricants, controlled release vehicles, diluents, emulsifying agents, hemectants, lubricants, dispersion media, coatings, antibacterial or antifungal agents, isotonic and absorption delaying agents, and the like, compatible with pharmaceutical administration.
- solvents solubilizers, fillers, stabilizers, binders, absorbents, bases, buffering agents, lubricants, controlled release vehicles, diluents, emulsifying agents, hemectants, lubricants, dispersion media, coatings, antibacterial or antifungal agents, isotonic and absorption delaying agents, and the like, compatible with pharmaceutical administration.
- diluents e
- a pharmaceutical composition of the invention is formulated to be compatible with its intended route of administration.
- routes of administration include parenteral, e.g., intravenous, intradermal, subcutaneous, oral (e.g., inhalation), transdermal (topical), transmucosal, vagina I, and rectal administration.
- Solutions or suspensions used for parenteral, intradermal, vaginally, or subcutaneous application can include the following components: a sterile diluent such as water for injection, saline solution, fixed oils, polyethylene glycols, glycerine, propylene glycol or other synthetic solvents; antibacterial agents such as benzyl alcohol or methyl parabens; antioxidants such as ascorbic acid or sodium bisulfate; chelating agents such as ethylenediaminetetraacetic acid; buffers such as acetates, citrates or phosphates and agents for the adjustment of tonicity such as sodium chloride or dextrose. pH can be adjusted with acids or bases, such as hydrochloric acid or sodium hydroxide.
- a sterile diluent such as water for injection, saline solution, fixed oils, polyethylene glycols, glycerine, propylene glycol or other synthetic solvents
- antibacterial agents such as benzyl alcohol or methyl parabens
- compositions suitable for injectable use include sterile aqueous solutions (where water soluble) or dispersions and sterile powders for the extemporaneous preparation of sterile injectable solutions or dispersion.
- suitable carriers include physiological saline, bacteriostatic water, Cremophor ELTM (BASF, Parsippany, N.J.) or phosphate buffered saline (PBS).
- the injectable composition should be sterile and should be fluid to the extent that easy syringability exists.
- the carrier can be a solvent or dispersion medium containing, for example, water, ethanol, polyol (for example, glycerol, propylene glycol, and liquid polyethylene glycol).
- EXAMPLE #1 Demonstration that digitoxin reduces the expression of mRNAs associated with fibrosis and epithelial differentiation.
- Cultures of cystic fibrosis lung epithelial IB3-1 and I83-1/S9 cells were grown exactly as described in [27]. IB3-1 cells were treated either with or without digitoxin.
- FIG. 2 shows that digitoxin treatment results in significant reduction in TGFBR2 protein and significant elevation in KRTB protein Example #3. Demonstration that digitoxin can be used as an adjuvant with other drugs.
- FRT Fisher Rat Thyroid
- FIG. 3 shows that compared to the control condition with vehicle (DMSO) alone, VX-770 (3 uM) alone activated chloride current by nearly 3- fold. Digitoxin (25 nM) modestly but significantly activated chloride conductance both as monotherapy and together VX-770.
- Table 1 Digitoxin and gene therapy-induced changes in mRNA signaling for inflammation, fibrosis and epithelial differentiation in 183-1 cells and I83-1/S9 cells (with AAV-[wildtype]CFTR) * *: CF cells were IB3-1 CF lung epithelial cells; gene therapy treated CF cells were IB3-1 cells (IB3-1/S9) that had been stably transfected with AAV- [wildtype]CFTR.
- REFERENCES 1 REFERENCES 1.
- PubMed PMID 7680480; PubMed Central PMCID: PMCPMC45970. 4. Verrecchia F, Mauviel A Transforming growth factor-beta and fibrosis. World journal of gastroenterology. 2007;13(22):3056-62. Epub 2007/06/26. PubMed PMID: 17589920; PubMed Central PMCID: PMCPMC4172611. 5. Babyatsky MW, Rossiter G, Podolsky DK. Expression of transforming growth factors alpha and beta in colonic mucosa in inflammatory bowel disease.
- PubMed PMID 28891814; PubMed Central PMCID: PMCPMC5617658.
- Pohlers D Brenmoehl J, Loffler I, Muller CK, Leipner C, Schultze-Mosgau S, et al. TGF- beta and fibrosis in different organs - molecular pathway imprints. Biochimica et biophysica acta. 2009;1792(8):746-56. Epub 2009/06/23. doi: 10.1016/j.bbadis.2009.06.004. PubMed PMID: 19539753. 10. Xu F, Liu C, Zhou D, Zhang L.
- PubMed PMID 31108916; PubMed Central PMCID: PMCPMC6566291.
- Principe DR DeCant B, Mascarinas E, Wayne EA, Diaz AM, Akagi N, et al. TGFbeta Signaling in the Pancreatic Tumor Microenvironment Promotes Fibrosis and Immune Evasion to Facilitate Tumorigenesis. Cancer research. 2016;76(9):2525-39. Epub 2016/03/17. doi: 10.1158/0008-5472.Can-15-1293.
- PubMed PMID 26980767; PubMed Central PMCID: PMCPMC4873388.
- Bhattacharyya S Wei J, Varga J.
- Srivastava M Eidelman 0, Zhang J, Paweletz C, Caohuy H, Yang Q, Jacobson KA, Heldman E, Huang W, Jozwik C, Pollard BS, Pollard HB.
- Digitoxin mimics gene therapy with CFTR and suppresses hypersecretion of IL-8 from cystic fibrosis lung epithelial cells. Proceedings of the National Academy of Sciences of the United States of America. 2004;101(20):7693-8. Epub 2004/201712. doi: 10.1073/pnas.0402030101. PubMed PMID: 15136726; PubMed Central PMCID: PMCPMC419668.
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Abstract
The present invention is directed to the use of cardiac cardenolide compounds, for the purposes of reducing fibrosis and enhancing epithelial differentiation. In one embodiment the cardiac cardenolide compounds of Figure 1 are administered to a mammal for the treatment of a fibrotic disease process. Examples of these compounds are oleandrin, digitoxin, digoxin, ouabain, digoxigenin, digitoxigenin, acetylstrophanthidin, and digitoxigenin di-acetyl.
Description
CARDIAC CARDENOLIDES TO REDUCE FIBROSIS AND ENHANCE EPITHELIAL DIFFERENTIATION FIELD OF THE INVENTION The present invention relates to the use of cardiac cardenolide compounds for the purposes of reducing fibrosis and enhancing epithelial differentiation BACKGROUND OF THE INVENTION 1.0. Introduction Injury in many tissues is followed by self-limiting fibrosis, which is followed by epithelial differentiation to repair functional damage. An example would be a scar, However, many disease entities are characterized by pathogenic fibrosis in which the process continues and does not stop before creating more damage. Furthermore, epithelial differentiation for detailed repair does not follow. Drugs to arrest fibrosis are thus considered necessary as therapeutics. Detailed repair processes depend on reconstitution of functional structure and cellular function. The subject of histology teaches that differentiated epithelial cells line the outside and inside cavities and lumina within the body. For example, these include the mouth, esophagus, small and large intestine, lungs, GI tract ( including liver and pancreas), the reproductive and urinary tracts, and the exocrine and endocrine glands, sweat glands, mammary glands and salivary glands, and include all organs whose discharges exit to outside the body. Thus drugs that promote epithelial differentiation following injury would be considered advantageous therapeutic entities.
2.0. Fibrosis is driven by Transforming Growth Factor Receptor beta Fibrosis is believed to be due to overactive signaling by Transforming Growth Factor beta (TGFbeta). TGF-beta is therefore referred to as the master regulator of the pro-fibrotic process [1]. Tissues and disease entities so affected include, but are not limited to, the kidney [2], including diabetic nephropathy [3]; gastrointestinal tract [4], including colitis/Crohn's Disease [5]; radiation-induced fibrosis [6]; salivary gland fibrosis [7]; heart, including /myocarditis [8]; rheumatoid arthritis [9]; liver , including hepatic fibrosis [10]; skeletal muscle fibrosis [11]; pancreatic fibrosis [12]; systemic scleroderma/systemic sclerosis [13]; and lung diseases, including chronic obstructive pulmonary disease (COPD), severe forms of asthma, and idiopathic pulmonary fibrosis (IPF), and cystic fibrosis (CF) [14]. The cellular source of TGFbeta in the normal human lung is believed to be the epithelial cells lining the airways [15]. Experimental and clinical suppression of TGFbeta signaling in some of these cases has apparently been accompanied by beneficial consequences. At least two small molecules, pirfenidone [16, 17] and losartan [18], have been approved to target this pathway. However, while both drugs contribute to enhanced progression-free survival in some of these diseases, neither are able to support a durable suppression of the disease process. Side effects are also a problem, and thus development of anti-fibrotic drugs targeting TGFbeta remain a life-limiting, unmet need. 3.0. Reducing TGFbeta expression reduces fibrosis in model systems. Reducing TGFbeta has been shown to be protective in model systems [19, 20]. For example, attempts have been made to reduce circulating TGFbeta levels with a TGFBR2-based ligand trap [21]. In another example, knockout of Tgfbr2 in mice confers increased survival and resistance to
bleomycin-induced pulmonary fibrosis [19]. So far, however, there has been no practical success [22]. Currently approved therapy for bronchiectasis includes inhaled glucocorticoids [23]. A classical example is beclomethasone propionate [24]. However, side effects from this therapeutic class have usually precluded chronic use. Thus for profibrotic disease in a single organ, or diseases in multiple organs, developing drugs that can potently and efficaciously suppress fibrosis remains a life-limiting unmet need. 4.0. Elevated KRT8 is expressed in cells undergoing epithelial differentiation for repair and reconstitution of damaged epithelial tissues. Epithelial differentiation from basal stem cells, known to be marked by Cytokeratin 8 (KRT8), is the mechanism for repair and reconstitution of normal epithelial morphology of damaged epithelial tissues. Organs expressing intrinsically high levels of KRT8 mRNA and protein include most endocrine tissues, bone marrow and immune system, lung, liver and gall bladder, pancreas, gastrointestinal tract, kidney and urinary bladder. In addition, high levels of KRT8 are found in male genital tissues (for example, testes, prostate, epididymis, seminal vesicles), and in female genital tissues (for example, fallopian tubes, breast, vagina, uterine cervix, endometrium, ovary and placenta). Details for KRT8 protein distribution are found in The Human Protein Atlas (protein atlas.erg). In the lung, for example, repair and reconstitution follows the embryological pathway, emphasizing reduction in TGFbeta and elevation in signals such as KRT8, which mark the conversion of basal stem cells to pseudostratified epithelial cells [25]. In the lung, Notch signaling drives basal stem cells to form KRT8-marked intermediate basal luminal precursor cells [26]. KRT8-marked precursor cells then differentiate into all other lung epithelial cells. In the proximal lung the latter include club, ciliated, neuroendocrine cells, and goblet cells. In the distal lung these KRT8- marked precursor cells differentiate into the alveolar Type I and Type II cells,
which mediate oxygen/CO2 exchange [25]. Thus KRT8 denotes cells associated with highly differentiated states in multiple epithelial tissues. Therefore drugs that elevate KRT8, and thus represent the process of epithelial differentiation, might have beneficial therapeutic value. SUMMARY OF THE INVENTION The present invention relates to the use of cardiac cardenolide compounds for the purposes of reducing fibrosis and enhancing epithelial differentiation. BRIEF DESCRIPTION OF THE DRAWINGS Figure 1. Examples of cardiac cardenolide drugs Figure 2. Digitoxin- induced changes in protein expression of TGFBR2 and KRT8. Figure 3. Digitoxin can be used as an adjuvant with other drugs DETAILED DESCRIPTION OF THE INVENTION The following detailed descriptions are presented to enable any person skilled in the art to make and use the invention. For purposes of explanation, specific nomenclature is set forth to provide a thorough understanding of the present invention. However, it will be apparent to one skilled in the art that these specific details are not required to practice the invention. Descriptions of specific applications are provided only as representative examples. Various modifications in the preferred embodiments will be readily apparent to one skilled in the art, and the general principals defined herein
may be applied to other embodiments.and applications without departing from the scope of the invention. The present invention is not intended to be limited to the embodiments shown, but is to be accorded the widest possible scope consistent with the principals and features disclosed herein. In one embodiment the cardiac cardenolide compounds of Figure 1 are administered to a mammal for the treatment of a fibrotic disease process. Examples of these compounds are oleandrin, digitoxin, digoxin, ouabain, digoxigenin, digitoxigenin, acetylstrophanthidin, and digitoxigenin di-acetyl. In another embodiment the cardiac cardenolide compounds of Figure 1 are administered to a mammal for the treatment of a fibrotic disease process, using one or more of the cardiac cardenolide compounds of Table 1. Another aspect of the invention provides a method for treating a pro- fibrotic disease, in a mammal by administering to a mammal a therapeutically effective amount of one or more cardiac cardenolide compounds typified by those shown in Figure 1 of the present invention. Administration of the cardiac cardenolide compounds(s) may occur prior to the manifestation of symptoms characteristic of fibrosis, such that the symptoms of fibrosis are prevented, or alternatively, delayed in its progression. Another aspect of the invention provides a method for being included as an adjuvant to existing anti-fibrotic drugs, or fibrotic drugs which may be developed in the future. The term "therapeutically effective amount," as used herein, is that amount that achieves at least partially a desired therapeutic or prophylactic
effect in the symptoms of a fibrotic disease process. The amount of cardiac cardenolide compound necessary to bring about prevention and/or therapeutic treatment of pro-fibrotoc disease, is not fixed per se. An effective amount is necessarily dependent upon the identity and the form of the cardiac cardenolide employed, the extent of the protection needed, or the severity of the pro-fibrotic condition. Cardiac cardenolide drugs include, but are not limited to, digitoxin, oleandrin, digoxin, and ouabain. The invention is further directed to pharmaceutical compositions comprising one or more cardiac cardenolide compound(s) of the present invention and a pharmaceutically acceptable carrier. As used herein the language "pharmaceutically acceptable carrier" is intended to include any and all solvents, solubilizers, fillers, stabilizers, binders, absorbents, bases, buffering agents, lubricants, controlled release vehicles, diluents, emulsifying agents, hemectants, lubricants, dispersion media, coatings, antibacterial or antifungal agents, isotonic and absorption delaying agents, and the like, compatible with pharmaceutical administration. The use of such media and agents for pharmaceutically active substances is well-known in the art. Except insofar as any conventional media or agent is incompatible with the active compound, use thereof in the compositions is contemplated. Supplementary agents can also be incorporated into the compositions. A pharmaceutical composition of the invention is formulated to be compatible with its intended route of administration. Examples of routes of administration include parenteral, e.g., intravenous, intradermal, subcutaneous, oral (e.g., inhalation), transdermal (topical), transmucosal, vagina I, and rectal administration. Solutions or suspensions used for parenteral, intradermal, vaginally, or subcutaneous application can include
the following components: a sterile diluent such as water for injection, saline solution, fixed oils, polyethylene glycols, glycerine, propylene glycol or other synthetic solvents; antibacterial agents such as benzyl alcohol or methyl parabens; antioxidants such as ascorbic acid or sodium bisulfate; chelating agents such as ethylenediaminetetraacetic acid; buffers such as acetates, citrates or phosphates and agents for the adjustment of tonicity such as sodium chloride or dextrose. pH can be adjusted with acids or bases, such as hydrochloric acid or sodium hydroxide. The parenteral preparation can be enclosed in ampoules, disposable syringes or multiple dose vials made of glass or plastic. Pharmaceutical compositions suitable for injectable use include sterile aqueous solutions (where water soluble) or dispersions and sterile powders for the extemporaneous preparation of sterile injectable solutions or dispersion. For intravenous administration, suitable carriers include physiological saline, bacteriostatic water, Cremophor ELTM (BASF, Parsippany, N.J.) or phosphate buffered saline (PBS). In all cases, the injectable composition should be sterile and should be fluid to the extent that easy syringability exists. It must be stable under the conditions of manufacture and storage and must be preserved against the contaminating action of microorganisms such as bacteria and fungi. The carrier can be a solvent or dispersion medium containing, for example, water, ethanol, polyol (for example, glycerol, propylene glycol, and liquid polyethylene glycol). EXAMPLES: EXAMPLE #1. Demonstration that digitoxin reduces the expression of mRNAs associated with fibrosis and epithelial differentiation. Experiment: Cultures of cystic fibrosis lung epithelial IB3-1 and I83-1/S9 cells were grown exactly as described in [27]. IB3-1 cells were treated either with or
without digitoxin. At the end of the culture period, the cultures were prepared for RNA-seq by standard methods. Parallel studies were also performed on I83-1/S9 cells that had been permanently transfected with AAV- [wildtype]CFTR; however, no digitoxin was added. Samples were analyzed, and log2fold changes in expression were calculated. False Discovery Rates (FDR) were calculated relative to IB3-1 cells that had been cultured without digitoxin. Changes in expression of TGFBR2 and KRTB mRNA and protein in 183-1 cells treated with digitoxin and in IB3-1/S9 cells, were calculated. Differences that were p < 0.05, were taken as significant. TGFBR2 and KRTB were surprisingly found to be significant. Results: Digitoxin treatment and gene therapy with AAV[wildtyope]CFTR resulted in significant reductions in mRNAs for TGFBR2 and KRTB (Table 1). EXAMPLE #2 Demonstration that digitoxin reduces the expression of TGFBR2 protein associated with fibrosis. and increases the expression of KRTB associated with epithelial differentiation. Experiment: IB3-1 cells were grown exactly as described in [27], and treated either with or without digitoxin. At the end of the culture periods, the cultures were assayed for TGFBR2 and KRTB by Western blot analysis. Three independent experiments were performed, and standard ANOVA statistics calculated. Changes that were p <0.05 were taken as significant. Results: Figure 2 shows that digitoxin treatment results in significant reduction in TGFBR2 protein and significant elevation in KRTB protein Example #3. Demonstration that digitoxin can be used as an adjuvant with other drugs. Experiment: Fisher Rat Thyroid (FRT) cells expressing the CF mutation
[G551DJ were grown as monolayers and assayed in a Ussing chamber for activation of CFTR by addition of 100 uM amiloride, 100 uM IBMX and 10 uM forskolin. Experiments were terminated by addition of 10 uM of the CFTR channel blocker CFTRinh-172. Results: Figure 3 shows that compared to the control condition with vehicle (DMSO) alone, VX-770 (3 uM) alone activated chloride current by nearly 3- fold. Digitoxin (25 nM) modestly but significantly activated chloride conductance both as monotherapy and together VX-770. Table 1. Digitoxin and gene therapy-induced changes in mRNA signaling for inflammation, fibrosis and epithelial differentiation in 183-1 cells and I83-1/S9 cells (with AAV-[wildtype]CFTR) *
*: CF cells were IB3-1 CF lung epithelial cells; gene therapy treated CF cells were IB3-1 cells (IB3-1/S9) that had been stably transfected with AAV- [wildtype]CFTR.
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Claims
CLAIMS 1. A cardiac cardenolide compound having structures such as, but not limited to, digitoxin, oleandrin, digoxin, ouabain, digitoxigenin, digoxigenin, acetyl-strophanthin, and digitoxigenin,3,12-diAc. 2. The cardiac cardenolide compound of claim 1, wherein said compound is used to treat fibrosis in kidney, including diabetic nephropathy; gastrointestinal tract, including colitis/Crohn's Disease; radiation-induced fibrosis; salivary glands; heart, including myocarditis;, rheumatoid arthritis; liver, including hepatic fibrosis; skeletal muscle fibrosis; pancreatic fibrosis; systemic scleroderma/systemic sclerosis; and lung diseases including chronic obstructive pulmonary disease (COPD), severe forms of asthma, and idiopathic pulmonary fibrosis (IPF), and cystic fibrosis (CF). 3. The pharmaceutical composition for treating fibrosis in comprising said cardiac cardenolide compound of Claim 1, and a pharmaceutically acceptable carrier. 4. The method of treating fibrosis in a mammal, comprising administering to said mammal a therapeutically effective amount of the pharmaceutical composition of Claim 3. 5. The method of Claim 4 wherein said disease to be treated is kidney, including diabetic nephropathy; gastrointestinal tract, including colitis/Crohn's Disease; radiation-induced fibrosis; salivary glands; heart, including myocarditis;, rheumatoid arthritis; liver, including hepatic fibrosis; skeletal muscle fibrosis; pancreatic fibrosis; systemic · scleroderma/systemic sclerosis; and lung diseases including chronic obstructive pulmonary disease
(COPD), severe forms of asthma, and idiopathic pulmonary fibrosis (IPF), and cystic fibrosis (CF). 6. The method of Claim 4, wherein said pharmaceutical composition is administered orally, intravascularly, vaginally, intramuscularly, subcutaneously, or intraperitoneally. 7. The cardiac cardenolide compound of claim 1, wherein said compound is used to treat kidney, including diabetic nephropathy; gastrointestinal tract, including colitis/Crohn's Disease; radiation-induced fibrosis; salivary glands; heart, including myocarditis; rheumatoid arthritis; liver, including hepatic fibrosis; skeletal muscle fibrosis; pancreatic fibrosis; systemic scleroderma/systemic sclerosis; and lung diseases including chronic obstructive pulmonary disease (COPD), severe forms of asthma, and idiopathic pulmonary fibrosis (IPF), and cystic fibrosis (CF). 8. A pharmaceutical composition for treating kidney, including diabetic nephropathy; gastrointestinal tract, including colitis/Crohn's Disease; radiation-induced fibrosis; salivary glands; heart, including myocarditis;, rheumatoid arthritis; liver, including hepatic fibrosis; skeletal muscle fibrosis; pancreatic fibrosis; systemic scleroderma/systemic sclerosis; and lung diseases including chronic obstructive pulmonary disease (COPD), severe forms of asthma, and idiopathic pulmonary fibrosis (IPF), and cystic fibrosis (CF). or a related condition, comprising said cardiac cardenolide compound of Claim 1, and a pharmaceutically acceptable carrier. 9. A method of treating kidney, including diabetic nephropathy; gastrointestinal tract, including colitis/Crohn's Disease; radiation-induced fibrosis; salivary glands; heart, including myocarditis; rheumatoid arthritis; liver, including hepatic fibrosis; skeletal muscle fibrosis; pancreatic fibrosis;
systemic scleroderma/systemic sclerosis; and lung diseases including chronic obstructive pulmonary disease (COPD), severe forms of asthma, and idiopathic pulmonary fibrosis (IPF), and cystic fibrosis (CF). or related condition in a mammal, comprising administering to said mammal a therapeutically effective amount of the pharmaceutical composition of Claim 3. 10. The method of Claim 4 wherein said disease to be treated is kidney, including diabetic nephropathy; gastrointestinal tract, including colitis/Crohn's Disease; radiation-induced fibrosis; salivary glands; heart, including myocarditis;, rheumatoid arthritis; liver, including hepatic fibrosis; skeletal muscle fibrosis; pancreatic fibrosis; systemic scleroderma/systemic sclerosis; and lung diseases including chronic obstructive pulmonary disease (COPD), severe forms of asthma, and idiopathic pulmonary fibrosis (IPF), and cystic fibrosis (CF). or a related condition. 11. The method of Claim 4, wherein said pharmaceutical composition is administered orally, intravascularly, vaginally, intramuscularly, subcutaneously, or intraperitoneally.
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| PCT/US2019/052217 WO2021054978A1 (en) | 2019-09-20 | 2019-09-20 | Cardiac cardenolides to reduce fibrosis and enhance epithelial differentiation |
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| US8361973B2 (en) * | 2008-08-07 | 2013-01-29 | Centrose, Llc | Glycoside compounds and pharmaceutical compositions thereof |
| US20170067897A1 (en) * | 2007-11-13 | 2017-03-09 | Phoenix Biotechnology, Inc. | Method of Treatment Employing Cardiac Glycoside |
| US20180064730A1 (en) * | 2014-11-05 | 2018-03-08 | Bette Pollard | Cardiac glycosides to reduce pulmonary exacerbations and other adverse events in cystic fibrosis and other related disorders |
| WO2018141678A1 (en) * | 2017-01-31 | 2018-08-09 | Medizinische Hochschule Hannover (Mhh) | Natural compounds and fibrosis |
| US20190038655A1 (en) * | 2016-02-02 | 2019-02-07 | James Allen CARDELLI | Chemicals and methods to prevent and treat tgf-beta mediated activation of fibroblasts to reduce and treat cancer and fibrosis |
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| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US20170067897A1 (en) * | 2007-11-13 | 2017-03-09 | Phoenix Biotechnology, Inc. | Method of Treatment Employing Cardiac Glycoside |
| US8361973B2 (en) * | 2008-08-07 | 2013-01-29 | Centrose, Llc | Glycoside compounds and pharmaceutical compositions thereof |
| US20180064730A1 (en) * | 2014-11-05 | 2018-03-08 | Bette Pollard | Cardiac glycosides to reduce pulmonary exacerbations and other adverse events in cystic fibrosis and other related disorders |
| US20190038655A1 (en) * | 2016-02-02 | 2019-02-07 | James Allen CARDELLI | Chemicals and methods to prevent and treat tgf-beta mediated activation of fibroblasts to reduce and treat cancer and fibrosis |
| WO2018141678A1 (en) * | 2017-01-31 | 2018-08-09 | Medizinische Hochschule Hannover (Mhh) | Natural compounds and fibrosis |
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