# US 2021/0275560 A1 · Azithromycin and roxithromycin derivtives as senolytic drugs > Agents: this data is available as JSON from the patentgrep API and MCP server — see https://patentgrep.com/llms.txt - Number: US 2021/0275560 A1 - Document ID: US20210275560A1 - Status: Published application - Kind: A1 - Application number: 17/282,212 - Filed: 2019-10-02 - Published: 2021-09-09 - Earliest claimed: 2018-10-02 - Assignees: Lunella Biotech, Inc. - Applicants: Lunella Biotech, Inc. - Inventors: Federica SOTGIA; Michael P. LISANTI - Web page: https://patentgrep.com/patent/US20210275560A1 ## CPC - A61K 31/7048: having oxygen as a ring hetero atom, e.g. leucoglucosan, hesperidin, erythromycin, nystatin {, digitoxin or digoxin} - A61K 31/65: Tetracyclines - A61K 31/7052: having nitrogen as a ring hetero atom, e.g. nucleosides, nucleotides - A61K 45/06: Mixtures of active ingredients without chemical characterisation, e.g. antiphlogistics and cardiaca - A61P 39/00: General protective or antinoxious agents - A61K: PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES (devices or methods specially adapted for bringing pharmaceutical products into particular physical or administering forms A61J3/00; chemical aspects of, or use of materials for deodorisation of air, for disinfection or sterilisation, or for bandages, dressings, absorbent pads or surgical articles A61L; soap compositions C11D) - A61P: SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS ## Abstract This disclosure describes the use of azithromycin, roxithromycin, and telithromycin, including derivatives thereof, as senolytic drugs. BrdU was used to induce senescence in model human fibroblast cell lines. Also disclosed are methods for screening compounds for senolytic activity. The SRB assay was used to measure cell viability through protein content. Azithromycin roxithromycin, and telithromycin, clinically-approved pharmaceuticals, were found to be senolytic drugs. However, the closely-related parent compound, erythromycin, showed no senolytic activity. Azithromycin strongly induced both aerobic glycolysis and autophagy in human fibroblasts, but showed bi-phasic effects including on mitochondrial oxygen consumption rates with inhibitory activity at 50 μM and stimulatory activity at 100 μM. The xCELLigence real-time assay system showed that azithromycin preferentially targets senescent cells, removing approximately 97% (nearly a 25-fold reduction in senescent cells). ## Claims ### Claim 1 A senolytic composition comprising a pharmaceutically effective amount of a compound having a formula selected from wherein R1 and R2 may be the same or different, and are selected from a membrane-targeting signal, a mitochondria-targeting signal, hydrogen, carboxyl, an alkane, a cyclic alkane, an alkane-based derivative, an alkene, a cyclic alkene, an alkene-based derivative, an alkyne, an alkyne-based derivative, a ketone, a ketone-based derivative, an aldehyde, an aldehyde-based derivative, a carboxylic acid, a carboxylic acid-based derivative, an ether, an ether-based derivative, an ester, an ester-based derivative, an amine, an amino-based derivative, an amide, an amide-based derivative, a monocyclic arene, a polycyclic arene, a heteroarene, an arene-based derivative, a heteroarene-based derivative, a phenol, a phenol-based derivative, a benzoic acid, and a benzoic acid-based derivative; and at least one of R1 and R2 is one of a membrane-targeting signal selected from palmitic acid, stearic acid, myristic acid, and oleic acid, a short-chain fatty acid, and a medium-chain fatty acid and a TPP-derivative. ### Claim 2 (depends on 1) The senolytic composition of claim 1, wherein at least one of R1 and R2 is myristic acid. ### Claim 3 (depends on 1) The senolytic composition of claim 1, wherein at least one of R1 and R2 is a membrane-targeting signal selected from a short-chain fatty acid and a medium-chain fatty acid. ### Claim 4 (depends on 3) The senolytic composition of the claim 3, wherein at least one of R1 and R2 is a TPP-derivative selected from 2-butene-1,4-bis-TPP; 2-chlorobenzyl-TPP; 3-methylbenzyl-TPP; 2,4-dichlorobenzyl-TPP; 1-naphthylmethyl-TPP; p-xylylenebis-TPP; a derivative of 2-butene-1,4-bis-TPP; a derivative of 2-chlorobenzyl-TPP; a derivative of 3-methylbenzyl-TPP; a derivative of 2,4-dichlorobenzyl-TPP; a derivative of 1-naphthylmethyl-TPP; and a derivative of p-xylylenebis-TPP. ### Claim 5 (depends on 1) The senolytic composition of claim 1, wherein the compound has a formula wherein either R1 is methyl and R2 is one of a membrane-targeting signal and a TPP-derivative, or R1 is one of a membrane-targeting signal and a TPP-derivative and NH—R2 is N(CH3)2. ### Claim 6 (depends on 1) The senolytic composition of claim 1, wherein the compound has a formula wherein either R1 is O—CH2—O—(CH2)2—OCH3 and R2 is one of a membrane-targeting signal and a TPP-derivative, or R1 is one of a membrane-targeting signal and a TPP-derivative and NH—R2 is N(CH3)2. ### Claim 7 (depends on 1) The senolytic composition of claim 1, wherein the compound has a formula wherein either R1 is and R2 is one of a membrane-targeting signal and a TPP-derivative, or R1 is one of a membrane-targeting signal and a TPP-derivative and NH—R2 is N(CH3)2. ### Claim 8 (depends on 5, 6, 7) The senolytic composition of any of claims 5-7, wherein either at least one of R1 and R2 is a membrane-targeting signal selected from palmitic acid, stearic acid, myristic acid, oleic acid, a short-chain fatty acid, and a medium-chain fatty acid, or at least one of R1 and R2 is a TPP-derivative selected from 2-butene-1,4-bis-TPP; 2-chlorobenzyl-TPP; 3-methylbenzyl-TPP; 2,4-dichlorobenzyl-TPP; 1-naphthylmethyl-TPP; p-xylylenebis-TPP; a derivative of 2-butene-1,4-bis-TPP; a derivative of 2-chlorobenzyl-TPP; a derivative of 3-methylbenzyl-TPP; a derivative of 2,4-dichlorobenzyl-TPP; a derivative of 1-naphthylmethyl-TPP; and a derivative of p-xylylenebis-TPP. ### Claim 9 A senolytic composition comprising a therapeutic amount of at least one senolytic agent selected from azithromycin, roxithromycin, telithromycin, an azithromycin analog, a roxithromycin analog, and a telithromycin analog. ### Claim 10 (depends on 9) The senolytic composition of claim 9, wherein the senolytic agent comprises one of an azithromycin analog, a roxithromycin analog, and a telithromycin analog, having a targeting signal comprising at least one of (i) a membrane-targeting signal selected from palmitic acid, stearic acid, myristic acid, oleic acid, a short-chain fatty acid, and a medium-chain fatty acid; and (ii) a TPP-derivative. ### Claim 11 (depends on 10) The senolytic composition of claim 10, wherein the targeting signal is a TPP-derivative selected from 2-butene-1,4-bis-TPP; 2-chlorobenzyl-TPP; 3-methylbenzyl-TPP; 2,4-dichlorobenzyl-TPP; 1-naphthylmethyl-TPP; p-xylylenebis-TPP; a derivative of 2-butene-1,4-bis-TPP; a derivative of 2-chlorobenzyl-TPP; a derivative of 3-methylbenzyl-TPP; a derivative of 2,4-dichlorobenzyl-TPP; a derivative of 1-naphthylmethyl-TPP; and a derivative of p-xylylenebis-TPP. ### Claim 12 (depends on 9, 10, 11) The senolytic composition of any of claims 9-11, further comprising a therapeutic amount of at least one of tetracycline, chlortetracycline, oxytetracycline, demeclocycline, methacycline, doxycycline, minocycline, and tigecycline. ### Claim 13 (depends on 9, 10, 11) The senolytic composition of any of claims 9-11, further comprising a therapeutic amount of at least one of pyrvinium, atovaquone, bedaquiline, irinotecan, sorafenib, niclosamide, stirpentol, chloroquine, and rapamycin. ### Claim 14 (depends on 9, 10, 11) The senolytic composition of any of claims 9-11, further comprising a therapeutic amount of at least one of a mitoriboscin, a mitoketoscin, a mitoflavoscin, 2-butene-1,4-bis-TPP; a derivative of 2-butene-1,4-bis-TPP; 2-chlorobenzyl-TPP; a derivative of 2-chlorobenzyl-TPP; 3-methylbenzyl-TPP; a derivative of 3-methylbenzyl-TPP; 2,4-dichlorobenzyl-TPP; a derivative of 2,4-dichlorobenzyl-TPP; 1-naphthylmethyl-TPP; a derivative of 1-naphthylmethyl-TPP; p-xylylenebis-TPP; and a derivative of p-xylylenebis-TPP. ### Claim 15 (depends on 9, 10, 11) The senolytic composition of any of claims 9-11, further comprising at least one of Vitamin C, berberine, caffeic acid phenyl ester, silibinin, brutieridin, and melitidin. ### Claim 16 (depends on 9) The senolytic composition of claim 9, wherein the senolytic composition is in the form of at least one of a cosmetic, a pill, a lotion, a shampoo, a cream, a soap, a skin cleaner, a shaving preparation, an after-shave, a gel, a stick, a paste, a spray, an aerosol, a powder, a liquid, an aqueous suspension, an aqueous solution, a foam, a transdermal patch, a tincture, and a vapor. ### Claim 17 A composition for use in senescence therapy, the composition comprising a therapeutic amount of a senolytic agent comprising at least one of azithromycin, roxithromycin, telithromycin, an azithromycin analog, a roxithromycin analog, and a telithromycin analog. ### Claim 18 (depends on 17) The composition of claim 17, wherein the senolytic agent comprises one of an azithromycin analog, a roxithromycin analog, and a telithromycin analog, having at least one of (i) a membrane-targeting signal selected from palmitic acid, stearic acid, myristic acid, oleic acid, a short-chain fatty acid, and a medium-chain fatty acid; and (ii) a TPP-derivative. ### Claim 19 (depends on 17, 18) The composition of any of claims 17-18, further comprising a therapeutic amount of at least one of tetracycline, chlortetracycline, oxytetracycline, demeclocycline, methacycline, doxycycline, minocycline, and tigecycline. ### Claim 20 (depends on 17, 18) The composition of any of claims 17-18, further comprising a therapeutic amount of at least one of pyrvinium, atovaquone, bedaquiline, irinotecan, sorafenib, niclosamide, stirpentol, chloroquine, and rapamycin. ### Claim 21 (depends on 17, 18) The composition of any of claims 17-18, further comprising a therapeutic amount of at least one of a mitoriboscin, a mitoketoscin, a mitoflavoscin, 2-butene-1,4-bis-TPP; a derivative of 2-butene-1,4-bis-TPP; 2-chlorobenzyl-TPP; a derivative of 2-chlorobenzyl-TPP; 3-methylbenzyl-TPP; a derivative of 3-methylbenzyl-TPP; 2,4-dichlorobenzyl-TPP; a derivative of 2,4-dichlorobenzyl-TPP; 1-naphthylmethyl-TPP; a derivative of 1-naphthylmethyl-TPP; p-xylylenebis-TPP; and a derivative of p-xylylenebis-TPP. ### Claim 22 (depends on 17, 18) The composition of any of claims 17-18, further comprising at least one of Vitamin C, berberine, caffeic acid phenyl ester, silibinin, brutieridin, and melitidin. ### Claim 23 A method for inducing death of senescent cells in a subject, the method comprising administering to the subject a therapeutic amount of a senolytic agent comprising at least one of azithromycin, roxithromycin, telithromycin, an azithromycin analog, a roxithromycin analog, and a telithromycin analog. ### Claim 24 (depends on 23) The method of claim 23, wherein the senolytic agent comprises one of an azithromycin analog, a roxithromycin analog, and a telithromycin analog, and at least one of (i) a membrane-targeting signal selected from palmitic acid, stearic acid, myristic acid, oleic acid, a short-chain fatty acid, and a medium-chain fatty acid; and (ii) a TPP-derivative. ### Claim 25 (depends on 23) The method of claim 23, further comprising administering a therapeutic amount of at least one of tetracycline, chlortetracycline, oxytetracycline, demeclocycline, methacycline, doxycycline, minocycline, and tigecycline. ### Claim 26 (depends on 23) The method of claim 23, further comprising administering a therapeutic amount of at least one of pyrvinium, atovaquone, bedaquiline, irinotecan, sorafenib, niclosamide, stirpentol, chloroquine, and rapamycin. ### Claim 27 (depends on 23) The method of claim 23, further comprising administering a therapeutic amount of at least one of a mitoriboscin, a mitoketoscin, a mitoflavoscin, 2-butene-1,4-bis-TPP; a derivative of 2-butene-1,4-bis-TPP; 2-chlorobenzyl-TPP; a derivative of 2-chlorobenzyl-TPP; 3-methylbenzyl-TPP; a derivative of 3-methylbenzyl-TPP; 2,4-dichlorobenzyl-TPP; a derivative of 2,4-dichlorobenzyl-TPP; 1-naphthylmethyl-TPP; a derivative of 1-naphthylmethyl-TPP; p-xylylenebis-TPP; and a derivative of p-xylylenebis-TPP. ### Claim 28 (depends on 23) The method of claim 23, further comprising administering at least one of Vitamin C, berberine, caffeic acid phenyl ester, silibinin, brutieridin, and melitidin. ### Claim 29 A method for delaying the onset of an age-related disease in a subject, the method comprising administering to the subject a therapeutic amount of a senolytic agent comprising at least one of azithromycin, roxithromycin, telithromycin, an azithromycin analog, a roxithromycin analog, and a telithromycin analog. ### Claim 30 (depends on 29) The method of claim 29, wherein the age-related disease comprises at least one of atherosclerosis, arthritis, cancer, cardiovascular disease, cataract, dementia, diabetes, hair loss, hypertension, inflammatory disease, kidney disease, muscular atrophy, neurological disease, osteoarthritis, osteoporosis, pulmonary disease, vertebral disc degeneration, and alopecia. ### Claim 31 (depends on 30) The method of claim 30, wherein the neurological disease comprises at least one of mild cognitive impairment, motor neuron dysfunction, Alzheimer's disease, Parkinson's disease, and macular degeneration. ### Claim 32 (depends on 29) The method of claim 29, wherein the senolytic composition comprises at least one of an oral composition, a topical composition, and an intravenous composition. ### Claim 33 (depends on 1, 29, 30, 31, 32) The method of any of claims 29-32, wherein the senolytic agent comprises the compound of claim 1. ### Claim 34 (depends on 29, 30, 31, 32) The method of any of claims 29-32, further comprising administering a therapeutic amount of at least one of tetracycline, chlortetracycline, oxytetracycline, demeclocycline, methacycline, doxycycline, minocycline, and tigecycline. ### Claim 35 (depends on 29, 30, 31, 32) The method of any of claims 29-32, further comprising administering a therapeutic amount of at least one of pyrvinium, atovaquone, bedaquiline, irinotecan, sorafenib, niclosamide, stirpentol, chloroquine, and rapamycin. ### Claim 36 (depends on 29, 30, 31, 32) The method of any of claims 29-32, further comprising administering a therapeutic amount of at least one of a mitoriboscin, a mitoketoscin, a mitoflavoscin, 2-butene-1,4-bis-TPP; a derivative of 2-butene-1,4-bis-TPP; 2-chlorobenzyl-TPP; a derivative of 2-chlorobenzyl-TPP; 3-methylbenzyl-TPP; a derivative of 3-methylbenzyl-TPP; 2,4-dichlorobenzyl-TPP; a derivative of 2,4-dichlorobenzyl-TPP; 1-naphthylmethyl-TPP; a derivative of 1-naphthylmethyl-TPP; p-xylylenebis-TPP; and a derivative of p-xylylenebis-TPP. ### Claim 37 (depends on 29, 30, 31, 32) The method of any of claims 29-32, further comprising administering at least one of Vitamin C, berberine, caffeic acid phenyl ester, silibinin, brutieridin, and melitidin. ### Claim 38 A composition for use in delaying the onset of an age-related disease, the composition comprising a therapeutic amount of a senolytic agent comprising at least one of azithromycin, roxithromycin, telithromycin, an azithromycin analog, a roxithromycin analog, and a telithromycin analog. ### Claim 39 (depends on 38) The composition of claim 38, wherein the age-related disease comprises at least one of atherosclerosis, arthritis, cancer, cardiovascular disease, cataract, dementia, diabetes, hair loss, hypertension, inflammatory disease, kidney disease, muscular atrophy, neurological disease, osteoarthritis, osteoporosis, pulmonary disease, vertebral disc degeneration, and alopecia. ### Claim 40 (depends on 38, 39) The composition of any of claims 38 and 39, wherein the senolytic agent further comprises a therapeutic amount of at least one of tetracycline, chlortetracycline, oxytetracycline, demeclocycline, methacycline, doxycycline, minocycline, tigecycline, pyrvinium, atovaquone, bedaquiline, irinotecan, sorafenib, niclosamide, stirpentol, chloroquine, rapamycin, a mitoriboscin, a mitoketoscin, a mitoflavoscin, 2-butene-1,4-bis-TPP; a derivative of 2-butene-1,4-bis-TPP; 2-chlorobenzyl-TPP; a derivative of 2-chlorobenzyl-TPP; 3-methylbenzyl-TPP; a derivative of 3-methylbenzyl-TPP; 2,4-dichlorobenzyl-TPP; a derivative of 2,4-dichlorobenzyl-TPP; 1-naphthylmethyl-TPP; a derivative of 1-naphthylmethyl-TPP; p-xylylenebis-TPP; a derivative of p-xylylenebis-TPP; Vitamin C, berberine, caffeic acid phenyl ester, silibinin, brutieridin, and melitidin. ### Claim 41 (depends on 1, 2, 3, 4, 5, 6, 7, 38) The composition of claim 38, wherein the senolytic agent comprises the compound of any of claims 1-7. ### Claim 42 A method for screening compounds for senolytic activity, the method comprising: exposing a cell population to a DNA damaging agent for a first duration to generate a senescent cell population; treating at least a portion of the senescent cell population with a candidate compound for a second duration to form a treated cell population; analyzing the treated cell population for at least one marker of senolytic activity. ### Claim 43 (depends on 42) The method of claim 42, wherein the DNA damaging agent comprises bromodeoxyuridine (BrdU). ### Claim 44 (depends on 42) The method of claim 42, wherein the at least one marker of senolytic activity comprises at least one of cell viability, aerobic glycolysis, autophagy, inhibitory activity, and cell proliferation reduction. ### Claim 45 (depends on 42) The method of claim 42, wherein the at least one marker of senolytic activity comprises a quantitatively measurement of autophagic LC3 proteins. ### Claim 46 (depends on 42) The method of claim 42, wherein the first duration is about 8 days. ### Claim 47 (depends on 42) The method of claim 42, wherein the second duration is about 3 days to about 5 days. ### Claim 48 (depends on 42) The method of claim 42, wherein the senescent cell population is exposed to BrdU during the second duration. ### Claim 49 (depends on 42) The method of claim 42, further comprising analyzing at least a portion of the senescent cell population for at least one marker of senolytic activity. ### Claim 50 (depends on 42) The method of claim 42, wherein analyzing the treated cell population for at least one marker of senolytic activity comprises at least one of performing the Sulphorhodamine B assay and measuring cell-induced electrical impedance.