Methods and compositions for initiating, modulating and modulating weight loss and therapeutic applications thereof

Abstract

Methods and systems as well as compounds and compositions for modulating or modulating weight and / or inducing weight loss and other diseases and conditions in a subject are provided. Various methods of administration for modulating or modulating or inducing weight loss and other diseases and conditions are provided. In addition, methods of weight reduction and methods for treating obesity are provided, e.g., by modulating one or more enzymes in the ferroptosis pathway.

Description

[0001] CROSS-REFERENCE TO RELATED APPLICATIONS

[0002] This application claims priority to U.S. Provisional Application No. 63 / 350,218, filed on June 8, 2022, which is incorporated herein by reference in its entirety.

[0003] Overview

[0004] Disclosed herein are methods and compositions for inducing weight loss, fat loss, or both in a subject, the methods comprising administering to the subject an amount of an ferroptosis inducer sufficient to induce weight loss, fat loss, or both. In certain embodiments, it is a method for inducing weight loss. In certain embodiments, it is a method for inducing fat loss. In certain embodiments, it is a method for inducing weight loss and fat loss. In certain embodiments, the method for this purpose further comprises administering a priming agent to the subject before administering an ferroptosis inducer. In certain embodiments, it is a method and composition for this purpose, wherein after administering an ferroptosis inducer to the subject, the adipose tissue in the subject is reduced in weight, volume, or both relative to the weight, volume, or both of the adipose tissue in the subject before administration. In certain embodiments, it is a method and composition for this purpose, wherein after administering an ferroptosis inducer to the subject, the adipose tissue in the subject is reduced in volume relative to the adipose tissue volume before administration, as determined by a spring clip test, by a dual x-ray absorptiometry (DEXA) scan, waist circumference reduction measurement, computer axial tomography (CAT) test, or any combination thereof. In certain embodiments, methods and compositions for this use, wherein after administering a ferroptosis inducer to the subject, the adipose tissue in the subject is reduced in volume relative to the volume of adipose tissue before administration, as determined by a spring clip test, by a dual x-ray absorptiometry (DEXA) scan, waist circumference reduction measurement, computer axial tomography (CAT) test, or any combination thereof. In certain embodiments, methods and compositions for this use, wherein after administering a ferroptosis inducer to the subject, the adipose tissue in the subject is reduced in weight relative to the weight of adipose tissue before administration. In certain embodiments, methods and compositions for this use, wherein the weight reduction is determined as follows: the total weight of the subject after administering the ferroptosis inducer is reduced compared to the total weight of the subject before administering the ferroptosis inducer. In certain embodiments, methods and compositions for this use, wherein the weight reduction is determined as follows: comparing the DEXA scan results of the subject obtained before and after administering the ferroptosis inducer to the subject. In certain embodiments, the method and composition for use further comprises, after administering the ferroptosis inducing agent to the subject, a decrease in the cell size, cell volume, or both of a plurality of cells in the adipose tissue of the subject compared to the cell size, cell volume, or both of a plurality of cells in the adipose tissue of the subject before administration. In certain embodiments, the method and composition for use, wherein the subject is a human.In certain embodiments, methods and compositions for use, wherein the ferroptosis inducing agent is administered orally. In certain embodiments, methods and compositions for use, wherein the administration occurs for about 1 day, 2 days, 3 days, 4 days, 5 days, 1 week, 10 days, 2 weeks, 15 days, 20 days, 3 weeks, 25 days, 30 days, one month, two months, three months, four months, five months, six months, one year, two years, three years, four years, five years, lifelong or as needed. In certain embodiments, methods and compositions for use, wherein the ferroptosis inducing agent is administered in an amount ranging from about 0.1 ng to about 25,000 mg. In certain embodiments, methods and compositions for use are provided, wherein the ferroptosis inducing agent is administered in an amount of about 1 ng, 10 ng, 100 ng, 1 microgram, 10 micrograms, 100 micrograms, 1 mg, 10 mg, 100 mg, 1 g, 2 g, 3 g, 4 g, 5 g, 6 g, 7 g, 8 g, 9 g, 10 g, 15 g, 20 g, or 25 g. In certain embodiments, methods and compositions for use are provided, wherein the ferroptosis inducing agent is administered in an amount ranging from about 1 ng / kg to 1,000 mg / kg, wherein mg is mg of the ferroptosis inducing agent, and kg is kg of the subject's body weight. In certain embodiments, methods and compositions for use are provided, wherein the ferroptosis inducing agent is administered orally. In certain embodiments, methods and compositions for use are provided, wherein the ferroptosis inducing agent is administered by one of the following routes: oral administration, subcutaneous, intradermal, intramuscular, inhalation, intravenous, intraperitoneal, intracranial, intrathecal, transdermal, rectal, or direct administration to adipose tissue. In certain embodiments, methods and compositions for use are provided, which treat at least one of the following: obesity, metabolic syndrome, elevated blood sugar, diabetes, type 2 diabetes, type 3 diabetes, insulin resistance, hypertension, cardiovascular disease, coronary artery disease, cerebrovascular disease, stroke, rheumatic heart disease, arteriosclerosis, atherosclerosis, liver disease, fatty liver disease, non-alcoholic fatty liver disease (NAFLD), non-alcoholic steatohepatitis (NASH), or any combination thereof. In certain embodiments, methods and compositions for use are provided, wherein the subject is a man. In certain embodiments, methods and compositions for use are provided, wherein the subject is a woman. In certain embodiments, methods and compositions for use are provided, wherein the subject is in the range of about 1 to about 18 years old. In certain embodiments, methods and compositions for use are provided, wherein the subject is in the range of about 18 to about 120 years old. In certain embodiments, methods and compositions for use are provided, wherein the subject is a subject in need thereof. In certain embodiments, methods and compositions for use are provided, wherein the ferroptosis inducing agent is contained in a pharmaceutical composition, and the pharmaceutical composition comprises a pharmaceutically acceptable: excipient, carrier, or diluent.In certain embodiments, methods and compositions for this purpose, wherein the pharmaceutical composition is in unit dosage form. In certain embodiments, methods and compositions for this purpose, further comprising administering another therapeutic agent to the subject. In certain embodiments, methods and compositions for this purpose, wherein the another therapeutic agent is administered in parallel with the ferroptosis inducer. In certain embodiments, methods and compositions for this purpose, wherein the another therapeutic agent is administered in conjunction with the ferroptosis inducer. In certain embodiments, methods and compositions for this purpose, wherein the ferroptosis inducer is administered directly to adipose tissue. In certain embodiments, methods and compositions for this purpose, wherein the administration is continuous administration. In certain embodiments, the method and composition for this purpose, wherein the administration lasts for about: 1 / 2 hour, 1 hour, 2 hours, 3 hours, 4 hours, 5 hours, 6 hours, 7 hours, 8 hours, 9 hours, 10 hours, 11 hours, 12 hours, 13 hours, 14 hours, 15 hours, 16 hours, 17 hours, 18 hours, 19 hours, 20 hours, 21 hours, 22 hours, 23 hours, 24 hours, 2 days, 3 days, 4 days, 5 days, 6 days or 1 week. In certain embodiments, the method and composition for this purpose, wherein the adipose tissue is heated to a temperature of about 23, 24, 25, 26, 27, 28, 29, 30, 31, 32 or 33 degrees Celsius. In certain embodiments, the method and composition for this purpose, wherein the ferroptosis inducer is administered in the form of an aqueous solution. In certain embodiments, the method and composition for this purpose, wherein the concentration of the ferroptosis inducer in the aqueous solution is about 0.1nM to about 500μM. In certain embodiments, methods and compositions for this purpose, wherein the administration results in a reduction in the size of fat cells or the number of fat cells in the subject, wherein the reduction is determined as follows: the size of fat cells or the number of fat cells is measured before and after the administration of a therapeutic amount of a ferroptosis regulator, optionally using a microscopic evaluation of a biopsy sample. In certain embodiments, methods and compositions for this purpose, wherein the method further comprises reducing the epididymal white adipose tissue of the subject. In certain embodiments, methods and compositions for this purpose, wherein the subject is overweight or obese. In certain embodiments, methods and compositions for this purpose, wherein the subject is obese and has a BMI of about 30.0 or more. In certain embodiments, methods and compositions for this purpose, wherein the subject is overweight and has a BMI of about 25.0 to about 30. In certain embodiments, methods and compositions for this purpose, wherein the other therapeutic agent has glucagon-like peptide-1 (GLP-1) receptor agonist activity.In certain embodiments are methods and compositions for use, wherein the second therapeutic agent comprises semaglutide, dulaglutide, liraglutide, exenatide, telpotide, or any combination thereof.

[0005] Also disclosed herein are methods and compositions for this purpose. In certain embodiments, disclosed herein are methods and compositions for inducing, regulating or adjusting weight loss, body composition or fat reduction or fat reduction in a subject. In certain embodiments, the method comprises administering to the subject a therapeutically effective amount of an iron death regulator, a glutamate-cysteine ​​ligase (GCL) regulator, an agent that binds GCL, an agent that inhibits GCL, or any combination thereof, thereby inducing, regulating or adjusting weight loss, body composition or fat reduction or fat reduction in the subject. In certain embodiments, it is a method and composition for this purpose, wherein the subject is a person. In certain embodiments, it is a method and composition for this purpose, wherein the subject is a man. In certain embodiments, it is a method and composition for this purpose, wherein the iron death regulator, the glutamate-cysteine ​​ligase (GCL) regulator, the agent that binds GCL, the agent that inhibits GCL, or any combination thereof, is delivered orally as a solution having a concentration of at least about 5 mg of agent / mL solution. In certain embodiments, the method and composition for use, wherein the administration is discontinuous, and the administration occurs daily for about 5 days, 10 days, 15 days, 20 days, 25 days, 30 days, two months, three months, four months, five months, six months or one year. In certain embodiments, the method and composition for use, wherein the ferroptosis inducing agent is selected from: (1S, 3R)-RSL3, ML-162, ML-210, JKE-1674, JKE-1716, erastin, jacaricacid, trigonelline, glutamate / glutamate / glutamate ester (glutamate), sulfasalazine, auranofin, brucein, sorafenib, sorafenib-d3, sorafenib tosylate, trigonelline, FIN56, FINO2, CIL56, dihydroisotanshinone I, GPX4-IN-3, sulfoximine, BSO and salts of any of these. In certain embodiments are methods and compositions for this use, in certain embodiments are methods and compositions for this use, wherein the administration results in a reduction in the size of the subject's adipocytes, wherein the reduction is determined by measuring the adipocyte size before and after administration of the ferroptosis inducer, optionally using a microscopic evaluation of a biopsy sample. In certain embodiments are methods and compositions for this use, wherein the administration results in a reduction in the number of adipocytes in the subject, wherein the reduction is determined by measuring the number of adipocytes before and after administration of a therapeutic amount of a ferroptosis modulator, optionally using a microscopic evaluation of a biopsy sample. In certain embodiments are methods and compositions for this use, wherein the method comprises reducing epididymal white adipose tissue in the subject.In certain embodiments, the method and composition for use are provided, wherein the subject is overweight or obese. In certain embodiments, the method and composition for use are provided, wherein the subject is obese and has a BMI of about 30.0 or more. In certain embodiments, the method and composition for use are provided, wherein the subject is overweight and has a BMI of about 25.0 to about 30. In certain embodiments, the method and composition for use are provided, wherein an additional agent or therapy is administered concurrently or consecutively to the subject. In certain embodiments, the method and composition for use are provided, wherein the additional agent or therapy is selected from: a GLP-1 agonist, a gastric inhibitory polypeptide analog, naltrexone-bupropion or a salt thereof, orlistat or a salt thereof, phentermine or a salt thereof, phentermine-topiramate or a salt thereof, selenium, exercise therapy, surgery, dietary modification, and any combination thereof. In certain embodiments, the method and composition for use, wherein the additional agent or therapy comprises administering a GLP-1 agonist comprising dulaglutide, exenatide, semaglutide, liraglutide, lixisenatide, or a salt of any of these. In certain embodiments, the method and composition for use, wherein the additional agent or therapy comprises administering a gastric inhibitory polypeptide comprising tirpotide or a derivative thereof. In certain embodiments, the method and composition for use, wherein the additional agent or therapy comprises surgery comprising a gastric sleeve, a gastric bypass, an adjustable gastric band, a ball placed in the stomach, or any combination thereof. In certain embodiments, the method and composition for use, wherein the subject has not been diagnosed with cancer. In certain embodiments, the method and composition for use, in certain embodiments, the method and composition for use, wherein the subject is a subject in need thereof. In certain embodiments, the method and composition for use, wherein the subject has type I diabetes, type II diabetes, type IIIa diabetes, or metabolic syndrome. In certain embodiments, the method and composition for use, wherein the medicament is in the form of a pharmaceutical composition, the pharmaceutical composition further comprising a pharmaceutically acceptable: carrier, excipient or diluent. In certain embodiments, the method and composition for use, wherein the pharmaceutical composition is in unit dosage form.

[0006] Also described herein are methods and compositions for this purpose, wherein the composition comprises i) a ferroptosis regulator, a glutamate-cysteine ​​ligase (GCL) regulator, an agent that binds to GCL, an agent that inhibits GCL; and ii) a GLP-1 agonist or a salt thereof, a gastric inhibitory polypeptide analog, naltrexone-bupropion or a salt thereof, orlistat or a salt thereof, phentermine or a salt thereof, and phentermine-topiramate or a salt thereof, or selenium or a salt thereof. In certain embodiments, methods and compositions for this purpose, wherein the composition comprises i) a glutamate-cysteine ​​ligase (GCL) regulator, and ii) a GLP-1 agonist or a salt thereof, a gastric inhibitory polypeptide analog, naltrexone-bupropion or a salt thereof, orlistat or a salt thereof, phentermine or a salt thereof, and phentermine-topiramate or a salt thereof, or selenium or a salt thereof. In certain embodiments, methods and compositions for this purpose, which are pharmaceutical compositions. In certain embodiments, methods and compositions for this purpose, wherein the composition is in unit dosage form. In certain embodiments, methods and compositions for treating a disease or condition. In certain embodiments, methods and compositions for treating a kidney disease or condition. In certain embodiments, methods and compositions for treating symptoms associated with a kidney disease or condition. In certain embodiments, methods and compositions for use, wherein the disease or condition is weight loss, weight maintenance, a disease or condition associated with iron metabolism, or a disease or condition associated with high, low, or abnormal cholesterol.

[0007] Also described herein are methods and compositions for regulating iron metabolism in a plurality of cells in a subject, the methods comprising administering to the subject a therapeutically effective amount or an effective amount of an inducer of ferroptosis, a glutamate-cysteine ​​ligase (GCL) modulator, an agent that binds to GCL, an agent that inhibits GCL, or any combination thereof, thereby regulating iron metabolism in the subject and thereby inducing ferroptosis in the subject. In certain embodiments are methods and compositions for this use, wherein immediately after the regulation, a majority of the plurality of cells remain viable.

[0008] Also described herein are methods and compositions for regulating iron metabolism in a plurality of cells in a subject, the methods comprising administering to the subject a therapeutically effective amount or an effective amount of an ferroptosis-inducing agent, thereby regulating iron metabolism in the subject. In certain embodiments are methods and compositions for this purpose, wherein immediately after the regulation, a majority of the plurality of cells remain viable.

[0009] Also described herein are methods and compositions for treating, inducing, regulating or adjusting iron metabolism or a disease or condition associated with iron metabolism in a subject, the method comprising administering to the subject a therapeutically effective amount or an effective amount of a ferroptosis regulator, a glutamate-cysteine ​​ligase (GCL) regulator, an agent that binds GCL, an agent that inhibits GCL, or any combination thereof, thereby treating, inducing, regulating or adjusting iron metabolism or a disease or condition associated with iron metabolism in the subject. In certain embodiments, methods and compositions for this purpose, wherein the subject is a human. In certain embodiments, methods and compositions for this purpose, wherein the subject is a subject in need thereof. In certain embodiments, methods and compositions for this purpose, wherein a therapeutically effective amount of a ferroptosis regulator is added.

[0010] Also described herein are methods and compositions for regulating cholesterol metabolism in a subject, the methods comprising administering to the subject a therapeutically effective amount or an effective amount of a ferroptosis regulator, a glutamate-cysteine ​​ligase (GCL) regulator, an agent that binds GCL, an agent that inhibits GCL, or any combination thereof, thereby regulating cholesterol metabolism in the subject. In certain embodiments, methods and compositions for this use, wherein the subject is a human. In certain embodiments, methods and compositions for this use, wherein the subject is a subject in need thereof. In certain embodiments, methods and compositions for this use, wherein the ferroptosis inducer is administered.

[0011] Also described herein are methods and compositions for regulating cholesterol metabolism in a subject, the methods comprising administering to the subject a therapeutically effective amount or an effective amount of an inducer of ferroptosis, thereby regulating cholesterol metabolism in the subject. In certain embodiments, methods and compositions for this use, wherein the subject is a human. In certain embodiments, methods and compositions for this use, wherein the subject is a subject in need thereof. In certain embodiments, methods and compositions for this use, wherein the inducer of ferroptosis is administered.

[0012] Also described herein are methods and compositions for treating, inducing, regulating, adjusting or alleviating cholesterol metabolism or a disease associated with cholesterol metabolism in a subject, the method comprising administering to the subject a therapeutically effective amount or an effective amount of a ferroptosis regulator, a glutamate-cysteine ​​ligase (GCL) regulator, an agent that binds GCL, an agent that inhibits GCL, or any combination thereof, thereby treating, inducing, regulating, adjusting or alleviating cholesterol metabolism or a disease or condition associated with cholesterol metabolism in the subject. In certain embodiments, methods and compositions for this purpose, wherein the subject is a human. In certain embodiments, methods and compositions for this purpose, wherein the subject is a subject in need thereof. In certain embodiments, methods and compositions for this purpose, wherein the ferroptosis regulator is administered.

[0013] Also described herein are methods and compositions for treating, inducing, regulating, adjusting, reducing or increasing reactive oxygen species or diseases or conditions associated with reactive oxygen species in a subject, the method comprising administering to the subject a therapeutically effective amount or an effective amount of a ferroptosis regulator, a glutamate-cysteine ​​ligase (GCL) regulator, an agent that binds to GCL, an agent that inhibits GCL, or any combination thereof, thereby treating, inducing, regulating, adjusting, reducing or increasing reactive oxygen species or diseases or conditions associated with reactive oxygen species in the subject. In certain embodiments, methods and compositions for this use, wherein the subject is a human. In certain embodiments, methods and compositions for this use, wherein the subject is a subject in need thereof. In certain embodiments, methods and compositions for this use, wherein the ferroptosis regulator is administered.

[0014] Also described herein are methods and compositions for increasing reactive oxygen species in a subject, the methods comprising administering to the subject a therapeutically effective amount or an effective amount of a ferroptosis modulator, a glutamate-cysteine ​​ligase (GCL) modulator, an agent that binds to GCL, an agent that inhibits GCL, or any combination thereof, thereby increasing reactive oxygen species in the subject. In certain embodiments, methods and compositions for this use, wherein the subject is a human. In certain embodiments, methods and compositions for this use, wherein the subject is a subject in need thereof. In certain embodiments, methods and compositions for this use, wherein the ferroptosis modulator is administered.

[0015] Also described herein are methods and compositions for promoting at least one of the following: treating, overcoming, regulating or reducing acute stress in a subject; chronic stress response; immune hypersensitivity; cachexia; kidney disease; neurodegeneration; cardiotoxicity; or cardiotoxicity caused at least in part by chemotherapy; the method comprises administering to the subject a therapeutically effective amount or an effective amount of a ferroptosis regulator, a glutamate-cysteine ​​ligase (GCL) regulator, an agent that binds to GCL, an agent that inhibits GCL, or any combination thereof, thereby treating, inducing, regulating, adjusting, reducing or increasing acute stress in the subject; chronic stress response; immune hypersensitivity; cachexia; kidney disease; neurodegeneration; cardiotoxicity; or cardiotoxicity caused at least in part by chemotherapy. In certain embodiments, methods and compositions for this use, wherein the subject is a human. In certain embodiments, methods and compositions for this use, wherein the subject is a subject in need thereof. In certain embodiments, methods and compositions for this use, wherein the ferroptosis regulator is administered.

[0016] Also described herein are methods and compositions for promoting at least one of the following: treating kidney disease in a subject, the method comprising administering to the subject a therapeutically effective amount or an effective amount of a ferroptosis regulator, a glutamate-cysteine ​​ligase (GCL) regulator, an agent that binds GCL, an agent that inhibits GCL, or any combination thereof, thereby treating kidney disease in the subject. In certain embodiments, methods and compositions for this purpose, wherein the subject is a human. In certain embodiments, methods and compositions for this purpose, wherein the subject is a subject in need thereof. In certain embodiments, methods and compositions for this purpose, wherein the ferroptosis regulator is administered.

[0017] Also described herein are methods and compositions for promoting at least one of the following: treating a kidney disease in a subject, the method comprising administering to the subject a therapeutically effective amount or an effective amount of an inducer of ferroptosis, thereby treating the kidney disease in the subject. In certain embodiments, methods and compositions for this use, wherein the subject is a human. In certain embodiments, methods and compositions for this use, wherein the subject is a subject in need thereof. In certain embodiments, methods and compositions for this use, wherein the ferroptosis inducer is administered. In certain embodiments, methods and compositions for this use, wherein the method comprises reducing epididymal white adipose tissue of the subject. In certain embodiments, methods and compositions for this use, wherein the subject is overweight or obese. In certain embodiments, methods and compositions for this use, wherein the subject is obese and has a BMI of 30.0 or more. In certain embodiments, methods and compositions for this use, wherein the subject is overweight and has a BMI of 25.0 to <30. In certain embodiments, methods and compositions for this use, wherein additional agents or therapies are administered to the subject in parallel or in a row. In certain embodiments, the method and composition for use are selected from the group consisting of: a GLP-1 receptor agonist, a gastric inhibitory polypeptide analog, naltrexone-bupropion or a salt thereof, orlistat or a salt thereof, phentermine or a salt thereof, phentermine-topiramate or a salt thereof, selenium, exercise therapy, surgery, dietary modification, and any combination thereof. In certain embodiments, the method and composition for use are selected from the group consisting of: a GLP-1 receptor agonist, a gastric inhibitory polypeptide analog, naltrexone-bupropion or a salt thereof, orlistat or a salt thereof, phentermine or a salt thereof, topiramate or a salt thereof, selenium, exercise therapy, surgery, dietary modification, and any combination thereof. In certain embodiments, the method and composition for use are selected from the group consisting of: a GLP-1 receptor agonist, a gastric inhibitory polypeptide analog, naltrexone-bupropion or a salt thereof, ... In certain embodiments, methods and compositions for use, wherein the subject is a subject in need thereof. In certain embodiments, methods and compositions for use, wherein the subject has type I diabetes, type II diabetes, type IIIa diabetes, or metabolic syndrome. In certain embodiments, methods and compositions for use, wherein the medicament is in the form of a pharmaceutical composition, and the pharmaceutical composition further comprises a pharmaceutically acceptable: carrier, excipient, or diluent. In certain embodiments, methods and compositions for use, wherein the pharmaceutical composition is in unit dosage form.In certain embodiments, the methods and compositions for use, wherein the pharmaceutical agent is a compound in Table 1 or a salt thereof. In certain embodiments, the methods and compositions for use, wherein the pharmaceutical agent is in the form of a pharmaceutical composition, the pharmaceutical composition further comprising a pharmaceutically acceptable: carrier, excipient or diluent. In certain embodiments, the methods and compositions for use, wherein the pharmaceutical composition is in unit dosage form.

[0018] Also described herein are methods and compositions for this use, including compositions comprising: i) ferroptosis modulators, glutamate-cysteine ​​ligase (GCL) modulators, agents that bind to GCL, agents that inhibit GCL; and ii) GLP-1 agonists or salts thereof, gastric inhibitory polypeptide analogs, naltrexone-bupropion or salts thereof, orlistat or salts thereof, phentermine or salts thereof, and phentermine-topiramate or salts thereof or selenium or salts thereof. In certain embodiments, methods and compositions for this use, wherein the additional agent or therapy comprises administering a GLP-1 receptor agonist comprising dulaglutide, exenatide, semaglutide, liraglutide, lixisenatide, or a salt of any of these. In certain embodiments, methods and compositions for this use, wherein the additional agent or therapy comprises administering a gastric inhibitory polypeptide comprising tirpotide or a derivative thereof. In certain embodiments, the method and composition for use, wherein the additional agent or therapy comprises a surgical procedure comprising a gastric sleeve, a gastric bypass, an adjustable gastric band, a balloon placed in the stomach, or any combination thereof. In certain embodiments, the method and composition for use, wherein the subject has not been diagnosed with cancer. In certain embodiments, the method and composition for use, wherein the subject is a subject in need thereof. In certain embodiments, the method and composition for use, wherein the subject has type I diabetes, type II diabetes, type IIIa diabetes, or metabolic syndrome. In certain embodiments, the method and composition for use, wherein the medicament is in the form of a pharmaceutical composition, the pharmaceutical composition further comprising a pharmaceutically acceptable: carrier, excipient, or diluent. In certain embodiments, the method and composition for use, wherein the pharmaceutical composition is in unit dosage form. In certain embodiments, the method and composition for use, wherein the medicament is a compound or a salt thereof in Table 1. In certain embodiments, the method and composition for use, wherein the medicament is in the form of a pharmaceutical composition, the pharmaceutical composition further comprising a pharmaceutically acceptable: carrier, excipient, or diluent. In certain embodiments are methods and compositions for such use, wherein the pharmaceutical composition is in unit dosage form.

[0019] Also described herein are methods and compositions for this purpose, including compositions comprising the following substances: i) ferroptosis regulators, glutamate-cysteine ​​ligase (GCL) regulators, agents that bind to GCL, agents that inhibit GCL; and ii) GLP-1 agonists or salts thereof, gastric inhibitory polypeptide analogs, naltrexone-bupropion or salts thereof, orlistat or salts thereof, phentermine or salts thereof, and phentermine-topiramate or salts thereof or selenium or salts thereof. In certain embodiments, methods and compositions for this purpose are pharmaceutical compositions. In certain embodiments, methods and compositions for this purpose are in unit dosage form. In certain embodiments, methods and compositions for treating diseases or conditions. In certain embodiments, methods and compositions for treating kidney diseases or conditions. In certain embodiments, methods and compositions for treating symptoms associated with kidney diseases or conditions. In certain embodiments, methods and compositions for this purpose, wherein the disease or condition is weight loss, weight maintenance, a disease or condition associated with iron metabolism, or a disease or condition associated with high, low or abnormal cholesterol.

[0020] Also disclosed herein are methods of inducing, regulating or adjusting weight loss in a subject. In certain embodiments, the method may comprise administering to the subject (e.g., a mammal, e.g., a mouse, e.g., a male) a therapeutically effective amount of a ferroptosis modulator, a glutamate-cysteine ​​ligase (GCL) modulator, an agent that binds to GCL, an agent that inhibits GCL, an agent that regulates an enzyme in a ferroptosis pathway, or any combination thereof, thereby inducing, regulating or adjusting weight loss.

[0021] Disclosed herein are methods of inducing, regulating or modulating weight loss in a subject. In certain embodiments, the method may comprise administering to the subject (e.g., a mammal, e.g., a human, e.g., a male) a therapeutically effective amount of a ferroptosis modulator, a glutamate-cysteine ​​ligase (GCL) modulator, an agent that binds to GCL, an agent that inhibits GCL, an agent that modulates an enzyme in a ferroptosis pathway, or any combination thereof, thereby inducing, modulating or modulating weight loss.

[0022] In certain embodiments, the agent can be delivered orally (e.g., as a pill or capsule or as a solution) or intravenously. When delivered as a solution, the solution can have a concentration of at least about 5 mg agent / mL solution.

[0023] In certain embodiments, the administration can be discontinuous. The administration can occur daily for about 5 days, 10 days, 15 days, 20 days, 25 days, 30 days, two months, three months, four months, five months, six months or one year. In certain embodiments, the subject can be a mammal, such as a mouse or a human, and can be male.

[0024] Also disclosed herein is a method for treating, inducing, regulating or adjusting iron metabolism or a disease or condition related to iron metabolism in a subject, the method comprising administering to the subject a therapeutically effective amount or an effective amount of a ferroptosis regulator, a glutamate-cysteine ​​ligase (GCL) regulator, an agent that binds to GCL, an agent that inhibits GCL, or any combination thereof, thereby treating, inducing, regulating or adjusting iron metabolism or a disease or condition related to iron metabolism in the subject.

[0025] Further disclosed herein is a method for treating, inducing, regulating, adjusting or reducing cholesterol metabolism or a disease or condition related to cholesterol metabolism in a subject, the method comprising administering to the subject a therapeutically effective amount or an effective amount of a ferroptosis regulator, a glutamate-cysteine ​​ligase (GCL) regulator, an agent that binds to GCL, an agent that inhibits GCL, or any combination thereof, thereby treating, inducing, regulating, adjusting or reducing cholesterol metabolism or a disease or condition related to cholesterol metabolism in the subject.

[0026] In addition, disclosed herein is a method for treating, inducing, regulating, adjusting, reducing or increasing reactive oxygen species or a disease or condition associated with reactive oxygen species in a subject, the method comprising administering to the subject a therapeutically effective amount of a ferroptosis regulator, a glutamate-cysteine ​​ligase (GCL) regulator, an agent that binds to GCL, an agent that inhibits GCL, or any combination thereof, thereby treating, inducing, regulating, adjusting, reducing or increasing reactive oxygen species or a disease or condition associated with reactive oxygen species in the subject.

[0027] Further disclosed herein is a method of promoting, overcoming, or regulating, or reducing, or treating a disease or condition in a subject, the disease or condition comprising: acute stress; chronic stress or stress response; resistance to apoptosis; immune hypersensitivity; cachexia; kidney disease; neurodegeneration, cardiotoxicity, or cardiotoxicity caused at least in part by chemotherapy; the method comprising administering to the subject a therapeutically effective amount or an effective amount of a ferroptosis regulator, a glutamate-cysteine ​​ligase (GCL) regulator, an agent that binds to GCL, an agent that inhibits GCL, or any combination thereof, thereby promoting, overcoming, or regulating, or reducing, or treating the disease or condition in the subject. In certain embodiments, the renal disease comprises one or more of the following disease states or conditions: polycystic kidney dysplasia, Fabry disease, cystinosis, glomerulonephritis, IgA nephropathy, lupus nephritis, atypical hemolytic uremic syndrome (aHUS), nephrotic syndrome, renal injury, polycystic kidney disease (PKD), renal artery stenosis, renal tubular acidosis, simple renal cysts or solitary kidney or single functional kidney. In certain embodiments, the compositions disclosed herein are used to treat renal disease or condition. In certain embodiments, the compositions disclosed herein are used to treat symptoms associated with renal disease or condition.

[0028] In certain embodiments, the pharmaceutical agent may be a compound in Table 1 or a salt thereof.

[0029] In certain embodiments, the agent can be selected from: (1S, 3R)-RSL3, ML-162, ML-210, JKE-1674, JKE-1716, erastin, jacaric acid, trigonelline, glutamate / glutamate / glutamate ester (glutamate), sulfasalazine, auranofin, brucea bruceiol, sorafenib, sorafenib-d3, sorafenib tosylate, trigonelline, FIN56, FINO2, CIL56, dihydroisotanshinone I, GPX4-IN-3, sulfoximine, buthionine sulfoximine (BSO) or a salt or pharmaceutically acceptable salt of any of these.

[0030] In certain embodiments, the administration can result in reducing the adipocyte size, cell number or both of the experimenter. In certain embodiments, the administration can result in reducing the adipocyte size of the experimenter. In certain embodiments, the administration can result in reducing the adipocyte number in the experimenter.

[0031] In certain embodiments, the reduction can be determined by measuring adipocyte size before and after administration of a therapeutic amount of a ferroptosis modulator. For example, the measurement can be performed microscopically on a biopsy sample from a subject.

[0032] In certain embodiments, the method can include reducing the epididymal white adipose tissue of the experimenter. In certain embodiments, the experimenter can be overweight or obese. In certain embodiments, the experimenter can be obese and have a body mass index (BMI) of 30.0 or higher. In certain embodiments, the experimenter can be overweight and have a BMI of 25.0 to <30.

[0033] In certain embodiments, the subject may be administered concurrently or consecutively with a second weight loss therapy. In certain embodiments, the second weight loss therapy may be selected from: GLP-1 agonists, gastric inhibitory polypeptide analogs, naltrexone-bupropion or a salt thereof, orlistat or a salt thereof, phentermine or a salt thereof and phentermine-topiramate or a salt thereof, selenium, exercise therapy, surgery, dietary modification, and any combination thereof.

[0034] In certain embodiments, the second therapy may comprise administration of a GLP-1 agonist comprising dulaglutide, exenatide, semaglutide, liraglutide, lixisenatide, an analog, derivative, or salt of any of these.

[0035] In certain embodiments, the second therapy may comprise administration of a gastric inhibitory polypeptide comprising telpotide or a derivative thereof.

[0036] In certain embodiments, the second therapy may comprise a surgical procedure including a gastric sleeve, gastric bypass, an adjustable gastric band, a balloon placed in the stomach, or any combination thereof.

[0037] In certain embodiments, the subject has not been diagnosed with cancer.

[0038] In certain embodiments, the subject may be a subject in need thereof. In certain embodiments, the subject in need thereof has, is suspected of having, or is diagnosed with type I diabetes, type II diabetes, type IIIa diabetes, or metabolic syndrome. In certain embodiments, the subject is at risk of developing type I diabetes, type II diabetes, type IIIa diabetes, or metabolic syndrome.

[0039] In certain embodiments, the medicament is in the form of a pharmaceutical composition, which further comprises a pharmaceutically acceptable carrier, excipient or diluent.

[0040] In certain embodiments, the pharmaceutical composition is in unit dosage form.

[0041] Also described herein are compositions comprising i) an agent, a ferroptosis modulator or inducer, a glutamate-cysteine ​​ligase (GCL) modulator, an agent that binds to GCL, or an agent that inhibits GCL; and ii) a GLP-1 agonist or a salt thereof, a gastric inhibitory polypeptide analog, naltrexone-bupropion or a salt thereof, orlistat or a salt thereof, phentermine or a salt thereof, and phentermine-topiramate or a salt thereof, or selenium or a salt thereof.

[0042] In certain embodiments, the composition may be a pharmaceutical composition. In certain embodiments, the pharmaceutical composition is in unit dosage form.

[0043] In certain embodiments, the administration of the pharmaceutical composition can be carried out in an effective amount or a therapeutically effective amount, and can be oral or intravenous administration. In certain embodiments, administration is once, twice, three times or four times a day for one day, one week, one month, six months, one year or longer, or as needed. BRIEF DESCRIPTION OF THE DRAWINGS

[0045] The novel features of the present invention are particularly set forth in the appended claims. A better understanding of the features and advantages of the present invention will be obtained by reference to the following detailed description and the accompanying drawings which set forth exemplary embodiments in which the principles of the present invention are utilized:

[0046] Figure 1 is a schematic diagram of the ferroptosis pathway.

[0047] Figure 2 It is shown that specific perturbations of ferroptosis regulation lead to ferroptosis that affects adipose lipid accumulation. The top panel shows images of wild-type (WT) and knockout (KO) adipose tissue. The bottom panel shows epididymal / gonadal white adipose tissue (mg) in WT and KO.

[0048] Figure 3 Schematic diagram showing the function of NADPH in lipid synthesis and defense against oxidative stress, for example as a cofactor for glutathione reductase.

[0049] Figures 4A-4D Shown are changes in weight loss of male controls and males administered a ferroptotic agent (FA). Figure 4A Shown is a graph of percent body weight change over time for male control mice and male mice treated with 5 mg / ml of a ferroptosis inducer. Figure 4B Graphs showing water intake over time for male controls and males administered ferroptosis agents; Figure 4C Shown are epididymal white adipose tissue (eWAT) weights of male controls and males administered with ferroptosis agents; water intake of male controls and male mice administered with ferroptosis agents. Figure 4D Shown are the levels of hepatic GSH concentrations in male controls and males administered a ferroptotic agent.

[0050] Figure 5 Images of fat cells in control mice and mice treated with 5 mg / ml ferroptosis agent are shown. Mice treated with 5 mg / mL ferroptosis agent showed reduced fat.

[0051] Figures 6A-6E showed that mice treated with ferroptosis agents did not gain weight when fed a high-fat diet (HFD). Fig. 6A Shown are weight changes of untreated and ferroptotic agent-treated diet-induced obese (DIO) mice when fed a HFD diet. Figure 6B Shown are weight changes of C57Bl / 6 mice treated with ferroptotic agents. Figure 6C Shown are eWAT weights of DIO and C57B1 / 6 treated and untreated mice. Figure 6D-6E Triglycerides in DIO and C57B1 / 6 treated and untreated mice are shown. Fig.6D ) and LDL cholesterol ( Fig. 6E )'s lipid profile.

[0052] Figure 7 A simplified diagram is shown illustrating the effects of perturbations in ferroptosis regulation on adipose lipid accumulation.

[0053] Figure 8 A simplified diagram is shown illustrating the targets that enable precise ferroptosis induction.

[0054] Fig. 9 An overview of ferroptosis resulting from a cell disintegration process due to uncontrolled lipid peroxidation is shown.

[0055] Fig.10 Experiments shown indicate that ferroptosis sensitivity is associated with apoptosis resistance in melanoma cancer cells (A375) and (PDX), pancreatic cancer cells (KP4), breast cancer cells (BT474), lung cancer cells (PC9), and ovarian cancer cells (Kuramochi). Cells were treated with different concentrations of glutathione peroxidase 4 inhibitor (GPX4i).

[0056] Fig.11 A schematic depicting that ferroptosis sensitivity can be driven by prosurvival stress is shown.

[0057] Fig.12 Examples are shown of glutathione regulation as an anchor for a variety of different cellular processes that can lead to disease, such as stress defense, neurodegeneration, selenium turnover, inflammation, infection, and redox signaling.

[0058] Fig.13 A schematic diagram showing the application of ferroptotic cell states to disease treatment.

[0059] Fig.14 Induction of ferroptosis in tumors in vivo is shown. Histological images of controls, ferroptosis inducers alone, and ferroptosis inducers and ferroptosis preventers in cells in vivo are shown. Ferropoptosis inducers alone mimic ferroptosis. Ferropoptosis inducers and ferroptosis preventers inhibit ferroptosis.

[0060] Detailed Description of the Disclosure

[0061] The following description and examples illustrate embodiments of the present invention in detail. It should be understood that the present invention is not limited to the specific embodiments described herein, and therefore can be varied. It will be appreciated by those skilled in the art that there are numerous variations and modifications within the scope of the present invention.

[0062] definition

[0063] Throughout the present disclosure, various embodiments can be presented in a range format. It should be understood that the description of the range format is only for convenience and simplicity, and should not be interpreted as a strict limitation of the scope of any embodiment. Therefore, the description of the scope should be considered to have specifically disclosed all possible sub-ranges and each numerical value within the range up to one-tenth of the lower limit unit, unless the context clearly indicates otherwise. For example, the description of the scope such as from 1 to 6 should be considered to have specifically disclosed sub-ranges such as from 1 to 3, from 1 to 4, from 1 to 5, from 2 to 4, from 2 to 6, from 3 to 6, and each value within the range, for example, 1.1, 2, 2.3, 5 and 5.9. Regardless of the width of the scope, this is applicable. The upper and lower limits of these insertion ranges can be independently included in a smaller range, and are also included in the present disclosure, except for any explicitly excluded limitation within the scope. In the case where the scope includes one or two restrictions, the scope excluding any one or two restrictions in those included restrictions is also included in the present disclosure, unless the context clearly indicates otherwise.

[0064] Unless otherwise specified or obvious from the context, the term "about" used herein is understood to be within the range generally tolerated in the art, such as within 2 standard deviations of the mean value. "About" can be understood to be within 15%, 14%, 13%, 12%, 11%, 10%, 9%, 8%, 7%, 6%, 5%, 4%, 3%, 2%, 1%, 0.5%, 0.1%, 0.05% or 0.01% of the value. Unless otherwise clear from the context, all numerical values ​​provided herein are modified by the term "about". In the case where the application and claims describe specific values, unless otherwise stated, the term "about" is implicit and refers to the acceptable error range of the specific value in this context.

[0065] Unless otherwise stated, structures depicted herein are also meant to include all isomeric (e.g., enantiomeric, diastereomeric, geometric (or conformational)) forms of the structure; for example, L and S designations for each asymmetric center, R and S configurations for each asymmetric center, (Z) and (E) double bond isomers, and (Z) and (E) conformational isomers. Therefore, single stereochemical isomers as well as enantiomeric, diastereomeric, and geometric (or conformational) mixtures of the compounds of the invention are within the scope of the present disclosure.

[0066] Unless otherwise stated, reference herein to compounds having one or more asymmetric centers includes enantiomerically pure compounds, diastereomeric compounds, and racemic mixtures thereof.

[0067] The term "adjacent" and its grammatical equivalents used herein mean next to the reference object. For example, in the context of cells or tissues, the term "adjacent" may mean that there are no other cells or tissues between the two.

[0068] As used herein, the term "analog" and its grammatical equivalents refer to molecules that are not identical but have similar structural features. An analog of a drug or agent is a drug or agent that is related to a reference agent (e.g., an agent provided in Table 1) but whose chemical structure may be different. In general, an analog exhibits similar activity to the reference drug or agent, but the activity may be increased, decreased, or otherwise improved. In general, an analog form of a compound or drug refers to a compound or drug in which the backbone core of the structure is modified or altered compared to the reference drug.

[0069] The term "cancer" and its grammatical equivalents used herein refer to excessive proliferation of cells, whose unique characteristics (loss of normal control) lead to unregulated growth, lack of differentiation, local tissue invasion and metastasis. For the methods provided herein, the cancer can be any cancer, including, but not limited to any of the following: acute lymphocytic cancer, acute myeloid leukemia, alveolar rhabdomyosarcoma, bladder cancer, bone cancer, brain cancer, breast cancer, anal cancer, anal canal cancer, rectal cancer, eye cancer, intrahepatic bile duct cancer, joint cancer, cervical cancer, gallbladder cancer or pleural cancer, nasal cancer, nasal cancer or middle ear cancer, oral cancer, vulvar cancer, chronic lymphocytic leukemia, chronic myeloid cancer, colon cancer, esophageal cancer, cervical cancer , fibrosarcoma, gastrointestinal cancer, Hodgkin's lymphoma, hypopharyngeal cancer, kidney cancer, laryngeal cancer, leukemia, liquid tumors, liver cancer, lung cancer, lymphoma, malignant mesothelioma, mast cell tumor, melanoma, multiple myeloma, nasopharyngeal cancer, non-Hodgkin's lymphoma, ovarian cancer, pancreatic cancer, peritoneal, omental and mesenteric cancer, pharyngeal cancer, prostate cancer, colorectal cancer, kidney cancer, skin cancer, small intestine cancer, soft tissue cancer, solid tumors, stomach cancer, testicular cancer, thyroid cancer, ureter cancer and / or bladder cancer. As used herein, the term "tumor" refers to an abnormal growth of cells or tissues, such as malignant or benign types.

[0070] The terms "effective amount" or "therapeutically effective amount" and grammatical equivalents thereof mean an amount sufficient to achieve or at least partially achieve a desired effect.

[0071] The term "expression" and its grammatical equivalents used herein refer to the biosynthesis of a gene product. For example, in the case of a structural gene, expression involves transcription of the structural gene into mRNA and translation of the mRNA into one or more polypeptides.

[0072] The term "ferroptosis" refers to a form of cell death that involves the production of reactive oxygen species mediated by iron and is characterized in part by lipid peroxidation. The term "ferroptosis inducer" or "ferroptosis activator" or "ferroptosis inducer" or "ferroptosis inducing compound" refers to an agent that promotes or activates ferroptosis in a cell.

[0073] As used herein, the term "hyperproliferative cell" and grammatical equivalents thereof refer to cells characterized by undesirable cell proliferation or an abnormally high rate or sustained cell division, unrelated or uncoordinated with cells of the surrounding normal tissue.

[0074] As used herein, the term "in vitro" and its grammatical equivalents refer to events that occur in an artificial environment (eg, in a test tube or reaction vessel, in cell culture, etc.) rather than in vivo in a multicellular organism.

[0075] As used herein, the term "in vivo" and its grammatical equivalents refer to events that occur within a multicellular organism, such as a non-human animal.

[0076] As used herein, the term "iron-dependent cell death agent" and its grammatical equivalents refer to an agent that induces, promotes or activates cell death mediated by iron. In some cases, in the present disclosure, the term "iron-dependent cell death agent" is used interchangeably with ferroptosis inducer.

[0077] As used herein, the term "normal cell" and grammatical equivalents thereof refer to a cell undergoing controlled cell division, controlled activation, or a quiescent cell.

[0078] The terms used herein are only used for the purpose of describing specific embodiments and are not intended to limit any embodiment. The singular forms "one", "a kind of" and "said" used herein are also intended to include plural forms, unless the context clearly indicates otherwise. It is further understood that the terms "comprising" and / or "including" when used in this specification specify the presence of stated features, integers, steps, operations, elements and / or components, but do not exclude the presence or addition of one or more other features, integers, steps, operations, elements, components and / or their groups. The term "and / or" used herein includes any combination and all combinations of one or more of the listed related items.

[0079] Overview

[0080] Provided herein are compositions and methods that can be used to treat diseases or conditions (such as overweight or obesity). Also provided herein are compositions for use using any of the methods described herein. Also provided herein are treatment regimens for treating various diseases or conditions (such as overweight or obesity). The treatment regimen may include administering an agent or ferroptosis regulator alone or in combination with a second therapy, each administered alone in an effective amount or a therapeutically effective amount, thereby treating the disease or condition. In the case of administering two or more agents or active agents, the administration may be sequential or concurrent and separate or in a single dosage form, which may be a fixed-dose combination drug.

[0081] Iron Death

[0082] Cell death is a cellular process involved in development, cell homeostasis, and prevention of proliferative diseases such as cancer. Programmed cell death can take different forms, such as apoptosis, mitotic catastrophe, necrosis, senescence, and autophagy. Although each of these processes ultimately leads to cell death, the pathways and mechanisms appear to be unique at the molecular and cellular levels.

[0083] Ferroptosis is a non-apoptotic oxidative form of regulated cell death, which involves the accumulation of lipid hydroperoxides and lipid peroxides at the cytoplasmic membrane. Cells undergoing ferroptosis do not show cell features or functions associated with apoptosis (a canonical form of cell death). Examples of apoptotic cell features include, for example, mitochondrial cytochrome c release, caspase activation, and chromatin fragmentation. Ferroptosis is also characterized by increased intracellular reactive oxygen species (ROS) levels, which can be prevented by genetic inhibition of iron chelation and cellular iron uptake. The addition of iron (rather than other divalent transition metal ions) can enhance ferroptosis signaling in cells.

[0084] Cellular components associated with and regulating ferroptosis include, among others, the cysteine-glutamate antiporter (system Xc), glutathione peroxidase 4 (GPX4), p53, the cargo receptor NCOA4, glutathione synthetase (GSH), glutamate-cysteine ​​ligase (GCL). Inactivation or inhibition of some of these molecules (e.g., system Xc, GPX4, or glutathione synthetase) can lead to iron-dependent cell death or ferroptosis.

[0085] It has been found that hyperproliferative cells in a drug-resistant state (such as, for example, drug-resistant cancer cells) exhibit dysregulation in apoptotic cell pathways. Surprisingly, hyperproliferative cells resistant to apoptotic agents can have an enhanced ability to undergo ferroptosis. Due to their "flammable" ferroptosis-sensitive state, apoptosis-resistant cells can be killed by ferroptosis induction.

[0086] In certain embodiments, perturbations in ferroptosis regulation can be used to treat diseases and conditions associated with reactive oxygen species, iron metabolism, lipid metabolism, glutathione biosynthesis, cholesterol metabolism, selenium metabolism, or any combination thereof.

[0087] In certain embodiments, ferroptosis regulatory perturbations can affect adipose lipid accumulation. Ferroptosis regulatory perturbations can reduce fat accumulation and triglycerides. In some cases, ferroptosis regulatory perturbations can reduce obesity and liver fat. Ferroptosis regulatory perturbations can lead to metabolic disease prevention. In some cases, selenium and / or ferroptosis regulatory perturbations can increase UCP1 expression. In some cases, selenium and / or ferroptosis regulatory perturbations can increase thermogenesis and energy expenditure, which can increase metabolic disease prevention.

[0088] Methods for characterizing ferroptosis-sensitive cells

[0089] Provided herein is a method for identifying and characterizing ferroptosis-sensitive cells in a subject. In certain embodiments, the characterization is performed before treating a subject with an ferroptosis-inducing agent provided herein. Ferroptosis-sensitive cells can be identified by the following properties provided herein: (1) a selenium concentration greater than the selenium concentration in the corresponding normal cells; (2) an iron concentration greater than the iron concentration in the corresponding normal cells; (3) a polyunsaturated fatty acid (PUFA) concentration greater than the PUFA concentration in the corresponding normal cells; (4) a peroxidation index (PI) greater than the PI in the corresponding normal tissue; and / or (5) one or more markers indicating mesenchymal state, and other morphological and histological features. Methods for measuring the analyte concentrations of selenium, iron, and PUFA include, for example, mass spectrometry, chromatography, immunoassays, immunosorbent assays, absorbance and colorimetric assays, and microwave plasma-atomic emission spectroscopy. Methods for measuring markers of mesenchymal cell states include, for example, immunoassays, polymerase chain reaction (PCR) assays, and sequencing assays.

[0090] (1) Selenium (Se) concentration and selenoprotein

[0091] Selenium (Se) is a micronutrient that contributes to the synthesis of selenoproteins in cells. Dietary selenium is present in meat, nuts, grains, mushrooms and vegetables. The selenium content in the human body is in the range of about 13 milligrams (mg) to 20 mg. Selenium is involved in the cellular process of selenoprotein synthesis and ferroptosis. Selenoproteins are rare proteins that contain selenocysteine ​​(Sec) residues to replace cysteine. Non-limiting examples of selenoproteins include GPX1, GPX2, GPX3, GPX4, GPX6, TXNRD1, TXNRD2 (TXRD2), TXNRD3, DIO1, DIO2, DIO3, SEPHS2, SEPS1, SEPP1, SEP15, SEPN1 (SELENON), SEPX1, SEPW1 (SELENOW), SEPT1, SELH, SELI, SELK, SELM (SELENOM), SELO and SELV. Selenoproteins exhibit biochemical activities such as redox, selenocysteine ​​synthesis and / or selenium transport. GPX4 is a phospholipid hydroperoxidase that catalyzes the reduction of hydrogen peroxide and organic peroxides, thereby protecting cells from membrane lipid peroxidation and oxidative stress. GPX4 is a regulator of the ferroptosis pathway, and inhibition of GPX4 induces ferroptotic cell death.

[0092] Provided herein is a method for identifying iron-death-sensitive cells in mammalian tissues by selenium concentration. In certain embodiments, provided herein is a method comprising measuring selenium (Se) concentration in a cell, a plurality of cells or a mammalian tissue. In certain embodiments, the Se concentration in a cell or a plurality of cells of a mammalian tissue is at least about 1%, 2%, 3%, 4%, 5%, 6%, 7%, 8%, 9%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95% or 100% greater than the Se concentration in the cells of healthy tissue. In certain embodiments, the Se concentration in multiple cells of mammalian tissue is 1%-10%, 10%-20%, 20%-30%, 30%-40%, 40%-50%, 50%-60%, 60%-70%, 70%-80%, 80%-90% or 90%-100% greater than the Se concentration in cells of healthy tissue. In certain embodiments, the methods provided herein comprise administering an effective amount of an ferroptosis inducing agent to a mammal, wherein the selenium concentration of multiple cells of mammalian tissue is greater than the selenium concentration of cells of normal or healthy tissue; and inducing ferroptosis in the multiple cells.

[0093] (2) Iron concentration

[0094] Ferroptosis is an iron-dependent cellular process, and ferroptosis-sensitive cells have increased intracellular iron concentrations compared to normal cells. Cells treated with deferoxamine (DFO) can inhibit cell death, which is an iron chelator used to treat iron overload and a medicament reported to block ferroptosis. Alternatively, iron loading into cells by treatment with ferric ammonium citrate (FAC) is sufficient to simulate particle treatment and induce ferroptosis in amino acid-starved cells. Increased iron uptake in cells can lead to depletion of glutathione, which may be due to increased ROS generation that leads to ferroptosis induction.

[0095] Provided herein is a method for identifying ferroptosis-sensitive cells in mammalian tissues by iron concentration. In certain embodiments, provided herein is a method comprising measuring the concentration of iron or iron oxide in a cell, a plurality of cells or a mammalian tissue. In certain embodiments, the ferroptosis-sensitive cells comprise an increased intracellular iron concentration of at least about 7 parts per billion (ppb) or more, about 8 ppb or more, about 9 ppb or more, about 10 ppb or more, about 20 ppb or more, about 30 ppb or more, about 40 ppb or more, about 50 ppb or more, about 60 ppb or more, about 70 ppb or more, about 80 ppb or more, about 90 ppb or more, about 100 ppb or more, about 110 ppb or more, about 120 ppb or more, about 130 ppb or more, about 140 ppb or more, about 150 ppb or more, about 160 ppb or more, at most 170 ppb. In certain embodiments, the ferroptosis-sensitive cells comprise an increased intracellular iron concentration that is at least about 2 micromolar (μM) or more, 2.5 μM or more, 3.0 μM or more, 4.0 μM or more, 5.0 μM or more, up to 10 μM higher than the intracellular iron concentration of normal cells. In certain embodiments, the iron concentration in one or more cells of a mammalian tissue is at least about 1%, 2%, 3%, 4%, 5%, 6%, 7%, 8%, 9%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, or 100% greater than the iron concentration in cells of healthy tissue. In certain embodiments, the iron concentration in multiple cells of mammalian tissue is 1%-10%, 10%-20%, 20%-30%, 30%-40%, 40%-50%, 50%-60%, 60%-70%, 70%-80%, 80%-90% or 90%-100% greater than the iron concentration in cells of healthy tissue. In certain embodiments, the methods provided herein include administering an effective amount of an ferroptosis inducing agent to a mammal, wherein the iron concentration of multiple cells of a mammalian tissue is greater than the iron concentration of cells of a normal or healthy tissue; and inducing ferroptosis in the multiple cells. In certain embodiments, the methods provided herein include administering an effective amount of an ferroptosis inducing agent to a mammal, wherein the iron concentration of multiple cells of a mammalian tissue is greater than the iron concentration of cells of a normal or healthy tissue, thereby affecting iron metabolism; and inducing ferroptosis in the multiple cells.

[0096] (3) PUFA status

[0097] Apoptosis-resistant cells gain an advantage by being in a ferroptosis-sensitive state with high levels of polyunsaturated fatty acids (PUFAs). Due to their "combustible" high-PUFA state, apoptosis-resistant cells can be killed by ferroptosis induction. The combustible state is defined by a high membrane abundance of PUFAs (as opposed to MUFAs, monosaturated fatty acids) that are prone to uncontrolled lipid peroxidation (free radical chain reactions of polyunsaturated fatty acids), which leads to ferroptotic cell death.

[0098] PUFAs are classified as ω-3 (n-3) and ω-6 (n-6) based on the position of the last double bond relative to the terminal methyl end of the molecule. Non-limiting examples of PUFAs include: hexadecatrienoic acid (HTA), alpha-linolenic acid (ALA), stearidonic acid (SDA), eicosatrienoic acid (ETE), eicosatetraenoic acid (ETA), eicosapentaenoic acid (EPA, eicosapentaenoic acid), heneicosapentaenoic acid (HPA), docosapentaenoic acid (DPA, ichthyic acid), docosahexaenoic acid (DHA, ichthyic acid), tetracosahexaenoic acid (herring acid), tetracosapentaenoic acid, linoleic acid (LA), gamma-linolenic acid (GLA), eicosadienoic acid, dihomo-gamma-linolenic acid (DGLA), arachidonic acid (AA), docosadienoic acid, adrenic acid (AdA), docosapentaenoic acid (osbond acid), tetracosatetraenoic acid, and tetracosapentaenoic acid. Humans can synthesize all fatty acids that are utilized by the body except linoleic acid (LA, C18:2n-6) and α-linolenic acid (ALA, C18:3n-3).

[0099] Provided herein is a method for identifying ferroptosis-sensitive cells in mammalian tissues by the concentration of PUFA. In certain embodiments, the method provided herein comprises administering an effective amount of an ferroptosis-inducing agent to a mammal, wherein the polyunsaturated fatty acid (PUFA) concentration of multiple cells of mammalian tissue is greater than the PUFA concentration of cells of normal or healthy tissue; and inducing ferroptosis in the multiple cells. In certain embodiments, the PUFA concentration in multiple cells of mammalian tissue is greater than the PUFA concentration in cells of healthy or non-malignant tissues of mammals. In certain embodiments, the PUFA concentration in multiple cells of mammalian tissue is at least about 1%, 2%, 3%, 4%, 5%, 6%, 7%, 8%, 9%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95% or 100% greater than the PUFA concentration in cells of healthy tissues. In certain embodiments, the concentration of PUFAs in a plurality of cells of a mammalian tissue is 1%-10%, 10%-20%, 20%-30%, 30%-40%, 40%-50%, 50%-60%, 60%-70%, 70%-80%, 80%-90%, or 90%-100% greater than the concentration of PUFAs in cells of a healthy tissue. In certain embodiments, the concentration of PUFAs in a plurality of cells of a mammalian tissue is greater than a predetermined PUFA concentration. In certain embodiments, the predetermined PUFA concentration is about 5, 10, 15, 20, 25, 30, 35, 40, 45, 50, 55, 60, 65, 70, 75, 80, 85, or 90 mole percent of total lipids. In certain embodiments, the predetermined PUFA concentration is about 10-20, 20-30, 30-40, 40-50, 50-60, 60-70, 70-80, or 80-90 mol % of total lipids. In certain embodiments, the predetermined PUFA concentration is about 20 mol % of total lipids.

[0100] (4) PI index

[0101] Cell membrane composition must contain a sufficient threshold of polyunsaturated fatty acyl chains to support enzymatic and / or non-enzymatic lipid peroxidation. The peroxidizability of polyunsaturated fatty acids (PUFAs) is linearly related to the number of diallyl positions present in the molecule. The sensitivity of cell membranes to lipid peroxidation can be estimated using a peroxidizability index (PI), which is calculated from the measured fatty acid composition (%, w / w) as follows: PI = (% dienoic acid x 1) + (% trienoic acid x 2) + (% tetraenoic acid x 3) + (% pentaenoic acid x 4) + (% hexaenoic acid x 5). Alternatively, PI can be calculated as: PI = (% monoenoic acid x 0.025) + (% dienoic acid x 1) + (% trienoic acid x 2) + (% tetraenoic acid x 4) + (pentaenoic acid x 6) + (hexaenoic acid x 8). Lipidomic measurements of cell membrane composition are used to determine the peroxidizability index. Cell lines with low PI values ​​(<50) are less sensitive to perturbations that induce ferroptosis (e.g., GPX4 inhibition, GSH depletion, addition of pro-oxidant compounds). As membrane PI values ​​increase, cells are more susceptible to ferroptosis.

[0102] Cells grown in vitro have a different fatty acid profile and lower PI levels than cells in vivo. Vertebrate cells cannot synthesize PUFAs de novo and rely on dietary sources for such molecules. Typical cell culture methods use culture media supplemented with serum (usually 10%, v / v), which is the only source of exogenous lipids and contains 1% of the PUFAs available to cells in vivo. Therefore, cells grown in culture have half the PUFA levels of cells in vivo and twice the amount of monounsaturated fatty acids (MUFAs).

[0103] The ferroptosis sensitivity of cell lines can be modulated by including fatty acids in the culture medium. Saturated fatty acids (SFA), monounsaturated fatty acids (MUFA), and deuterated PUFA protect cells from ferroptosis, while the addition of PUFA increases the sensitivity of cells to perturbations that induce ferroptosis. Supplementing cell culture media with exogenous PUFA can mimic in vivo PUFA concentrations and induce membrane compositions with higher PI values. Regulatory profiling assays using fatty acid supplementation and ferroptosis inducers allow experimental determination of specific membrane PUFA content and PI values ​​required for ferroptosis in a given cell line. For example, due to the preferential uptake of PUFA by sarcomas and other cancer cells, their peroxidation index (PI) is greater than that of non-malignant tissues. Many sarcomas preferentially take up PUFAs and incorporate polyunsaturated fatty acyl chains into membrane lipids, resulting in higher membrane peroxidation index values ​​(PI>100) relative to non-malignant tissues (average PI=91). Differences in membrane peroxidation provide a therapeutic window for ferroptosis induction to selectively target sarcoma cells relative to non-malignant tissues. The more peroxidative membrane state is consistent with the observation of higher levels of lipid peroxidative stress in primary bone and soft tissue sarcomas. Exogenous PUFA addition can increase oxidative stress in osteogenic sarcoma cells and exhibit selective cytotoxic effects.

[0104] Provided herein is a method for identifying ferroptosis-sensitive cells in mammalian tissues by peroxidation index (PI). In certain embodiments, the method provided herein comprises administering an effective amount of an ferroptosis-inducing agent to a mammal, wherein the PI of multiple cells of a mammalian tissue is greater than the PI in cells of a normal or healthy tissue; and inducing ferroptosis in the multiple cells. In certain embodiments, the PI in multiple cells of a mammalian tissue is greater than a predetermined PI. In certain embodiments, the predetermined PI is about 50, 60, 70, 80, 90, 100, 110, 120, 130, 140, or 150. In certain embodiments, the predetermined PI is about 90. In certain embodiments, the PI in multiple cells of a mammalian tissue is greater than the PI in cells of a healthy or non-malignant tissue by about 1%-10%, 10%-20%, 20%-30%, 30%-40%, 40%-50%, 50%-60%, 60%-70%, 70%-80%, 80%-90%, or 90%-100%.

[0105] (5) Mesenchymal cell status

[0106] Treatment-resistant cells have three cell characteristics of high mesenchymal state and patient-derived characteristics. The first cell characteristic is the expression of mesenchymal cell markers. Ferropyrosis-sensitive cells exhibit one or more markers of mesenchymal cell state. Mesenchymal cell state markers that can be used to identify ferroptosis-sensitive cells include, but are not limited to: ZEB1, ACSL4, FADS2, PPARγ, Fsp1, SLC7A11, SLC3A2 and LPCAT3. The second cell characteristic of ferroptosis-sensitive cells is the reduced expression of endothelial cell markers compared to normal cells. Non-limiting examples of endothelial cell markers include: vimentin, E-cadherin and β (β)-actin. The third cell characteristic of ferroptosis-sensitive cells is sensitivity to GPX4 knockdown, resulting in cell death. Compared with normal mesenchymal cell lines, GPX4 dependence is more obvious in cancer cells that present a treatment-resistant mesenchymal state. Methods for reducing or silencing GPX4 expression can be achieved, for example, by CRISPR / Cas9, siRNA or shRNA.

[0107] Provided herein is a method for identifying ferroptosis-sensitive cells in mammalian tissues by the expression of one or more mesenchymal cell state markers. In certain embodiments, the method provided herein comprises administering an effective amount of an ferroptosis-inducing agent to a mammal, wherein multiple cells of a mammalian tissue express one or more markers of a mesenchymal cell state; and inducing ferroptosis in the multiple cells. In certain embodiments, the expression of mesenchymal cell markers in multiple cells of a mammalian tissue is greater than the expression of mesenchymal cell markers in cells of healthy or non-malignant tissues by about 1%-10%, 10%-20%, 20%-30%, 30%-40%, 40%-50%, 50%-60%, 60%-70%, 70%-80%, 80%-90% or 90%-100%.

[0108] (6) Other morphological characteristics of ferroptosis

[0109] Cells undergoing ferroptosis are morphologically characterized by the presence of smaller-than-normal mitochondria with condensed mitochondrial membrane density, reduced or absent mitochondrial cristae, and ruptured mitochondrial outer membranes. Histology and immunoassays can be used to determine whether a tissue is cancerous, whether it exhibits hyperplasia or fibrosis, and to identify ferroptosis-sensitive cells within mammalian tissues. The cell membranes of cells in a ferroptotic state lack the ruptures and blebbing of the plasma membrane that are typically associated with apoptosis. The nuclear size of ferroptotic cells is normal and chromatin condensation is lacking.

[0110] In certain embodiments, the method provided herein includes the step of obtaining a biological sample (e.g., a blood sample or a tissue biopsy sample) from a subject. In certain embodiments, the method provided herein further includes fixing, processing, embedding, slicing and staining biological samples for histological analysis. In certain embodiments, the tissue comprises histological abnormalities. In certain embodiments, the histological abnormalities are determined by tissue biopsy before or during the targeted continuous administration of an ferroptosis inducing agent to the tissue. In certain embodiments, the histological abnormalities are hyperplasia, angiogenesis / angiogenesis or fibrosis. Hyperplasia is identified by the number of cells increased in the tissue compared to normal healthy tissue. Angiogenesis and angiogenesis in tissue samples are identified by immunoassays for vascular markers (e.g., vascular endothelial growth factor (VEGF) and angiopoietin-2 (Ang2)). Fibrosis is characterized by abnormal collagen deposition between cells identified in tissue samples, such as by Masson's trichrome, picrosirius red or collagen staining.

[0111] Cell Death and Ferroptosis Inducers

[0112] Provided herein are methods of inducing ferroptosis in a tissue in a subject, wherein the method comprises: (a) continuously administering a therapeutic amount of a ferroptosis inducing agent; (b) contacting the in vivo tissue with an effective amount of an iron-dependent cell death agent for a period of time; and / or (c) contacting a mammalian tissue with the initiator and then contacting the in vivo mammalian tissue with an effective amount of a ferroptosis inducing agent for a period of time. Exemplary targets in the ferroptosis pathway are provided in Figure 1 middle.

[0113] The methods provided herein include administering to cells, tissues, or subjects an agent that regulates cell death. In certain embodiments, the administration induces cell death. In certain embodiments, the administration inhibits cell death or saves cells from cell death. In certain embodiments, the administration regulates ferroptosis. In certain embodiments, the administration induces ferroptosis in vivo. In certain embodiments, the administration inhibits ferroptosis in vivo. In certain embodiments, the agent is an inducer of ferroptosis. In certain embodiments, the agent is an inducer of cell death that is iron-dependent. Agents that can be used to induce ferroptosis in vivo and for treating diseases or disorders are discussed in further detail below.

[0114] (1) Ferroptosis regulators

[0115] Provided herein is an agent for regulating ferroptosis in a subject. In certain embodiments, the agent is a small molecule, a peptide or a nucleic acid. In certain embodiments, the ferroptosis regulator is a glutathione peroxidase 4 (GPX4), a glutathione synthetase, a glutamate-cysteine ​​ligase, a phosphoseryl-TRNA kinase (PSTK), a eukaryotic elongation factor selenocysteine-TRNA specificity (EEFSEC), a selenophosphate synthetase 2 (SEPHS2), a Sep (O-phosphoserine) TRNA: an inhibitor of a Sec (selenocysteine) TRNA synthase (SEPSECS) or a SECIS binding protein 2 (SECISBP2).

[0116] In certain embodiments, the agent is an inhibitor of glutathione peroxidase 4 (GPX4). Glutathione peroxidase 4 (GPX4), also known as MCSP, SMDS, GPx-4, PHGPx, snGPx, GSHPx-4, snPHGPx, belongs to the glutathione peroxidase family, whose members catalyze the reduction of hydrogen peroxide, organic hydroperoxides and lipid hydroperoxides, and thereby protect cells from oxidative damage. GPX4 activation directly reduces the level of phospholipid hydroperoxides in the cell membrane. There are several isozymes of this gene family in vertebrates, which vary in cellular location and substrate specificity. GPX4 has a high preference for lipid hydroperoxides and protects cells from membrane lipid peroxidation and cell death. This isozyme is also a selenoprotein containing the rare amino acid selenocysteine ​​(Sec) at its active site. Representative human GPX4 cDNA and human GPX4 protein sequences are publicly available from the National Center for Biotechnology Information (NCBI). Human glutathione peroxidase 4 peroxidase isoform B precursor (NM_001039847.3 and NP_001034936.1), isoform C (NM_001039848.4 and NP_001034937.1), isoform D (NM_001367832.1 and NP_001354761.1), isoform A precursor (NM_002085.5 and NP_002076.2).

[0117] Depletion of GPX4 can induce lipid peroxidation-dependent cell death. Cancer cells in drug-induced treatment resistance states have an enhanced dependence on the lipid peroxidase activity of GPX4 to prevent ferroptosis cell death. Lipophilic antioxidants such as ferrostatin can save cells from ferroptosis induced by GPX4 inhibition. For example, cells knocked out of mesenchymal state GPX4- can survive in the presence of ferrostatin, but when the supply of ferrostatin is terminated, these cells will experience ferroptosis. GPX4 inhibition can be rescued by blocking other components of the ferroptosis pathway, such as lipid ROS scavengers (ferrostatin, liproxstatin), lipoxygenase inhibitors, iron chelators and caspase inhibitors, while apoptosis inhibitors will not rescue. Therefore, GPX4 inhibitors can be used to induce ferroptosis cell death.

[0118] In certain embodiments, the agent is an inhibitor of glutathione synthetase (GSS). Glutathione synthetase (GSS), also known as GSHS, HEL-S-64p, HEL-S-88n, is a homodimer that catalyzes the second step of glutathione biosynthesis, i.e., the ATP-dependent conversion of γ-L-glutamyl-L-cysteine ​​to glutathione. Representative human GSScDNA and human GSS protein sequences can be publicly obtained from the National Center for Biotechnology Information (NCBI). Human glutathione synthetase (NM_000178.4 and NP_000169.1, NM_001322494.1 and NP_001309423.1, NM_001322495.1 and NP_001309424.1).

[0119] In certain embodiments, the agent is an inhibitor of glutamate-cysteine ​​ligase (GCL). Glutamate-cysteine ​​ligase (GCL), a central node of the ferroptosis pathway, has been overlooked as a target. The loss of GCL activity induces ferroptosis in sensitive cells and kills only the cells that are most sensitive to ferroptosis. Representative human GCL cDNA and human GCL protein sequences are publicly available from the National Center for Biotechnology Information (NCBI). Human glutamate-cysteine ​​ligase catalytic subunit isoform b (NM_001197115.2 and NP_001184044.1, compared to variant 1, it lacks an in-frame exon in the 5' coding region. This produces a shorter protein (isoform b) compared to isoform a) and glutamate-cysteine ​​ligase catalytic subunit isoform a (NM_001498.4 and NP_001489.1, which represent longer transcripts and encode longer isoforms (a)).

[0120] In certain embodiments, the agent is an inhibitor of phospho-seryl-tRNA kinase (PSTK). PSTK is an enzyme that recruits selenocysteine, encoded by UGA. Sec is formed in the tRNA-dependent conversion of serine attached to tRNA Sec by seryl-tRNA synthetase. PSTK phosphorylates Ser-tRNA Sec to Sep-tRNA Sec, which is then converted into Sec-tRNA Sec by Sep-tRNA:Sec-tRNA synthase (SepSecS). Representative human PSTK cDNA and human PSTK protein sequences are well known in the art and can be publicly obtained from the National Center for Biotechnology Information (NCBI). Human L-seryl-tRNA(Sec) kinase isoform 1 (NM_001363531.2 and NP_001350460.1) and L-seryl-tRNA(Sec) kinase isoform 2 (NM_153336.3 and NP_699167.2).

[0121] In certain embodiments, the inhibitor is an inhibitor of eukaryotic elongation factor selenocysteine-TRNA specific (EEFSEC). EEFSEC is also known as selenoprotein translation factor selb. Representative human EEFSEC cDNA and human EEFSEC protein sequences are publicly available from the National Center for Biotechnology Information (NCBI) as follows: selenocysteine-specific elongation factor (NM_021937.5 and NP_068756.2), selenocysteine-specific elongation factor isoform X4 (XM_024453695.1 and XP_024309463.1), selenocysteine-specific elongation factor isoform X3 (XM_024453694.1 and XP_024309462.1), selenocysteine-specific elongation factor isoform X1 (XM_024453692.1 and XP_024309463.1), selenocysteine-specific elongation factor isoform X2 (XM_024453693.1 and XP_024309464.1), selenocysteine-specific elongation factor isoform X3 (XM_024453694.1 and XP_024309465.1), selenocysteine-specific elongation factor isoform X4 (XM_024453695.1 and XP_024309465.1), selenocysteine-specific elongation factor isoform X3 (XM_024453694.1 and XP_0243094667.1), selenocysteine-specific elongation factor isoform X1 (XM_024453693.1 and XP_0243094667.1), seleno EEFSEC is a specialized translation elongation factor that is responsible for the co-translational incorporation of selenocysteine ​​into proteins by recoding of the UGA stop codon in the presence of a downstream mRNA hairpin loop.

[0122] In certain embodiments, the agent is an inhibitor of selenophosphate synthase 2 (SEPHS2). Selenophosphate synthase 2 (SEPHS2) catalyzes the generation of monoselenophosphate (MSP) from selenide and ATP. MSP is a selenium donor required for the synthesis of selenocysteine ​​(Sec), which is co-translationally incorporated into selenoproteins at the UGA codon in the frame that usually signals the termination of translation. The protein itself is a selenoprotein containing a Sec residue at its active site, indicating the existence of an autoregulatory mechanism. SEPHS2 is preferentially expressed in tissues related to the synthesis of selenoproteins and in blood cell development sites. In addition, genome-scale cancer dependency mapping analysis identifies selenoprotein synthases as targets for inducing ferroptosis. The loss of selenoprotein synthases induces ferroptosis in sensitive cells. In addition, the loss of selenophosphate synthase 2 (SEPHS2) exhibits a novel two-pronged mechanism of ferroptosis. The loss of SEPHS2 induces ferroptosis much faster than the loss of other selenoprotein biosynthetic enzymes. SEPHS2 inhibitors can induce ferroptosis in certain diseases. For example, aggressive liver cancer can be selectively targeted by SEPHS2 inhibition. Representative human SEPHS2 cDNA and human SEPHS2 protein sequences are well known in the art and publicly available from the National Center for Biotechnology Information (NCBI). Selenide, water dikinase 2 (NM_012248.4 and NP_036380.2).

[0123] In certain embodiments, the agent is an inhibitor of Sep(O-phosphoserine)TRNA:Sec(selenocysteine)TRNA synthase (SEPSECS). Sep(O-phosphoserine)TRNA:Sec(selenocysteine)TRNA synthase (SEPSECS) catalyzes the third step in the synthesis of selenocysteine, namely the conversion of O-phosphoseryl-tRNA(Sec) to selenocysteinyl-tRNA(Sec). Representative human SEPSECS cDNA and human SEPSECS protein sequences are publicly available from the National Center for Biotechnology Information (NCBI) as follows: O-phosphoseryl-tRNA(Sec)selenotransferase (NM_016955.4 and NP_058651.3), O-phosphoseryl-tRNA(Sec)selenotransferase isoform X1 (XM_017008277.1 and XP_016863766.1), O-phosphoseryl-tRNA(Sec)selenotransferase isoform X5 (XM_017008278.1 and XP_016863766.1). XP_016863767.1), O-phosphoseryl-tRNA(Sec)selenotransferase isoform X4 (XM_011513848.1 and XP_011512150.1), O-phosphoseryl-tRNA(Sec)selenotransferase isoform X2 (XM_011513846.2 and XP_011512148.1) and O-phosphoseryl-tRNA(Sec)selenotransferase isoform X3 (XM_011513847.2 and XP_011512149.1).

[0124] In certain embodiments, the agent is an inhibitor of SECIS binding protein 2 (SECISBP2). SECISBP2 is one of the polypeptide components of the mechanism involved in the co-translational insertion of selenocysteine ​​(Sec) into selenoproteins. Sec is encoded by the UGA codon, which normally signals translation termination. Recoding of UGA as a Sec codon requires a Sec insertion sequence (SECIS) element; this element is present in the 3' untranslated region of eukaryotic selenoprotein mRNA. The protein specifically binds to the SECIS element, which is stimulated by Sec-specific translation elongation factors.Representative human SECISBP2 cDNA and human SECISBP2 protein sequences are publicly available from the National Center for Biotechnology Information (NCBI) as follows: selenocysteine ​​insertion sequence-binding protein 2 isoform 2 (NM_001282688.2 and NP_001269617.1), selenocysteine ​​insertion sequence-binding protein 2 isoform 3 (NM_001282689.2 and NP_001269618.1), selenocysteine ​​insertion sequence-binding protein 2 isoform 4 (NM_001282690.1 and NP_001269619.1), selenocysteine ​​insertion sequence-binding protein 2 isoform 5 (NM_001354696.2 and NP_001341696.1). 25.1), selenocysteine ​​insertion sequence-binding protein 2 isoform 6 (NM_001354697.2 and NP_001341626.1), selenocysteine ​​insertion sequence-binding protein 2 isoform 7 (NM_001354698.2 and NP_001341627.1), selenocysteine ​​insertion sequence-binding protein 2 isoform 8 (NM_001354702.2 and NP_001341631.1), selenocysteine ​​insertion sequence-binding protein 2 isoform 1 (NM_024077.5 and NP_076982.3), selenocysteine ​​insertion sequence-binding protein 2 isoform X9 (XM_024447669.1 and X P_024303437.1), selenocysteine ​​insertion sequence-binding protein 2 isoform X8 (XM_024447667.1 and XP_024303435.1), selenocysteine ​​insertion sequence-binding protein 2 isoform X5 (XM_017015122.2 and XP_016870611.1), selenocysteine ​​insertion sequence-binding protein 2 isoform X6 (XM_024447666.1 and XP_024303434.1), selenocysteine ​​insertion sequence-binding protein 2 isoform X9 (XM_024447668.1 and XP_024303436.1), selenocysteine ​​insertion sequence-binding protein 2 isoform X1 (XM_011519000.2 and XP_011517302.1), selenocysteine ​​insertion sequence-binding protein 2 isoform X2 (XM_011519001.1 and XP_011517303.1), selenocysteine ​​insertion sequence-binding protein 2 isoform X3 (XM_011519002.1 and XP_011517304.1), selenocysteine ​​insertion sequence-binding protein 2 isoform X4 (XM_011519003.1 and XP_011517305.1) and selenocysteine ​​insertion sequence-binding protein 2 isoform X7 (XM_006717282.2 and XP_006717345.1).

[0125] In certain embodiments, the agent is an inhibitor of nuclear factor-erythroid factor 2-related factor 2 (NRF2). NRF2 is a member of the cap 'n' neck (CNC) subfamily of basic region leucine zipper (bZip) transcription factors. NRF2 mediates the induction of a range of drug metabolizing enzymes such as glutathione S-transferase (GST) and NAD(P)H:quinone oxidoreductase 1 (NQO1) by antioxidants and electrophiles. NRF2 also regulates GPX4 protein levels, intracellular free iron levels, and mitochondrial function, thereby regulating ferroptosis. NRF2 protein sequences are publicly available from the National Center for Biotechnology Information (NCBI) as follows: Nrf2 [Homo sapiens], GenBank: AAB32188.1; and transcription factor Nrf2-human, PIR: I59340.

[0126] In certain embodiments, the agent is an inhibitor of the cystine transporter SLC7A11 (also known as xCT). SLC7A11 (also commonly referred to as xCT) functions to import cystine for glutathione biosynthesis and antioxidant defense and is overexpressed in a variety of human cancers. The SLC7A11 (xCT) protein sequence is publicly available from the National Center for Biotechnology Information (NCBI) as follows: cystine / glutamate transporter [Homo sapiens] NP_055146.1 and cystine / glutamate transporter isoform X1 [Homo sapiens] XP_011530104.1.

[0127] In certain embodiments, the agent is an inhibitor of system Xc. System Xc-also known as the cystine / glutamate antiporter, is an intracellular antioxidant element composed of the light chain SLC7A11 (xCT) and the heavy chain SLC3A2 (4F2hc), and functions as a raw material for the synthesis of glutathione (GSH). System Xc protein sequences are publicly available from the National Center for Biotechnology Information (NCBI) as follows: cystine / glutamate transporter [Homo sapiens] NP_055146.1, cystine / glutamate transporter isoform X1 [Homo sapiens] XP_011530104.1, 4F2 cell-surface antigen heavy chain isoform f [Homo sapiens] NP_001013269.1, 4F2 cell-surface antigen heavy chain isoform c [Homo sapiens] NP_002385.3, 4F2 cell-surface antigen heavy chain isoform b [Homo sapiens] NP_001012680.1, and 4F2 cell-surface antigen heavy chain isoform e [Homo sapiens] NP_001012682.1.

[0128] In certain embodiments, the agent is an inhibitor of thioredoxin reductase (TXNRD). TRXNRD is involved in the reversible S-nitrosylation of cysteine ​​in certain proteins. TRXNRD protein sequences are publicly available from the National Center for Biotechnology Information (NCBI) as follows: thioredoxin reductase [Homo sapiens] AAB35418.1, thioredoxin reductase [Homo sapiens] AAF15900.1 GI: 6538774, thioredoxin reductase [Homo sapiens] AAD25167.1, thioredoxin reductase [Homo sapiens] AAD19597.1, and thioredoxin reductase [Homo sapiens] CAA04503.1.

[0129] In certain embodiments, the ferroptosis-inducing agent is a class of molecules that includes at least one of the following (for molecules in this class): a salt, a pharmaceutically acceptable salt, a solvate, a hydrate, an enantiomer, a diastereomer, a racemate, a crystalline form, or any combination of these of the molecule.

[0130] In certain embodiments, the initiator is a class of molecules that includes at least one of the following (for molecules in that class): a salt, a pharmaceutically acceptable salt, a solvate, a hydrate, an enantiomer, a diastereomer, a racemate, a crystalline form, or any combination of these of the molecule.

[0131] In certain embodiments, the agent is a statin. Exemplary statins include, but are not limited to, atorvastatin, cerivastatin, fluvastatin, lovastatin, mevastatin, pitavastatin, pravastatin, rosuvastatin, and simvastatin.

[0132] In certain embodiments, the agent that induces or modulates tissue or systemic ferroptosis is selected from Table 1. Table 1 provides exemplary ferroptosis modulators and their formulas, chemical identifiers, and respective targets and / or mechanisms of action.

[0133] Table 1. Ferroptosis regulators

[0134]

[0135]

[0136]

[0137]

[0138]

[0139]

[0140]

[0141]

[0142]

[0143]

[0144]

[0145]

[0146]

[0147]

[0148]

[0149]

[0150]

[0151]

[0152]

[0153]

[0154]

[0155]

[0156]

[0157]

[0158]

[0159] In certain embodiments, the ferroptosis regulator is selected from: (1S, 3R)-RSL3, ML-162, ML-210, JKE-1674, JKE-1716, elastin, jacaric acid, buthionine sulfoximine (BSO), trigonelline, glutamate / glutamate / glutamate ester (glutamate), sulfasalazine, auranofin, brucein, sorafenib, sorafenib-d3, sorafenib tosylate, trigonelline, FIN56, FINO2, CIL56, dihydroisotanshinone I, GPX4-IN-3, its analogs, salts or derivatives. In certain embodiments, the agent in Table 1 is a pharmaceutically acceptable salt form of the small molecule.

[0160] (2) Initiator

[0161] Provided herein is a method for inducing targeted cell death in a mammalian tissue in vivo, the method comprising: (a) contacting the mammalian tissue with the initiator; (b) contacting the mammalian tissue in vivo with an effective amount of an inducer of ferroptosis for a period of at least 4 hours when a plurality of cells within the mammalian tissue are responsive to the initiator, wherein the responsiveness is determined by detecting the following items in the mammalian tissue: (i) a plurality of cells comprising a selenium concentration greater than the selenium concentration in the mammalian tissue prior to contact with the initiator; (ii) a plurality of cells comprising an iron concentration greater than the iron concentration in the mammalian tissue prior to contact with the initiator; (iii) a plurality of cells comprising a PUFA concentration greater than the PUFA concentration in the mammalian tissue prior to contact with the initiator; (iv) a plurality of cells expressing one or more markers indicative of a mesenchymal state; (v) a plurality of cells comprising a peroxidation index (PI) greater than the PI in the mammalian tissue prior to contact with the initiator; and / or (vi) excessive proliferation of cells in the mammalian tissue, wherein the inducer of ferroptosis induces targeted cell death in the mammalian tissue in vivo.

[0162] In certain embodiments, an initiator is administered before administering an ferroptosis inducer provided herein. In certain embodiments, the initiator is administered in vivo, in vitro, or ex vivo. An initiator is an agent that prepares a subject or tissue for administration of a therapeutically effective dose of an ferroptosis inducer or ferroptosis regulator provided herein. In certain embodiments, the initiator is an inhibitor of ferroptosis. In certain embodiments, the initiator makes cells in a tissue sensitive to ferroptosis. In certain embodiments, the initiator is a lipophilic antioxidant or a free radical trap. In certain embodiments, the initiator is a polyunsaturated fatty acid. In certain embodiments, the initiator is an iron chelator. In certain embodiments, the initiator is a lipid peroxidation inhibitor. In certain embodiments, the initiator regulates blood oxygen levels. In certain embodiments, the initiator is a hydroperoxide. In certain embodiments, the initiator is selected from the group consisting of: liproxstatin-1, ferrostatin-1, deferoxamine (DFO), iron, selenium, vitamin E, erythropoietin, polyunsaturated fatty acids, N-acetylcysteine, pifithrin-α-HBr, and methylnaphthalene-4-propionate endoperoxide (MNPE). In certain embodiments, the polyunsaturated fatty acid is selected from the group consisting of hexadecatrienoic acid (HTA), alpha-linolenic acid (ALA), stearidonic acid (SDA), eicosatrienoic acid (ETE), eicosatetraenoic acid (ETA), eicosapentaenoic acid (EPA, eicosapentaenoic acid), heneicosapentaenoic acid (HPA), docosapentaenoic acid (DPA, ichthyic acid), docosahexaenoic acid (DHA, ichthyic acid), tetracosahexanoic acid (herring acid), tetracospentaenoic acid, linoleic acid (LA), gamma-linolenic acid (GLA), eicosadienoic acid, dihomo-gamma-linolenic acid (DGLA), arachidonic acid (AA), docosadienoic acid, adrenic acid (AdA), docosapentaenoic acid (osbond acid), tetracosatetraenoic acid and tetracospentaenoic acid. Table 2 provides non-limiting examples of initiators.

[0163] Table 2. Initiators.

[0164]

[0165]

[0166]

[0167]

[0168]

[0169] In certain embodiments, the methods provided herein comprise administering any one of the agents listed in Table 2. Further provided herein are pharmaceutical compositions, wherein the composition comprises a ferroptosis inducer or a ferroptosis regulator and an initiator. In certain embodiments, the pharmaceutical composition further comprises a chemotherapeutic agent.

[0170] In certain embodiments are compositions for use in methods for reducing the number of adipocytes in a subject in need thereof. In certain embodiments are compositions for use in regulating, adjusting or inducing weight loss or fat loss. In certain embodiments are compositions for use in regulating or adjusting iron metabolism. In certain embodiments are compositions for use in treating obesity. In certain embodiments are compositions for use in treating kidney disease. In certain embodiments are compositions for use in treating kidney disease. In certain embodiments are compositions for use in treating diseases or conditions associated with iron metabolism.

[0171] In certain embodiments are compositions comprising an agent in Table 1 for use in the preparation of a medicament for treating a disease or condition described herein such as a kidney disease or kidney condition. In certain embodiments are compositions comprising an agent in Table 1 for use in the preparation of a medicament for treating elevated cholesterol. In certain embodiments are compositions comprising an agent in Table 1 for use in the preparation of a medicament for inducing ferroptosis in a subject. In certain embodiments are compositions comprising an inducer of ferroptosis for use in the preparation of a medicament for treating a kidney disease, a kidney condition, a disease or condition associated with fat metabolism, or a disease or condition associated with elevated cholesterol.

[0172] (3) Other treatments

[0173] In certain embodiments, the methods provided herein comprise administering to the subject at least one additional treatment (e.g., a second weight loss therapy). In some cases, the second weight therapy is selected from: GLP-1 agonists, gastric inhibitory polypeptide analogs, naltrexone-bupropion or a salt thereof, orlistat or a salt thereof, phentermine or a salt thereof and phentermine-topiramate or a salt thereof, selenium, exercise therapy, surgery, dietary modification, and any combination thereof. In some cases, the GLP-1 agonist comprises dulaglutide, exenatide, semaglutide, liraglutide, lixisenatide, or a salt of any of these. In some cases, the gastric inhibitory polypeptide comprises tirpotide or a derivative thereof. In some cases, the surgery is a gastric sleeve, a gastric bypass, an adjustable gastric band, or any combination thereof. In some cases, selenium may comprise inorganic selenium (selenate and selenite) organic selenium (selenomethionine and selenocysteine) or any combination thereof.

[0174] In certain embodiments, the methods provided herein include administering at least one additional treatment to a subject. In certain embodiments, the additional treatment is surgery. In certain embodiments, the additional treatment is radiation therapy. In certain embodiments, the additional treatment is a dietary supplement. Non-limiting examples of dietary supplements include: probiotics, selenium, iron, vitamins (e.g., vitamin A, vitamin C, vitamin E), curcumin, fish oil, beta carotene, hydrogen sulfide, fatty acids, methionine, cysteine, homocysteine, taurine, cystine or di-cysteine. In certain embodiments, the dietary supplement is a high-selenium nutritional supplement.

[0175] In certain embodiments, the additional treatment is an additional therapeutic agent. In certain embodiments, the methods provided herein include administering an additional agent in combination with an ferroptosis inducer, an iron-dependent cell death inducer and / or an initiator provided herein. In certain embodiments, the additional agent is a cell death inducer. In certain embodiments, the additional agent is an anticancer agent. In certain embodiments, the anticancer agent is a chemotherapeutic agent. Chemotherapeutic agents or compounds are any agents or compounds that can be used to treat cancer. Cancer chemotherapeutics that can be used in combination with ferroptosis inducers or iron-dependent cell death agents provided herein include, but are not limited to, mitotic inhibitors (vinca alkaloids). These include vincristine, vinblastine, vindesine and novelbine TM (Vinorelbine, 5'-noranhydroblastine). In other cases, cancer chemotherapeutic agents include topoisomerase I inhibitors, such as camptothecin compounds. "Camptothecin compounds" as used herein include Camptosar TM (irinotecan hydrochloride) and Hycamtin TM(topotecan hydrochloride) and other compounds derived from camptothecin and its analogs. Another class of cancer chemotherapeutics that can be used in the methods and compositions disclosed herein are podophyllotoxin derivatives, such as etoposide, teniposide and mitozazide. The present disclosure further encompasses other cancer chemotherapeutics known as alkylating agents, which alkylate the genetic material in tumor cells. These include, but are not limited to, cisplatin, cyclophosphamide, nitrogen mustard, trimethylenethiophosphoramide, carmustine, busulfan, chlorambucil, belustine, uracil mustard, chlomaphazin and dacarbazine. The present disclosure encompasses antimetabolites as chemotherapeutic agents. Examples of these types of agents include cytosine arabinoside, fluorouracil, methotrexate, mercaptopurine, azathioprine and procarbazine. Another class of cancer chemotherapeutics that can be used in the methods and compositions disclosed herein include antibiotics. Examples include, but are not limited to, doxorubicin, bleomycin, dactinomycin, daunorubicin, mithramycin, mitomycin, cytomycin C and daunomycin. There are many commercially available liposomal formulations of these compounds. The present disclosure further contemplates other cancer chemotherapeutic agents, including, but not limited to, anti-tumor antibodies, dacarbazine, azacytidine, amsacrine, melphalan, ifosfamide, and mitoxantrone.

[0176] The disclosed medicaments provided herein can be co-administered with other anti-tumor agents, which include cytotoxic agents / antitumor agents and anti-angiogenic agents. Cytotoxic agents / antitumor agents can be defined as agents that attack and kill cancer cells. Some cytotoxic agents / antitumor agents can be alkylating agents that alkylate the genetic material in tumor cells, for example, cisplatin, cyclophosphamide, nitrogen mustard, trimethylene thiophosphoramide, carmustine, busulfan, chlorambucil, belustine, uracil nitrogen mustard, chlomaphazin and dacarbazine. Other cytotoxic agents / antitumor agents can be antimetabolites of tumor cells, for example, cytosine arabinoside, fluorouracil, methotrexate, mercaptopurine, thiopurine and procarbazine. Other cytotoxic agents / antitumor agents can be antibiotics, for example, doxorubicin, bleomycin, dactinomycin, daunorubicin, mithramycin, mitomycin, cytomycin C and daunomycin. There are numerous commercially available liposome preparations for these compounds. Still other cytotoxic / antitumor agents may be inhibitors of mitosis (vinca alkaloids). These include vincristine, vinblastine and etoposide. Various cytotoxic / antitumor agents include taxol and its derivatives, L-asparaginase, antitumor antibodies, dacarbazine, azacytidine, amsacrine, melphalan, VM-26, ifosfamide, mitoxantrone and vindesine.

[0177] Anti-angiogenic agents may also be used. Suitable anti-angiogenic agents for use in the disclosed methods and compositions include anti-VEGF antibodies, including humanized and chimeric antibodies, anti-VEGF aptamers and antisense oligonucleotides. Other angiogenesis inhibitors include angiostatin, endostatin, interferon, interleukin 1 (including α and β) interleukin 12, retinoic acid, and tissue inhibitors of metalloproteinases-1 and -2 (TIMP-1 and -2). Small molecules may also be used, including topoisomerases such as razoxane, a topoisomerase II inhibitor with anti-angiogenic activity.

[0178] Other anticancer agents that can be used in combination with the ferroptosis inducing agents provided herein can include, but are not limited to: acivicin; aclarubicin; acodazole hydrochloride; aclonine; adolesin; aldesleukin; hexamethylmelamine; ambomycin; ametrine acetate; aminoglutethimide; amsacrine; anastrozole; anthramycin; asparaginase; triclinicin; avastin; azacitidine; azadepa; azomycin; batimastat; benzodepa; bevacizumab; Carutamide; bisantrene hydrochloride; binefad dimesylate; bisoprolol; bleomycin sulfate; buquina sodium; bropiridamine; busulfan; actinomycin C; captestosterone; caracetamide; carbetim; carboplatin; carmustine; carrubicin hydrochloride; cazelexin; cedifingol; chlorambucil; sirolimus; cisplatin; cladribine; clinatoyl mesylate; cyclophosphamide; cytarabine; dacarbazine; dactinomycin; daunorubicin hydrochloride; decitabine; dextromethorphan Maplatin; Dezaguanine; Dezaguanine mesylate; Diazanone; Docetaxel; Doxorubicin; Doxorubicin hydrochloride; Droloxifene; Droloxifene citrate; Drostanone propionate; Dazomycin; Edatrexate; Eflornithine hydrochloride; Elsamirucin; Enloplatin; Enprobamate; Epipiperidone; Epirubicin hydrochloride; Erbulazole; Esorubicin hydrochloride; Estramustine; Estramustine sodium phosphate; Etanidazole; Etoposide; Etoposide phosphate; Chlorpheniramine; Hydrochloride Fadrozole; fazarabine; fenretinide; floxuridine; fludarabine phosphate; fluorouracil; flucitabine; folinic acid; fosquatone; fostramoxine sodium; gemcitabine; gemcitabine hydrochloride; hydroxyurea; idarubicin hydrochloride; ifosfamide; imofosine; interleukin II (including recombinant interleukin II or rIL2), interferon α-2a; interferon α-2b; interferon α-n1; interferon α-n3; interferon β-Ia; interferon γ-I b; iproplatin; irinotecan hydrochloride; lanreotide acetate; letrozole; leuprolide acetate; lirazole hydrochloride; lometrexol sodium; lomustine; losoxantrone hydrochloride; masoprodol; maytansine (maytansine); nitrogen mustard hydrochloride; megestrol acetate; melengestrol acetate; melphalan; menolipir; mercaptopurine; methotrexate; methotrexate sodium; chlorpheniramine; metodepa; mibutidamide; mitokacin; mitoglobin; mitoxantrone; mitomycin; mitospe; mitotane; mitoxantrone hydrochloride; mycophenolic acid; nocodazole; noramycin; ormaplatin; oxysulen; paclitaxel; pepstatin Asparagase; Pelimycin; Nitrogen mustard pentahydrochloride; Pelimycin sulfate; Perfosfamide; Pipobroman; Piposufan; Pyroxantrone hydrochloride; Plicamycin; Promestane; Porfimer sodium; Porfiromycin; Prednimustine; Procarbazine hydrochloride; Puromycin; Puromycin hydrochloride; Pyrazofurine; Liboadenosine; Roglulimide; Safingol; Safingol hydrochloride; Semustine; Simtrazine; Sodium phosphoacetylaspartate; Sparamycin; Spirogermanium hydrochloride; Spiromustine; Spiroplatin; Streptozocin; Streptozocin; Sulfonyl urea; Talithromycin; Tecogalan sodium; Tegafur; Tiloxantrone hydrochloride; Temoporfin; Teniposide;Tiroxilon; testolactone; thioimidone; thioguanine; thiotepa; thiazole carboxamide nucleoside; tirapazamine; toremifene citrate; triptolon acetate; triciribine phosphate; trimetrexate; trimetrexate glucuronide; triptorelin; tobradazole hydrochloride; uracil mustard; uredepa; vapreotide; verteporfin; vinblastine sulfate; vincristine sulfate; vindesine; vindesine sulfate; vinpitidine sulfate; vinca glyceride sulfate; vinrosin sulfate; vinorelbine tartrate; vinprodine sulfate; vinorelbine sulfate; vorozole; zeniplatin; ninstatin; zorubicin hydrochloride. Other anticancer agents include, but are not limited to: 20-epi-1,25 dihydroxyvitamin D3; 5-ethynyluracil; abiraterone; aclarubicin; acylfulvene; adecypenol; adolesin; aldesleukin; ALL-TK agonists or antagonists; hexamethylmelamine; aminostine; 2,4-dichlorophenoxyacetic acid (amidox); amifostine; aminolevulinic acid; amrubicin; amsacrine; anagrelide; anastrozole; andrographolide; angiogenesis inhibitors; antagonist D; antagonist G; anrelix; anti-dorsal morphogenetic protein-1; anti-androgen, prostate cancer; anti-estrogen; antineoplastons; antisense oligonucleotides; glycine aphidicolin ; apoptosis gene regulator; apoptosis regulator; apurinic nucleic acid; cytarabine DP-DL-PTBA; arginine deaminase; asulacrine; atamestane; amustine; axinastatin 1; axinastatin 2; axinastatin 3; azasetron; azatoxin; diazotyrosine; baccatin III derivative; balanol; batimastat; BCR / ABL antagonist or agonist; benzodihydrochlorin; benzoylstaurosporine; beta-lactam derivative; beta-alethine; betaclamycin B; betulinic acid; bFGF inhibitors; bicalutamide; bisantrene; bisaziridinylspermine; binefad; bistratene A; bizeresin; breflate; bropiridin; budotitan; buthionine sulfoximine; calcipotriol; calciphosphotin C; camptothecin derivatives; canarypox IL-2; capecitabine; carboxyamido-amino-triazole; carboxyamido-triazole; CaRest M3; CARN 700; cartilage-derived inhibitors; kazeresin; casein kinase inhibitor (ICOS); castanospermine; bacillin B; cetrorelix; chlorins; chloroquinoxaline sulfonamide; cicaprost; cis-porphyrin; cladribine; clomiphene analogs; clotrimazole; collismycin A; collismycin B; combretastatin A4; combretastatin analog; conagenin; crambescidin816; crenato; Nostoc cyclotide 8;A derivative of Nostoc cyclotide A; curacin A; cyclopentanthraquinone; cycloplatam; cypemycin; cytarabine octadecyl phosphate; cytolytic factor; cytostatin; dacillin; decitabine; dehydrodeoxanthin B; deslorelin; dexamethasone; ifosfamide; dexrazoxane; dexverapamil; diazocone; deoxanthin B; didox; diethylnorspermine; dihydro-5-azacytidine; dihydrotaxol, 9-; dioxamycin; diphenylspiromustine; docetaxel; docosinol; dora Setron; doxifluridine; droloxifene; dronabinol; duocarmycin SA; ebselen; ecomustine; edilfosine; edrolomab; eflornithine; elemene; ethidium bromide; epirubicin; eristeride; estramustine analogs; estrogen agonists; estrogen antagonists; etanidazole; etoposide phosphate; exemestane; fadrozoles; fazarabine; fenretinide; filgrastim; finasteride; flavopiridol; fluzolastine; flusterone; fludarabine; fluorodaunorunicin hydrochloride; fofolmek; formestane; fostramcin; fomastine; tesarin gadolinium; gallium nitrate; galocitabine; ganirelix; gelatinase inhibitors; gemcitabine; glutathione inhibitors; hepsulfam; heregulin; hexamethylenediethylamide; hyperforin; ibandronic acid; idarubicin; idoxifene; idemenone; imofosine; ilomastat; imidazoacridone; imiquimod; immunostimulatory peptides; insulin-like growth factor-1 receptor inhibitors; interferon agonists; interferons; interleukins; iodobenzylguanidine; iododexorubicin; 4-iodopyralidone; iropra; irsogladine; isobengazole; isohomohalicondrin B; itasetron; jasplakinolide; kahalalide F; lamellarin-N triacetate; lanreotide; ranamycin; lenograstim; lentinan sulfate; leptolstatin; letrozole; leukemia inhibitory factor; interferon-α; leuprolide + estrogen + progesterone; leuprolide; levamisole; liarozole; linear polyamine analogs; lipophilic disaccharide peptides; lipophilic platinum compounds; lissoclinamide 7; lobaplatin; earthworm phospholipids; lometrexol; lonidamine; losoxantrone; lovastatin; loxoribine; lutetamine; lutetamine; lysofylline; lytic peptide; metansine; ganciclovir; marimastat; masorol; mammofilin; matrix lytic factor inhibitor; matrix metalloproteinase inhibitor; menolide; mebarone; metirol; methioninase; metoclopramide; MIF inhibitor; mifepristone; miltefosine; milistostatin; mismatched double-stranded RNA; mitoguanidine; dibromodole; mitomycin analog; mitonadine; mitotoxin fibroblast growth factor-saporin; mitoxantrone; mofarotin; morasporastatin;monoclonal antibody, human chorionic gonadotropin; monophosphoryl lipid A + mycobacterial cell wall sk; mopidarol; multidrug resistance gene inhibitor; multiple tumor suppressor gene 1-based therapy; mustard anticancer agent; mycaperoxide B; mycobacterial cell wall extract; myriaporone; N-acetyl pyridazine; N-substituted benzamide; nafarelin; nagrestip; naloxone + pentazocine; napavin; naphterpin; narostem; nedaplatin; nemorubicin; neridronic acid; neutral endopeptidase; nilutamide; nisamycin; nitric oxide modulator; nitrous oxide antioxidant; nitrullyn; O6-benzylguanine; octreotide; okicenone; oligonucleotide; onapristone; ondansetron; ondansetron; oracin; oral cell Cytokine inducers; ormaplatin; oxatetron; oxaliplatin; oxaunomycin; paclitaxel; paclitaxel analogs; paclitaxel derivatives; palauamine; palmitoyl rhizoxin; pamidronate; panaxytriol; panomifen; parabactin; parazeptin; pegaspargase; perdecan; pentosan polysulfate sodium; pentostatin; pentrozole; perflurane; perfosfamide; perillyl alcohol; phenazinomycin; phenylacetate; phosphatase inhibitors; bisibanib; pilocarpine hydrochloride; pirarubicin; pirexine; placetin A; placetin B; plasminogen activator inhibitor; platinum complex; platinum compound; platinum-triamine complex; porfimer sodium; porfibrin; prednisone; propyl di-acridone; prostaglandin J2; proteasome inhibitor; protein A-based immunomodulator; protein kinase C inhibitor; microalgae protein kinase C inhibitor; protein tyrosine phosphatase inhibitor; purine nucleoside phosphorylase inhibitor; purpurin; pyrazoloacridine; pyridoxylated hemoglobin polyoxyethylene conjugate; raf antagonist; raltitrexed; ramosetron; ras farnesyl protein transferase inhibitor; ras inhibitor; ras-GAP inhibitor; demethylated retiptine; etidronate rhenium Re 186; rhizoxin; ribozyme; RII retinamide; roglulimide; roxitomine; romotide; roquimek; rubiginone B1; ruboxyl; safengo; saintopin; SarCNU; muscle phytol A; sargramostim; Sdi 1 mimetic; semustine; senescence-derived inhibitor 1; sense oligonucleotide; signal transduction inhibitor; signal transduction regulator; single-chain antigen binding protein; sizoran; sobuzosine; borcarbazine sodium; sodium phenylacetate; solverol; somatomedin binding protein; sonarmin; fospartate; spicamycin D; spiromustine; spleen pentapeptide; spongestatin 1; squalamine; stem cell inhibitor; stem cell division inhibitor;stipiamide; stromelysin inhibitor; sulfinosine; superactive vasoactive intestinal peptide antagonist; suradista; suramin; swainsonine; synthetic glycosaminoglycans; tamustine; tamoxifen methyl iodide; tauromustine; tazarotene; tecogalan sodium; tegafur; tellurapyrylium; telomerase inhibitor; temoporfin; temozolomide; teniposide; tetrachlorodecoxide; tetrazomine; thaliblastine; thiocoratine; thrombopoietin; Thrombopoietin mimetics; thymofasin; thymopoietin receptor agonists; thymosin; thyrotropin; oxazolidinone; tirapazamine; dichlorotitanium; topsentin; toremifene; totipotent stem cell factor; translation inhibitors; retinoic acid; triacetyluridine; triciribine; trimetrexate; triptorelin; tropisetron; torosterol; tyrosine kinase inhibitors; tyrosine phosphorylation inhibitors; UBC inhibitors; ubenimex; urogenital sinus-derived growth inhibitory factor; urokinase receptor agonists or antagonists; vapreotide; variolin B; erythrocyte gene therapy; vilaresol; veratrumine; verdins; verteporfin; vinorelbine; vinxaltine; vitaxin; vorozole; zanoteron; zeniplatin; benzyl retinol; and zestatin. Any of the foregoing chemotherapeutic agents may be administered at a clinically effective dose. The chemotherapeutic agent may also be administered about -14, -13, -12, -11, -10, -9, -8, -7, -6, -5, -4, -3, -2, -1, 0, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13 days or up to about 14 days after administration of an agent provided herein. In some cases, the subject may have a refractory cancer that is unresponsive to chemotherapy. ;

[0179] Pharmaceutical composition

[0180] Provided herein is a pharmaceutical composition, wherein the pharmaceutical composition comprises a medicament selected from Table 1 or a combination of medicaments selected from Table 1 and / or Table 2; and a pharmaceutically acceptable excipient. In certain embodiments, the pharmaceutical composition further comprises a cell death inducing agent. In certain embodiments, the pharmaceutical composition further comprises a chemotherapeutic agent. In certain embodiments, provided herein is a pharmaceutical composition as a suspension, optionally a uniform suspension. In certain embodiments, provided herein is a pharmaceutical composition in the form of an emulsion. In certain embodiments, provided herein is a pharmaceutical composition comprising a salt form of any one of the medicaments provided herein. In certain embodiments, the salt is a mesylate.

[0181] Also provided herein is a pharmaceutical composition comprising an ferroptosis inducer or an iron-dependent cell death agent provided herein. Also provided herein is a pharmaceutical composition comprising a ferroptosis regulator. In some cases, the pharmaceutical composition may be in unit dosage form. In certain embodiments, the medicament provided herein is combined with a pharmaceutically acceptable salt, excipient and / or carrier to form a pharmaceutical composition. Pharmaceutical salts, excipients and carriers may be selected based on the route of administration, the location of the target problem and the time course of drug delivery. Pharmaceutically acceptable carriers or excipients may include solvents, dispersion media, coating agents, antibacterial and antifungal agents, isotonic agents and absorption delaying agents, etc., that are compatible with drug administration.

[0182] In certain embodiments, the pharmaceutical composition is in the form of a solid, semisolid, liquid or gas (aerosol). According to known techniques, an injectable preparation, such as a sterile injectable aqueous or oily suspension, can be prepared using a suitable dispersant or wetting agent and a suspending agent. The sterile injectable preparation can also be a sterile injectable solution, suspension or emulsion in a non-toxic parenteral acceptable diluent or solvent. Acceptable vehicles and solvents that can be used include water, Ringer's solution (U.S. Pharmacopoeia) and isotonic sodium chloride solution. In addition, a sterile, non-volatile oil is used as a solvent or suspension medium. For this purpose, any mild non-volatile oil can be used, including synthetic monoglycerides or diglycerides. In addition, fatty acids such as oleic acid are used in the preparation of injectable preparations. The injectable preparation can be sterilized, for example, by filtering through a bacterial retention filter, or by incorporating a sterilizing agent in the form of a sterile solid composition, which can be dissolved or dispersed in sterile water or other sterile injectable media before use.

[0183] Exemplary carriers and excipients can include dextrose, sodium chloride, sucrose, lactose, cellulose, xylitol, sorbitol, maltitol, gelatin, polymers, polyethylene glycol (PEG), polyvinyl pyrrolidone (PVP), and any combination thereof. In certain embodiments, excipients such as dextrose or sodium chloride can be in a percentage of about 0.5%, 1%, 1.5%, 2%, 2.5%, 3%, 3.5%, 4%, 4.5%, 5%, 5.5%, 6%, 6.5%, 7%, 7.5%, 8%, 8.5%, 9%, 9.5%, 10%, 10.5%, 11%, 11.5%, 12%, 12.5%, 13%, 13.5%, 14%, 14.5%, or up to about 15%.

[0184] Solid dosage forms for oral administration include capsules, tablets, pills, powders and granules. In such solid dosage forms, the encapsulated or unencapsulated conjugate is mixed with at least one inert, pharmaceutically acceptable excipient or carrier such as sodium citrate or dicalcium phosphate and / or (a) fillers or extenders such as starches, lactose, sucrose, glucose, mannitol and silicic acid, (b) binders such as carboxymethylcellulose, alginates, gelatin, polyvinyl pyrrolidone, sucrose and acacia, (c) humectants such as glycerol, (d) disintegrants such as agar, calcium carbonate, potato or tapioca starch, alginic acid, certain silicates and sodium carbonate, (e) solution retarders such as paraffin, (f) absorption accelerators such as quaternary ammonium compounds, (g) wetting agents such as cetyl alcohol and glyceryl monostearate, (h) absorbents such as kaolin and bentonite clay, and (i) lubricants such as talc, calcium stearate, magnesium stearate, solid polyethylene glycols, sodium lauryl sulfate, and mixtures thereof. In the case of capsules, tablets and pills, the dosage form may also comprise buffering agents.

[0185] Tablets may have a film coating or an enteric coating according to methods known in the art. Liquid products for oral administration may take the form of, for example, solutions, syrups or suspensions, or they may be present as dry products for reconstitution with water or other suitable vehicles before use. Such liquid preparations may be prepared by conventional means with pharmaceutically acceptable carriers and additives, such as suspending agents, for example, sorbitol syrup, cellulose derivatives or hydrogenated edible fats; emulsifiers, such as lecithin or gum arabic; non-aqueous vehicles, such as almond oil, oily esters, ethanol or fractionated vegetable oils; and preservatives, such as methyl or propyl parabens or sorbic acid. The products may also contain buffer salts, flavoring agents, colorants and / or sweeteners as needed. If desired, products for oral administration may be appropriately formulated to produce controlled release of the active compound.

[0186] Formulations suitable for buccal (sublingual) administration include, for example, lozenges containing the active compound in a flavored basis, usually sucrose and acacia or tragacanth; as well as pastilles containing the compound in an inert basis such as gelatin and glycerin or sucrose and acacia.

[0187] The ferroptosis inducers or ferroptosis modulators provided herein can be formulated as rectal compositions, e.g., suppositories or retention enemas, e.g., containing conventional suppository bases, e.g., cocoa butter or other glycerides, or a gel-forming agent such as carbomer.

[0188] The pharmaceutical compositions may also be administered via controlled release formulations and / or delivery devices (see, eg, US Pat. No. 5,733,566).

[0189] Various delivery vehicles are known and can be used to administer the ferroptosis inducers or ferroptosis regulators provided herein, such as but not limited to encapsulation in liposomes, microparticles, microcapsules, nanoparticles, vectors and recombinant cells. Liposomes and / or nanoparticles can also be used with the administration of the compositions herein. Liposomes are formed by phospholipids dispersed in an aqueous medium and spontaneously form multilayer concentric bilayer vesicles (also referred to as multilayer vesicles (MLVs)). MLVs typically have a diameter of 25nm to 4μm. Ultrasonic treatment of MLVs results in the formation of small unilamellar vesicles (SUVs) with a diameter in the range of 200 to 500 angstroms, which contain an aqueous solution in the core.

[0190] Phospholipids can form a variety of structures other than liposomes when dispersed in water, depending on the molar ratio of lipid to water. At low ratios, liposomes form. The physical properties of liposomes depend on the presence of pH, ionic strength and divalent cations. Liposomes can show low permeability to ions and polar substances, but undergo phase transitions at elevated temperatures, which significantly change their permeability. The phase transition involves changing from a tightly packed ordered structure (known as a gel state) to a loosely packed less ordered structure (known as a fluid state). This occurs at a characteristic phase transition temperature and results in an increase in the permeability to ions, sugars and drugs.

[0191] Liposomes interact with cells through different mechanisms: endocytosis by phagocytic cells of the reticuloendothelial system (such as macrophages and neutrophils); adsorption to the cell surface, which is through nonspecific weak hydrophobic or electrostatic forces, or through specific interactions with cell surface components; fusion with the plasma membrane by inserting the lipid bilayer of the liposome into the plasma membrane, while releasing the liposomal contents into the cytoplasm; and by transferring liposomal lipids to the cell membrane or subcellular membrane, or vice versa, without any binding to the liposomal contents. Changing the liposome formulation can change the mechanism of action, although more than one can work at the same time. Nanocapsules can usually capture compounds in a stable and reproducible manner. In order to avoid side effects caused by intracellular polymer overload, such ultrafine particles (about 0.1 μm in size) should be designed using polymers that can be degraded in vivo. Biodegradable polyalkyl-cyanoacrylate nanoparticles can also be used as delivery vehicles.

[0192] The nanoparticle carriers specifically targeting tissues provided herein can also be used as pharmaceutically acceptable carriers. In certain embodiments, the nanoparticles are gold nanoparticles, platinum nanoparticles, iron oxide nanoparticles, lipid nanoparticles, selenium nanoparticles, the glycol chitosan nanoparticles (CNP) of tumor targeting, the nanoparticles sensitive to cathepsin B, hyaluronic acid nanoparticles, paramagnetic nanoparticles or polymer nanoparticles.

[0193] The pharmaceutical preparation of the ferroptosis inducer or ferroptosis regulator suitable for transdermal application includes an effective amount of the agent and a carrier. The carrier includes an absorbable pharmacologically acceptable solvent to assist the agent to pass through the subject's skin. For example, the transdermal device is in the form of a bandage or patch, which includes a backing member, a reservoir containing a compound (optionally with a carrier), an optional rate-controlling barrier (for delivering the compound to the host's skin at a controlled and predetermined rate over an extended period of time) and a tool for fixing the device to the skin. Matrix transdermal preparations can also be used. Preparations suitable for topical application, such as to the skin and eyes, are preferably aqueous solutions, ointments, creams or gels well known in the art. The preparation may contain solubilizers, stabilizers, tonicity enhancers, buffers and preservatives.

[0194] In certain embodiments, the ferroptosis inducers or ferroptosis regulators provided herein are formulated as reservoir compositions. Such long-acting preparations can be administered by implantation (e.g., subcutaneously or intramuscularly) or by intramuscular injection. The ferroptosis inducers or ferroptosis regulators can be formulated with suitable polymers or hydrophobic materials (e.g., as an emulsion in an acceptable oil), ion exchange resins, biodegradable polymers, or as slightly soluble derivatives (e.g., as slightly soluble salts).

[0195] In certain embodiments, one or more medicaments provided herein are formulated as a pharmaceutical food composition (also referred to as medical food). The food composition can be edible for mammals (e.g., humans or non-human mammals). The medicaments provided herein can be formulated as dietary supplements or medical foods. In certain embodiments, the medicaments provided herein are administered together with food ingredients. Food ingredients are any products, compositions or food components known to have or disclosed as having nutritional effects. Food can include various meats (e.g., beef, pork, poultry, fish, etc.), dairy products (e.g., milk, cheese, eggs), fruits, vegetables, cereals, bread, etc. and components thereof. Food can be fresh or preserved, for example, by canning, dehydration, freezing or smoking. Food can be provided in a raw, unprocessed and / or natural state, or in a cooked, processed and / or mixed state. In certain embodiments, the food ingredients are selected from fat, carbohydrates, proteins, fibers, nutritional balance agents and mixtures thereof. In certain embodiments, the pharmaceutical food composition provided herein further comprises one or more of protein or amino acids. In certain embodiments of any of the aspects, the pharmaceutical food composition further comprises adenine, one or more vitamins (eg, vitamin E), potassium, fatty acids, and / or calcium carbonate.

[0196] Method of administering the drug

[0197] A method of administering a treatment regimen to a subject having a disease or disorder (e.g., cancer, autoimmune disease, obesity, or fibrosis) can be provided herein. In certain embodiments, the administration is a continuous administration of a therapeutically effective amount of an ferroptosis inducer or ferroptosis modulator. In certain embodiments, the continuous administration of an ferroptosis inducer comprises providing a tissue with sufficient to achieve at least about 10 ng / mm 2 The distribution of the ferroptosis inducing agent lasts for at least 4 hours, thereby inducing ferroptosis in the tissue. In certain embodiments, the continuous administration further forms a gradient of a subtherapeutic amount of ferroptosis inducing agent near the site of administration in the tissue. In certain embodiments, the continuous administration of ferroptosis inducing agent or ferroptosis regulator comprises another administration step. In certain embodiments, the ferroptosis inducing agent or the ferroptosis regulator is administered more than once. In certain embodiments, the administration is performed by a system provided herein. In certain embodiments, the administration is local administration in the tissue. In certain embodiments, the tissue is contacted with an effective amount of an iron-dependent cell death agent in vivo for a period of at least 4 hours. In certain embodiments, the administration comprises contacting mammalian tissue with the initiator, and contacting mammalian tissue with an effective amount of ferroptosis inducing agent provided herein, wherein the ferroptosis inducing agent induces targeted cell death in mammalian tissue in vivo. In certain embodiments, the administration is local administration or systemic administration. In certain embodiments, the administration or contacting step is by intratumoral injection, oral administration, transdermal injection, inhalation, nasal administration, topical administration, vaginal administration, ophthalmic administration, intracerebral administration, rectal administration.

[0198] In some cases, the medicament or medicament combination provided herein is used as a unit dosage form. Many medicaments can be used as liquids, capsules, tablets or chewable tablets for oral administration. Since the oral route is the most convenient and usually the safest and cheapest, it is the most commonly used route. However, since the drug usually moves through the approach of the digestive tract, it has limitations. For orally administered medicaments, absorption may start in the mouth and stomach. However, most medicaments are usually absorbed from the small intestine. The drug passes through the intestinal wall and arrives at the liver, and is then transported to its target site by the bloodstream. The intestinal wall and the liver can chemically change (metabolize) many medicaments, thereby reducing the amount of the drug arriving in the bloodstream. Therefore, these medicaments are usually administered in smaller doses when injected intravenously to produce the same effect.

[0199] In certain embodiments, the medicament provided herein is formulated for oral administration. In certain embodiments, the medicament provided herein is formulated for administration / for administration by subcutaneous, intradermal, intramuscular, inhalation, intravenous, intraperitoneal, intracranial, intrathecal, intratumoral or oral routes. For the subcutaneous route, the needle is inserted into the adipose tissue just below the skin. After the drug is injected, it then enters the small blood vessels (capillaries) and is taken away by the blood flow. Alternatively, the drug reaches the blood flow through the lymphatic vessels. When a larger volume of drug product is needed, the intramuscular route is preferred over the subcutaneous route. Since the muscle is located below the skin and adipose tissue, a longer needle is used. The medicament is usually injected into the muscle of the upper arm, thigh or buttocks. The speed at which the drug is absorbed into the bloodstream depends in part on the blood supply to the muscle: the scarcer the blood supply, the longer the time required for drug absorption. For the intravenous route, the needle is directly inserted into the vein. The solution containing the drug can be administered in a single dose or by continuous infusion. For infusion, the solution is moved by gravity (from a collapsible plastic bag), or more commonly by an infusion pump through a thin, flexible tube into a tube (catheter) inserted into a vein (usually in the forearm). In some cases, the medicament or treatment regimen is administered as an infusion. The infusion can occur over a period of time. For example, the infusion can be the administration of the medicament or treatment regimen over a period of about 5 minutes to about 5 hours. The infusion can occur over a period of about 5 min, 10 min, 20 min, 30 min, 40 min, 50 min, 1 hour, 1.5 hours, 2 hours, 2.5 hours, 3 hours, 3.5 hours, 4 hours, 4.5 hours, or up to about 5 hours.

[0200] In certain embodiments, intravenous administration is used to deliver accurate doses to the whole body quickly and in a well-controlled manner. It is also used for irritant solutions that can cause pain and damage tissue if administered by subcutaneous or intramuscular injection. Intravenous injection can be more difficult to administer than subcutaneous or intramuscular injection because it may be difficult to insert a needle or catheter into a vein, especially for obese people. When administered intravenously, the drug is immediately delivered to the bloodstream and tends to take effect faster than being administered by any other route. Therefore, health care practitioners closely monitor the people who receive intravenous injections to observe signs that the drug works or causes undesirable side effects. In addition, the effects of drugs administered by this route tend to last for a short time. Therefore, some medicaments must be administered by continuous infusion to keep their effects constant. For the intrathecal route, a needle is inserted between two vertebrae of the lower spine and into the space around the spinal cord. The drug is then injected into the spinal canal. A small amount of local anesthetic is often used to numb the injection site. This route is used when a drug is needed to have a rapid or local effect on the brain, spinal cord, or the layers of tissue that cover them (meninges) (for example, to treat an infection in these structures).

[0201] For administration by inhalation, the ferroptosis inducers or ferroptosis modulators provided herein can be conveniently delivered in the form of an aerosol spray delivery from a pressurized bag or nebulizer, and a suitable propellant, for example, dichlorodifluoromethane, trichlorofluoromethane, dichlorotetrafluoroethane, carbon dioxide or other suitable gas, can be used. For pressurized aerosols, the dosage unit can be determined as follows: a valve is provided to deliver a metered amount. Capsules and tubes (e.g., gelatin capsules and tubes) for use in inhalers or insufflators can be formulated, containing a powder mixture of the compound and a suitable powder matrix (e.g., lactose or starch). Medicaments administered by oral inhalation can be atomized into droplets that are smaller than those administered by the nasal route, so that the medicament can pass through the trachea and enter the lungs. The depth to which the medicament enters the lungs depends on the size of the droplets. Smaller droplets enter deeper, which increases the amount of absorbed drug. In the lungs, they are absorbed into the bloodstream.

[0202] Medications applied to the skin are usually used for their local action and are therefore most commonly used to treat superficial skin disorders such as psoriasis, eczema, skin infections (viral, bacterial and fungal), itching and dry skin. The drug is mixed with an inactive substance. Depending on the consistency of the inactive substance, the formulation can be an ointment, cream, lotion, solution, powder or gel.

[0203] In some cases, the dosage of the treatment regimen may be determined based on the subject's weight. For subjects who are considered obese (BMI>35), actual weight may need to be used. BMI is calculated as follows: BMI = weight (kg) / [height (m)] 2 .

[0204] In some cases, the treatment regimen can be administered together with a carrier or excipient. The ferroptosis inducer or ferroptosis regulator provided herein can be administered sequentially or in parallel with one or more of the second agent, by the same route of administration or by different routes of administration. When administered sequentially, the time between administrations is selected to facilitate the therapeutic effect and / or safety of the combined therapy, in particular. In certain embodiments, the agent provided herein may be administered first, followed by the second agent, or alternatively, the second agent may be administered first, followed by the agent of the present disclosure (e.g., the ferroptosis inducer / regulator of Table 1). As an example and not as a limitation, the time between administrations is about 1 hour, about 2 hours, about 4 hours, about 6 hours, about 12 hours, about 16 hours, or about 20 hours. In certain embodiments, the time between administrations is about 1, about 2, about 3, about 4, about 5, about 6, or about 7 days or more. In certain embodiments, the time between administrations is about 1 week, 2 weeks, 3 weeks, or 4 weeks or more. In certain embodiments, the time between administrations is about 1 month or 2 months or more.

[0205] In certain embodiments, the ferroptosis inducing agents provided herein are contacted with mammalian tissue for at least about 4 hours, at least about 6 hours, at least about 10 hours, at least about 12 hours, at least about 14 hours, at least about 16 hours, at least about 18 hours, at least about 20 hours, at least about 22 hours, at least about 24 hours, at least about 26 hours, at least about 28 hours, at least about 30 hours, at least about 36 hours, at least about 48 hours, up to 72 hours. In certain embodiments, the ferroptosis inducing agents provided herein are contacted with mammalian tissue for about 4 hours. In certain embodiments, the ferroptosis inducing agents provided herein are contacted with mammalian tissue for about 6 hours. In certain embodiments, the ferroptosis inducing agents provided herein are contacted with mammalian tissue for about 10 hours. In certain embodiments, the ferroptosis inducing agents provided herein are contacted with mammalian tissue for about 12 hours. In certain embodiments, the ferroptosis inducing agents provided herein are contacted with mammalian tissue for about 24 hours. In certain embodiments, the ferroptosis inducing agents provided herein are contacted with mammalian tissue for about 48 hours. In certain embodiments, a ferroptosis-inducing agent provided herein is contacted with a mammalian tissue for about 72 hours.

[0206] When concurrently administered, the agent can be administered separately from the second agent by the same or different routes at the same time, or administered in a single pharmaceutical composition by the same route. In certain embodiments, the amount and frequency of administration of the second agent can use the standard dose and standard administration frequency used for a specific compound.

[0207] Drug administration and tissue distribution

[0208] The methods provided herein include administering to a subject an amount of an agent or pharmaceutical composition provided herein that effectively induces ferroptosis in tissues in vivo. For ease of administration and uniform dosage, agents and pharmaceutical compositions for administration to subjects in need thereof can be formulated in dosage unit form. The dosage unit form is a physically discrete unit of the composition provided herein that is suitable for the subject to be treated. However, it should be understood that the total dosage of the composition provided herein will be determined by the attending physician within the scope of reasonable medical judgment. For any composition provided herein, the therapeutically effective dose can be initially estimated in a cell culture assay or in an animal model (such as a mouse, rabbit, dog, pig, or non-human primate). Animal models can also be used to achieve a desired concentration range and route of administration. Such information can then be used to determine useful doses and routes for administration in humans. The therapeutic effects and toxicity of the compositions provided herein can be determined in cell cultures or experimental animals by standard pharmaceutical procedures, for example, ED 50 (the dose that is therapeutically effective in 50% of the population) and LD 50(The dose is lethal to 50% of the population.) The dose ratio of toxic to therapeutic effects is the therapeutic index, and it can be expressed as LD 50 / ED 50 Ratio. Pharmaceutical compositions that exhibit large therapeutic indices may be useful in certain embodiments. Data obtained from cell culture assays and animal studies can be used in formulating a range of dosage for human use.

[0209] A typical human dose of an agent provided herein (e.g., an ferroptosis inducer / modulator) can be about 10 μg / kg body weight / day to 10,000 mg / kg / day. In certain embodiments, the dose of an agent provided herein is about 0.1 mg / kg to about 1000 mg / kg, 1 mg / kg to 1000 mg / kg, 1 mg / kg to 800 mg / kg, about 1 mg / kg to about 700 mg / kg, about 2 mg / kg to about 500 mg / kg, about 3 mg / kg to about 400 mg / kg, 4 mg / kg to about 300 mg / kg, or about 5 mg / kg to about 200 mg / kg. In certain embodiments, a suitable dose of the agent can be about 1 mg / kg, 5 mg / kg, 10 mg / kg, 15 mg / kg, 20 mg / kg, 25 mg / kg, 30 mg / kg, 35 mg / kg, 40 mg / kg, 45 mg / kg, 50 mg / kg, 60 mg / kg, 70 mg / kg, 80 mg / kg, 90 mg / kg, 100 mg / kg, 125 mg / kg, 150 mg / kg, 175 mg / kg, 200 mg / kg In some embodiments, the dosage of the medicament provided herein is about 100mg / kg / day to about 6,400mg / kg / day, 4 times a day. In some embodiments, the dosage of the medicament provided herein is about 50mg / kg / day to about 25mg / kg / day. In some embodiments, the dosage of the medicament provided herein is about 400mg / kg / day to about 800mg / kg / day. In certain embodiments, the dose of the agent may be administered once daily or divided into sub-doses and administered in multiple doses, for example, two, three, or four times daily.

[0210] In certain embodiments, the agent provided herein is administered in an amount of at least about 10 nanograms (ng) or more, about 20ng or more, about 30ng or more, about 40ng or more, about 50ng or more, about 60ng or more, about 70ng or more, about 80ng or more, about 90ng or more, at most 100ng. In certain embodiments, the agent is administered in an amount of at least about 1 microgram (μg) or more, about 5μg or more, about 10μg or more, about 20μg or more, about 30μg or more, about 40μg or more, about 50μg or more, about 60μg or more, about 70μg or more, about 80μg or more, about 90μg or more, at most 100μg.

[0211] In certain embodiments, the amount of the active ingredient is at least about 0.1 micromolar (μM) or more, about 1 μM or more, about 2 μM or more, about 3 μM or more, about 4 μM or more, about 5 μM or more, about 6 μM or more, about 7 μM or more, about 8 μM or more, about 9 μM or more, about 10 μM or more, about 15 μM or more, about 20 μM or more, about 25 μM or more, about 30 μM or more, about 35 μM or more, about 40 μM or more, about 45 μM or more, about 50 μM or more, about 55 μM or more, about 60 μM or more, about 65 μM or more, about 70 μM or more, about 75 μM or more, about 76 μM or more, about 77 μM or more, about 78 μM or more, about 79 μM or more, about 80 μM or more, about 81 μM or more, about 82 μM or more, about 83 μM or more, about 84 μM or more, about 86 μM or more, about 88 μM or more, about 89 μM or more, about 90 μM or more, about 91 μM or more, about 92 μM or more, about 94 μM or more, about 95 μM or more, about 100 μM or more, about 110 μM or more, about 111 μM or more, about 112 μM or more, about 113 μM or more, about 114 μM or more, about 115 μM or more In some embodiments, the agent provided herein is administered at a concentration of at least about 0.1 μM, at most about 500 μM. In some embodiments, the agent provided herein is administered at a concentration of at least about 1 μM, at most about 500 μM. In some embodiments, the agent provided herein is administered at a concentration of at least about 1 μM, at most about 500 μM. In certain embodiments, an agent provided herein is administered at a concentration of at least about 0.1 μM, up to 10 μM. In certain embodiments, an agent provided herein is administered at a concentration of at least about 1 μM, up to 10 μM.

[0212] In certain embodiments, the ferroptosis inducing agent or ferroptosis modulating agent provided herein is administered intravenously. In certain embodiments, at least about 20 mg / kg, about 25 mg / kg, about 30 mg / kg, about 35 mg / kg, about 40 mg / kg, about 45 mg / kg, about 50 mg / kg, about 60 mg / kg, about 70 mg / kg, about 80 mg / kg, about 90 mg / kg, about 100 mg / kg, about 200 mg / kg, about 300 mg / kg, about 400 mg / kg, about 500 mg / kg, about 600 mg / kg The ferroptosis inducers or ferroptosis modulators provided herein are administered intravenously at a concentration of about 700 mg / kg, about 800 mg / kg, about 900 mg / kg, about 1000 mg / kg, about 1100 mg / kg, about 1200 mg / kg, about 1300 mg / kg, about 1400 mg / kg, about 1500 mg / kg, about 2000 mg / kg, about 2200 mg / kg, about 2400 mg / kg, up to about 2500 mg / kg.

[0213] In certain embodiments, the ferroptosis inducing agent or ferroptosis modulating agent provided herein is orally administered. In certain embodiments, at least about 20 mg / kg, about 25 mg / kg, about 30 mg / kg, about 35 mg / kg, about 40 mg / kg, about 45 mg / kg, about 50 mg / kg, about 60 mg / kg, about 70 mg / kg, about 80 mg / kg, about 90 mg / kg, about 100 mg / kg, about 200 mg / kg, about 300 mg / kg, about 400 mg / kg, about 500 mg / kg, about 600 mg / kg, about 70 mg / kg, about 80 mg / kg, about 90 mg / kg, about 100 mg / kg, about 200 mg / kg, about 300 mg / kg, about 400 mg / kg, about 500 mg / kg, about 600 mg / kg g, about 700mg / kg, about 800mg / kg, about 900mg / kg, about 1000mg / kg, about 1100mg / kg, about 1200mg / kg, about 1300mg / kg, about 1400mg / kg, about 1500mg / kg, about 2000mg / kg, about 2200mg / kg, about 2400mg / kg, up to about 2500mg / kg concentration oral administration provided herein of ferroptosis inducer or ferroptosis regulator. In certain embodiments, once a day, about 25mg / kg concentration oral administration provided herein of ferroptosis inducer or ferroptosis regulator. In certain embodiments, twice a day, about 25mg / kg concentration oral administration provided herein of ferroptosis inducer or ferroptosis regulator. In certain embodiments, at a concentration of about 1300mg / kg / day, about ferroptosis inducer or ferroptosis regulator is orally administered. In certain embodiments, a ferroptosis inducer or ferroptosis modulator provided herein is orally administered at a concentration of about 2400 mg / kg / day.

[0214] The methods provided herein may be characterized by or further comprising measuring the distribution of the agent in the target tissue. 2 ) or cubic millimeters (mm 3 The distribution of the agents provided herein can be determined by measuring the amount or concentration of the agent in the tissue of the tumor. For example, for local administration of the agent to a tumor, the tissue can be about 6 to 7 mm in diameter, 36 to 42 mm in diameter, or 10 to 20 mm in diameter. 2 or 216 to 294 mm 3 Data obtained from animal studies can be used to develop drug distribution ranges in mammalian tissues. Methods for determining tissue distribution of a drug or pharmaceutical agent include, for example, mass spectrometry, chromatography, imaging techniques, and immunoassays. The systems provided herein can be used to determine the distribution of the pharmaceutical agents provided herein.

[0215] In certain embodiments, a therapeutic amount of a ferroptosis inducing agent is administered to a tissue, wherein the administration comprises providing an amount of a ferroptosis inducing agent to the tissue sufficient to achieve a desired drug distribution. In certain embodiments, the agent provided herein reaches at least about 1 ng / mm 2 or more, about 5ng / mm 2 or more, about 10ng / mm 2 Or more, about 15ng / mm 2 or more, about 20ng / mm 2 or more, about 25ng / mm 2 or more, about 30ng / mm 2 or more, about 35ng / mm 2 or more, about 40ng / mm 2 or more, about 45ng / mm 2 or more, about 50ng / mm 2 or more, about 55ng / mm 2 or more, about 60ng / mm 2 Or more, about 65ng / mm 2 or more, about 70ng / mm 2 or more, about 75ng / mm 2 or more, about 80ng / mm 2 Or more, about 85ng / mm 2 or more, about 90ng / mm 2 or more, about 95ng / mm 2 or more, about 100ng / mm 2 or more, about 110ng / mm 2 or more, about 120ng / mm 2 or more, about 130ng / mm 2 or more, about 140ng / mm2 or more, about 150ng / mm 2 Or more, about 160ng / mm 2 or more, about 170ng / mm 2 or more, about 180ng / mm 2 Or more, about 190ng / mm 2 or more, about 200ng / mm 2 or more, about 300ng / mm 2 or more, about 400ng / mm 2 or more, up to 500ng / mm 2 In certain embodiments, the distribution of the agents provided herein reaches at least about 1 ng / mm 3 or more, about 5ng / mm 3 or more, about 10ng / mm 3 Or more, about 15ng / mm 3 or more, about 20ng / mm 3 or more, about 25ng / mm 3 or more, about 30ng / mm 3 or more, about 35ng / mm 3 or more, about 40ng / mm 3 or more, about 45ng / mm 3 or more, about 50ng / mm 3 or more, about 55ng / mm 3 or more, about 60ng / mm 3 Or more, about 65ng / mm 3 or more, about 70ng / mm 3 or more, about 75ng / mm 3 or more, about 80ng / mm 3 Or more, about 85ng / mm 3 or more, about 90ng / mm 3 or more, about 95ng / mm 3 or more, about 100ng / mm 3 or more, about 110ng / mm 3 or more, about 120ng / mm 3 or more, about 130ng / mm 3 or more, about 140ng / mm 3 or more, about 150ng / mm 3 Or more, about 160ng / mm 3 or more, about 170ng / mm 3 or more, about 180ng / mm 3Or more, about 190ng / mm 3 or more, about 200ng / mm 3 or more, about 300ng / mm 3 or more, about 400ng / mm 3 or more, up to 500ng / mm 3 distribution.

[0216] In certain embodiments, the ferroptosis inducer or ferroptosis regulator provided herein is administered at least about once a day, twice a day, three times a day, four times a day, or five times a day. In certain embodiments of any aspect, the ferroptosis inducer or ferroptosis regulator is administered at least about once a week, at least about every 2 weeks, or at least about every 3 weeks. The amount of the drug administered depends on the size of the tissue, the type of disease being treated, and the type of administration (e.g., local administration of tissues in vivo using the system provided herein). The effective dose will vary, depending on the type of disease being treated, the route of administration, the use of excipients, and the possibility of co-use with other therapeutic treatments.

[0217] Effectiveness

[0218] The therapeutic effect of medicament and / or pharmaceutical composition provided herein can be determined by evaluating and contrasting the symptoms and quality of life of the patient before and after application.No matter the mode of administration, such method is applicable.In certain embodiments, before application, it is represented that the patient's symptoms and quality of life are evaluated before the treatment starts, and after application, it is represented that at least 1,2,3,4,5,6,7,8,9,10,11,12,13,14,15,16,17,18,19,20 weeks of evaluation of patient's symptoms and quality of life are evaluated after the treatment starts.In a specific embodiment, after about 2-8,2-6,4-6 or 4 weeks of application are carried out after the treatment starts.In a specific embodiment, clinically and by questionnaire assessment, evaluate the symptoms (for example, the symptoms related to cancer, fibrosis, obesity or autoimmune disease) and quality of life of the patient before and after application.

[0219] The agents and methods provided herein can be used to reduce cancer cell proliferation or survival in vivo or in vitro. Methods for evaluating tumor progression or cell proliferation are known in the art. In certain embodiments, the total response is evaluated from the time point response assessment (based on tumor load) as follows:

[0220] ● Complete response (CR): Disappearance of all target lesions. Any pathological lymph node (whether target or non-target) must have a short axis reduction to <10 mm.

[0221] ● Partial response (PR): A decrease of at least 30% in the sum of the diameters of the target lesions, based on the baseline sum of the diameters.

[0222] Progressive Disease (PD): An increase of at least 20% in the sum of diameters of target lesions, taking the smallest sum on study as reference (including the baseline sum if smallest on study). In addition to the 20% relative increase, the sum must also show an absolute increase of at least 5 mm. (Note: the appearance of one or more new lesions is also considered progression).

[0223] ●Stable disease (SD): Not shrinking enough to meet the criteria for PR or increasing enough to meet the criteria for PD, based on the smallest sum of diameters in the study.

[0224] In certain embodiments, the effectiveness of one or more ferroptosis inducing agents provided herein is assessed using an in vitro cell proliferation assay. The compositions and methods provided herein result in a reduction in the proliferation or survival of multiple cells. For example, after treatment with one or more agents provided herein, cell proliferation or survival is reduced by 5% or more (e.g., 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90% or more) compared to cell proliferation or survival before treatment.

[0225] In certain embodiments, an animal model is used to evaluate the in vivo efficacy of one or more ferroptosis inducers or ferroptosis regulators provided herein. The ferroptosis inducers and methods provided herein can result in a reduction in the size or volume of a hyperproliferative tissue (e.g., a tumor). For example, after treatment, the size of the tissue is reduced by 5% or more (e.g., 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90% or more) relative to its size before treatment. The size of a tissue (e.g., a tumor) can be measured by any reproducible measurement method. The size of the tissue can be measured by the diameter of the tumor or by any reproducible measurement method. Ferropoptosis inhibitors (e.g., agents in Table 2) can be used to determine the efficacy of a specific test agent (also referred to herein as an active agent) for inducing ferroptosis in tissues. For example, a combination of ferroptosis inducers paired with ferroptosis inhibitors (e.g., liproxstatin-1) can be used to determine whether the test agent targets proteins or nucleic acids involved in the ferroptosis pathway (see Figure 1). Further provided herein is a method of saving one or more cells from cell death and / or ferroptosis in vivo, the method comprising: administering a ferroptosis inhibitor to a subject. In certain embodiments, the method further comprises administering a ferroptosis inducer. Further provided herein is a method of screening for ferroptosis sensitivity of multiple cells in a tissue, the method comprising: contacting the tissue with a ferroptosis inducer and a ferroptosis inhibitor; and measuring one or more parameters indicative of ferroptosis. In certain embodiments, the ferroptosis inducer is an agent or a test agent in Table 1. In certain embodiments, the ferroptosis inhibitor is any agent listed in Table 2. In certain embodiments, the ferroptosis inhibitor is liproxstatin-1. In certain embodiments, the one or more parameters indicative of ferroptosis are PUFA concentration, PI index, regulation of mesenchymal cell state marker expression, or regulation of iron or selenium concentration. The screening method provided herein can be easily expanded to perform high-throughput analysis, which allows evaluation or prediction of the ferroptosis inducing activity of the test agent. Similarly, the screening method can be performed in an animal model as discussed above in the presence and absence of a ferroptosis inhibitor.

[0226] Treatment of diseases or disorders (e.g., cancer, obesity or fibrotic diseases) can further lead to a reduction in the number of hyperproliferative tissues and / or adipose tissues (e.g., tumors or fat). For example, after treatment, the number of hyperproliferative tissues or tumors is reduced by 5% or more (e.g., 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90% or more) compared to the number before treatment. The number of tumors, cells or tissues provided herein can be measured by any reproducible measurement method. By counting tumors, cells or tissues visible to the naked eye or at a specific magnification (e.g., 2 times, 3 times, 4 times, 5 times, 10 times or 50 times), the number of tumors, cells or tissues can be measured. In certain embodiments, the methods and ferroptosis inducing agents provided herein reduce the number of tumors in subjects.

[0227] In certain embodiments, when administered to a subject in need thereof, provided herein is an ferroptosis inducing agent or a ferroptosis regulator that reduces the number of fat cells in a subject. In some cases, after treatment, the number of fat cells in a subject may be reduced by about 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90% or more relative to the amount before treatment. In some cases, relative to before treatment, the amount of fat cells after treatment may be reduced by about 5%-99%, 25-75% or 40-80%. In some cases, relative to before treatment, the amount of epididymal white adipose tissue in a subject after treatment may be reduced by about 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90% or more. In some cases, relative to the amount before treatment, the amount of epididymal white adipose tissue after treatment may be reduced by about 5%-99%, 25-75% or 40-80%. In some cases, the reduction can be determined by measuring the amount of epididymal white adipose tissue before and after administration of a therapeutic amount of a ferroptosis modulator.

[0228] In some cases, after treatment, the size of the adipocytes may be reduced. In some cases, the size of the adipocytes may be reduced by about 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90% or more after treatment compared to the size before treatment. In some cases, the size of the adipocytes may be reduced by about 5%-99%, 25-75% or 40-80% after treatment compared to the size before treatment. In some cases, the reduction can be determined by measuring the size of the adipocytes before and after administering a therapeutic amount of a ferroptosis modulator.

[0229] In certain embodiments, the methods and ferroptosis inducers provided herein increase the number or activity of leukocytes in the tumor microenvironment. In certain embodiments, the leukocytes specifically target cancer cells that have high PUFA concentrations compared to normal cells.

[0230] Treatment of cancer can result in a reduction in the number of metastatic nodules in other tissues or organs away from the primary tumor site. For example, after treatment, the number of metastatic nodules is reduced by 5% or more (e.g., 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90% or more) relative to the number before treatment. The number of metastatic nodules can be measured by any reproducible measurement method. The number of metastatic nodules can be measured by counting the visible metastatic nodules to the naked eye or at a specific magnification (e.g., 2 times, 10 times or 50 times).

[0231] Compared to the colony of untreated subjects, treating a disease or disorder (e.g., cancer) can result in an increase in the mean survival time of a colony of subjects treated according to the present invention. For example, the mean survival time increases by more than 30 days (more than 60 days, 90 days, 120 days or longer). By any reproducible means, the increase in the mean survival time of a colony can be measured. For example, by calculating the mean survival time after treatment with the compound of the present invention begins for a colony, the increase in the mean survival time of a colony can be measured. For example, by calculating the mean survival length of a colony after completing the first round of treatment with the compound of the present invention, the increase in the mean survival time of a colony can also be measured.

[0232] Treating a disease or disorder (e.g., cancer) can also result in a reduction in the mortality rate of a population of treated subjects compared to an untreated population. For example, the mortality rate is reduced by more than 2% (e.g., more than 5%, 10%, 25% or more). The reduction in the mortality rate of a population of treated subjects can be measured in any reproducible manner, for example, by calculating the average number of disease-related deaths per unit time of a population after starting treatment with a compound of the present invention. For example, a reduction in the mortality rate of a population can also be measured by calculating the average number of disease-related deaths per unit time of a population after completing the first round of treatment with a ferroptosis inducer.

[0233] Treatment of a disease or disorder can also result in a reduction in at least one symptom associated with the disease, disorder or condition. In certain embodiments, the methods provided herein reduce at least one symptom of a disease or disorder by at least 10%, 20%, 30%, 40%, 50%, 70%, 80%, 90% or more compared to the number before treatment. In certain embodiments, after contacting with mammalian tissue or administering an ferroptosis inducing agent, cell death can be detected at a time point when or after contacting the mammalian tissue with the ferroptosis inducing agent. In certain embodiments, the methods provided herein increase cell death by at least 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90% or more compared to the number before treatment.

[0234] Therapeutic applications

[0235] Provided herein are methods for treating a disease or disorder in a subject. Also provided herein are compositions for use of any of the methods described herein. In certain embodiments, the subject has, is suspected of having, or is at risk of developing a hyperproliferative disease or condition. In certain embodiments, provided herein are methods further comprising obtaining a biopsy sample of tissue for histological analysis. In certain embodiments, the tissue comprises a histological abnormality, wherein the histological abnormality is hyperplasia or fibrosis.

[0236] Provided herein are methods for treating diseases such as obesity or overweight. In some cases, the methods provided herein induce weight loss in a subject by administering a therapeutic amount of a ferroptosis regulator. In some cases, the method may comprise reducing the size of the subject's adipocytes. In some cases, the reduction is determined by measuring the adipocyte size before and after administering a therapeutic amount of a ferroptosis regulator. In some cases, the method comprises reducing the subject's epididymal white adipose tissue. In some cases, the reduction is determined by measuring the amount of epididymal white adipose tissue before and after administering a therapeutic amount of a ferroptosis regulator.

[0237] In certain embodiments, the subject has, is suspected of having a metabolic disease, or is at risk of developing a metabolic disease. In certain embodiments, the metabolic disease is weight gain or obesity. In certain embodiments, the subject has, is suspected of having a weight gain, or is at risk of developing a weight gain. In certain embodiments, the subject is obese. In some cases, the subject has not been diagnosed with cancer. In certain embodiments, the subject is overweight. In certain embodiments, the subject has, is suspected of having a disease or condition associated with obesity, or is at risk of developing a disease or condition associated with obesity. In some cases, the subject has a BMI of 25.0 to <30, which is overweight. In some cases, the subject has a BMI of 30.0 or higher, which is obese. In some cases, the subject can be Class 1 obesity (BMI of 30 to <35), Class 2 obesity (BMI of 35 to <40) or Class 3 obesity (BMI of 40 or higher).

[0238] In certain embodiments, the subject may have, be suspected of having, be diagnosed with, or be at risk of developing Type I diabetes, Type II diabetes, Type IIIa diabetes, or metabolic syndrome.

[0239] In certain cases, the pharmaceutical or therapeutic agent or molecule may be a glucagon-like peptide 1 (GLP-1) agonist.

[0240] In certain embodiments, the subject has, is suspected of having, or is at risk of developing a disease or condition associated with abnormal angiogenesis or vascularization. Diseases or conditions associated with abnormal angiogenesis or vascularization may include, but are not limited to: ocular neovascularization, macular degeneration, retinopathy, sarcoma, polycystic kidney disease, benign hyperplasia, leiomyoma, adenoma, lipoma, hemangioma, fibroma, vascular occlusion, restenosis, atherosclerosis, pre-neoplastic lesions, carcinoma in situ, and cancer. In certain embodiments, the subject has, is suspected of having, or is at risk of developing an autoimmune disease. Non-limiting examples of related autoimmune diseases include: rheumatoid arthritis, inflammatory bowel disease, osteoarthritis, oral hairy leukoplakia, and psoriasis. In certain embodiments, the subject has, is suspected of having, or is at risk of developing fibrosis. Non-limiting examples of diseases and conditions associated with fibrosis include: keloids, hypertrophic scars, systemic sclerosis, pulmonary hypertension, cardiac fibrosis, hypertrophic cardiomyopathy, valvular disease, myelofibrosis, myelodysplastic syndrome, chronic myeloid leukemia, portal hypertension, hepatocellular carcinoma, retroperitoneal fibrosis, intestinal fibrosis, enteropathy, subretinal fibrosis, preretinal fibrosis, cystic fibrosis, emphysema, pancreatic fibrosis, chronic pancreatitis, ductal obstruction, arthrofibrosis, renal fibrosis, nephrogenic systemic fibrosis, renal anemia, chronic kidney disease, Dupuytren's disease, Ledderhose disease (plantar fibromatosis), primary biliary cholangitis (PBC), nonalcoholic steatohepatitis (NASH), scleroderma, diabetic neuropathy, hypertensive nephrosclerosis, allograft nephropathy, cirrhosis of the liver, and pulmonary fibrosis. In certain embodiments, the subject has, is suspected of having, or is at risk of developing a disease or condition associated with polycystic kidney dysplasia, Fabry disease, cystinosis, glomerulonephritis, IgA nephropathy, lupus nephritis, atypical hemolytic uremic syndrome (aHUS), nephrotic syndrome, renal injury, polycystic kidney disease (PKD), renal artery stenosis, renal tubular acidosis, simple renal cyst, or solitary or single functional kidney.

[0241] In certain embodiments, the subject has, is suspected of having cancer, or is at risk of developing cancer. In certain embodiments, the subject has a benign tumor. In certain embodiments, the subject has a precancerous lesion. In certain embodiments, the subject has a basal cell carcinoma (BCC) or a squamous cell carcinoma (SCC). In certain embodiments, the subject has a metastatic tumor. In certain embodiments, the cancer is a solid cancer or a blood cancer. In certain embodiments, the blood cancer is a leukemia or a lymphoma. In certain embodiments, the subject has a solid tumor. In certain embodiments, the solid tumor is a carcinoma, a melanoma, or a sarcoma. In certain embodiments, the melanoma is a dedifferentiated melanoma or amelanotic melanoma. In certain embodiments, the subject suffers from a melanoma with a B-Raf proto-oncogene, a serine / threonine kinase (BRAF) mutation. In certain embodiments, the subject suffers from a sarcoma with a Kirsten rat sarcoma (KRAS) mutation. In certain embodiments, the sarcoma is a soft tissue sarcoma. In certain embodiments, the sarcoma is a leiomyosarcoma. In certain embodiments, the cancer is colon adenocarcinoma.

[0242] Non-limiting examples of cancers that can be treated with the agents provided herein include: acoustic neuroma; adenocarcinoma; adrenal cancer; anal cancer; angiosarcoma (e.g., lymphangiosarcoma, lymphangioendothelioma, angiosarcoma); appendix cancer; benign monoclonal gammopathy; bile duct cancer (e.g., cholangiocarcinoma); bladder cancer; breast cancer (e.g., adenocarcinoma of the breast, papillary carcinoma of the breast, breast cancer, medullary carcinoma of the breast); brain cancer (e.g., meningioma, glioblastoma, glioma (e.g., astrocytoma, oligodendroglioma), medulloblastoma); bronchial carcinoma; carcinoid tumor; cervical cancer (e.g., cervical adenocarcinoma); choriocarcinoma; chordoma; craniopharyngioma; Connective tissue cancer; epithelial cancer; ependymoma; endothelial sarcoma (e.g., Kaposi's sarcoma, multiple idiopathic hemorrhagic sarcomas); endometrial cancer (e.g., uterine cancer, uterine sarcoma); esophageal cancer (e.g., adenocarcinoma of the esophagus, Barrett's adenocarcinoma); Ewing's sarcoma; eye cancer (e.g., intraocular melanoma, retinoblastoma); familial eosinophilia; gallbladder cancer; gastric cancer (e.g., gastric adenocarcinoma); gastrointestinal mesenchymal tumor (GIST); germ cell carcinoma; head and neck cancer (e.g., head and neck squamous cell carcinoma, mouth cancer (e.g., oral squamous cell carcinoma), laryngeal cancer (e.g., laryngeal cancer, pharyngeal cancer, nasopharyngeal carcinoma, oropharyngeal cancer)); hematopoietic system cancer ( For example, leukemias such as acute lymphocytic leukemia (ALL) (e.g., B cell ALL, T cell ALL), acute myeloid leukemia (AML) (e.g., B cell AML, T cell AML), chronic myeloid leukemia (CML) (e.g., B cell CIVIL, T cell CML) and chronic lymphocytic leukemia (CLL) (e.g., B cell CLL, T cell CLL)); lymphomas such as Hodgkin lymphoma (HL) (e.g., B cell HL, T cell HL) and non-Hodgkin lymphoma (NHL) (e.g., B cell NHL such as diffuse large cell lymphoma (DLCL) (e.g. , diffuse large B-cell lymphoma), follicular lymphoma, chronic lymphocytic leukemia / small lymphocytic lymphoma (CLL / SLL), mantle cell lymphoma (MCL), marginal zone B-cell lymphoma (e.g., mucosa-associated lymphoid tissue (MALT) lymphoma, nodal marginal zone B-cell lymphoma, splenic marginal zone B-cell lymphoma), primary mediastinal B-cell lymphoma, Burkitt lymphoma, lymphoplasmacytic lymphoma (e.g., Waldenstrom's macroglobulinemia), hairy cell leukemia (HCL), immunoblastic large cell lymphoma, precursor B lymphoblastic lymphoma, and primary central nervous system (CNS) lymphoma;and T-cell NHL such as precursor T-lymphoblastic lymphoma / leukemia, peripheral T-cell lymphoma (PTCL) (e.g., cutaneous T-cell lymphoma (CTCL) (e.g., mycosis fungoides, Sezary syndrome), angioimmunoblastic T-cell lymphoma, extranodal natural killer T-cell lymphoma, enteropathy-type T-cell lymphoma, subcutaneous panniculitis-like T-cell lymphoma, and anaplastic large cell lymphoma); a mixture of one or more leukemias / lymphomas as described above; and multiple myeloma (MM)), heavy chain disease (e.g., alpha chain disease, gamma chain disease, mu chain disease); hemangioblastoma; hypopharyngeal cancer; inflammatory myofibroblastic tumor; immune cell amyloidosis; kidney cancer (e.g., Wilms tumor, also known as Wilman tumor, renal cell carcinoma); liver cancer (e.g., hepatocellular carcinoma (HCC), malignant hepatoma); lung cancer (e.g., Bronchogenic carcinoma, small cell lung cancer (SCLC), non-small cell lung cancer (NSCLC), lung adenocarcinoma); leiomyosarcoma (LMS); mastocytosis (e.g., systemic mastocytosis); myoma; myelodysplastic syndrome (MDS); mesothelioma; myeloproliferative disorders (MPDs) (e.g., polycythemia vera (PV), essential thrombocythemia (ET), angiogenic myeloid metaplasia (AMM) also known as myelofibrosis (MF), chronic idiopathic myelofibrosis, chronic myelocytic leukemia (CIVIL), chronic neutrophilic leukemia (CNL), hypereosinophilic syndrome (HES); neuroblastoma; neurofibromas (e.g., neurofibromatosis type 1 or type 2 (NF), schwannomatosis); neuroendocrine cancers (e.g., gastroenteropancreatic neuroendocrine tumors (GEPs) NET), carcinoid tumor); osteosarcoma (e.g., bone cancer); ovarian cancer (e.g., cystadenocarcinoma, ovarian embryonal carcinoma, ovarian adenocarcinoma); papillary adenocarcinoma; pancreatic cancer (e.g., pancreatic adenocarcinoma, intraductal papillary mucinous neoplasm (IPMN), islet cell tumor); penile cancer (e.g., Paget's disease of the penis and scrotum); pinealoma; primary neuroectodermal tumor (PNT); plasmacytoma; paraneoplastic syndrome; intraepithelial neoplasm; prostate cancer (e.g., prostate adenocarcinoma); colorectal cancer; rhabdomyosarcoma; salivary gland cancer; skin cancer (e.g., squamous cell carcinoma (SCC), horn cancer acanthoma (KA), melanoma, basal cell carcinoma (BCC)); small intestinal cancer (e.g., appendix cancer); soft tissue sarcomas (e.g., malignant fibrous histiocytoma (MFH), liposarcoma, malignant peripheral nerve sheath tumor (MPNST), chondrosarcoma, fibrosarcoma, myxosarcoma); sebaceous gland carcinoma; small intestinal cancer; sweat gland carcinoma; synovioma; testicular cancer (e.g., seminoma, testicular embryonal carcinoma); thyroid cancer (e.g., papillary carcinoma of the thyroid, papillary thyroid carcinoma (PTC), medullary thyroid carcinoma); urethral cancer; vaginal cancer; and vulvar cancer (e.g., Paget's disease of the vulva). ;

[0243] Provided herein is a method of administering an ferroptosis inducing agent to a tissue, wherein the tissue comprises different cell types. In certain embodiments, the tissue comprises a heterogeneous cell population, wherein the heterogeneous cell population comprises at least one of a precancerous cell and a non-cancerous cell. In certain embodiments, the tissue comprises a heterogeneous cell population, wherein the heterogeneous cell population comprises an immune cell population.

[0244] Provided herein is a method for inducing immune cell recruitment to a tumor, the method comprising: administering to a subject an ferroptosis inducing agent provided herein by any method provided herein. In certain embodiments, the administration is continuous administration for at least about 10 hours, thereby recruiting immune cells to the tumor site. In certain embodiments, the immune cells are leukocytes. In certain embodiments, after contacting with mammalian tissue or administering an ferroptosis inducing agent, immune cell recruitment can be detected at a time point when or after contacting the mammalian tissue with the ferroptosis inducing agent. In certain embodiments, the administration reduces the size of the tumor and / or increases the number of leukocytes in the tumor.

[0245] Systems and Kits

[0246] Provided herein is a system for delivering an ferroptosis inducer or an iron-dependent cell death inducer provided herein. Provided herein is a system for delivering an ferroptosis regulator provided herein. Further provided herein is a system for inducing ferroptosis in vivo, the system comprising: an implantable microdevice configured for local administration to a tissue, the microdevice comprising: (a) a cylindrical support structure having at least one micropore formed on the surface or inside of the support structure; (b) a microdose of ferroptosis inducer in the at least one micropore; and (c) a compound release mechanism for sustained administration, which is used to control the release of the ferroptosis inducer from the micropore, wherein the microdose of ferroptosis inducer forms a gradient of a subtherapeutic dose of ferroptosis inducer at the administration site within the tissue for a period of at least 4 hours, wherein the microdevice is configured to allow implantation into the tissue using a catheter, cannula or biopsy needle, and wherein the microdevice is further configured to release the ferroptosis inducer from the at least one micropore to the administration site within the apoptosis-resistant tissue adjacent to the at least one micropore.

[0247] Further provided herein is a system for identifying ferroptosis induction in an animal model, the system comprising: (a) an animal model comprising a target tissue of interest; (b) a microdevice constructed to allow implantation into tissue in the animal model using a catheter, cannula or biopsy needle, the microdevice comprising: (i) at least one microwell containing one or more active agents; (ii) a microdose of one or more active agents in the at least one microwell; and (iii) a compound release mechanism comprising a polymer matrix for controlling the release of the one or more active agents from the microwell into the tissue; wherein the system measures the results of ferroptosis induction in the animal model after administering the one or more active agents to the tissue relative to a baseline tissue to which the one or more active agents are not administered, and identifies the one or more active agents that induce ferroptosis in the tissue.

[0248] Further provided herein is a system for in vivo screening of ferroptosis-induced cell death, the system comprising: (a) an animal model comprising a target tissue of interest; (b) a microdevice configured to allow implantation into the tissue in the animal model using a catheter, cannula or biopsy needle, the microdevice comprising: (i) at least one microwell containing one or more active agents; (ii) at least one microwell containing one or more ferroptosis inhibitors; (ii) a microdose of one or more active agents, and / or one or more ferroptosis inhibitors in the at least one microwell; and (iii) a compound release mechanism comprising a polymer matrix for controlling the release of the one or more active agents from the microwells into the tissue; wherein the system measures the results of ferroptosis induction in the animal model after administering the one or more active agents to the tissue relative to a baseline tissue to which the one or more active agents are not administered, wherein the system measures the results of ferroptosis induction in the animal model relative to administration of the one or more active agents and one or more ferroptosis inhibitors to the tissue after administering the one or more active agents to the tissue, and identifies the one or more active agents as inducing ferroptosis in the tissue.

[0249] The system provided herein generally includes a plurality of micropores arranged on or within a support structure. The micropores contain one or more active agents, either alone or in combination, with one or more doses and / or release pharmacokinetics. Preferably, the device is configured to deliver a microdose amount, thereby almost eliminating the overlap of active agents released from different micropores in the tissue. In certain embodiments, the device is configured to facilitate implantation and recovery in the target tissue. In an exemplary embodiment, the device has a cylindrical shape, with symmetrical holes on the outside of the device, each hole containing one or more drugs, with one or more concentrations. The size of the device allows placement using a catheter, cannula or probe. In a preferred embodiment, the device has a guide wire to assist placement and recovery. The device may also include features that help maintain the spatial stability of the tissue removed with the device, such as fins or stabilizers that can be extended from the device before or when removed. Optionally, the device has optical fibers, sensors and / or interactive features, such as remote accessibility (such as Wi-Fi), to provide modification of on-site information retrieval and device release performance. In the most preferred embodiment, the optical fiber and / or sensor can be accessed separately to discrete holes.

[0250] In certain embodiments, the system provided herein is formed by biocompatible silicon, metal, ceramic or polymer. They may include materials such as radiopaque materials or materials that can be used for ultrasound or MRI imaging. They can be manufactured using techniques such as deep ion etching, nanoimprint lithography, micromachining, laser etching, three-dimensional printing or stereolithography. The drug can be loaded as follows: the solution or suspension is injected into the hole, and then the solvent is removed by drying, evaporation or lyophilization, or the drug in tablet or microparticle form is placed in the hole. In a preferred embodiment, the drug is loaded on the top of the hydrogel pad in the micropore. The hydrogel pad will expand during implantation to deliver the drug to the surrounding tissue. Drug release pharmacokinetics vary with drug solubility, excipients, pore size and the tissue into which the device is implanted (the release rate in the tissue with a higher degree of vascularization is greater than the release rate in the tissue with a lower degree of vascularization).

[0251] In certain embodiments, the system provided herein is directly implanted in a solid tumor or tissue to be biopsied. After implantation, the system provided herein can locally release a series of active agents in microdoses. The analysis of the response of the tumor to a series of active agents subsequently can be used to identify specific drugs, drug combinations and / or dosages effective for treating solid tumors in patients. By locally delivering a series of drugs in microdoses, the micro-assay device can be used to test the response of patients to multiple regimens quickly and under actual physiological conditions without inducing systemic toxicity. These data are optionally used in conjunction with genomic data to accurately predict systemic drug responses.

[0252] Without limitation, the systems provided herein can administer the agents provided herein according to any of the methods provided herein. For example, the systems provided herein can be used to deliver microdoses of agents to tissues in vivo. The systems described herein can continuously administer a therapeutic amount of an ferroptosis inducing agent to a tissue, wherein the sustained administration of the therapeutic amount comprises providing the tissue with sufficient to achieve at least about 10 ng / mm 2 The distribution of the ferroptosis inducing agent over a period of at least 4 hours, thereby inducing ferroptosis in the tissue. In certain embodiments, the continuous administration further forms a gradient of a subtherapeutic amount of ferroptosis inducing agent near the administration site within the tissue. In certain embodiments, the continuous administration of a therapeutic amount of ferroptosis inducing agent is at least 10 hours. In certain embodiments, the therapeutic amount of ferroptosis inducing agent is a concentration of at least about 1 μM to 10 μM. In certain embodiments, the system provided herein is implanted in a tumor. In certain embodiments, the system delivers one or more ferroptosis inducing agents to a tumor.

[0253] Provided herein are kits comprising ferroptosis modulators or ferroptosis inducers. In some cases, the kits herein can be in a container. In some cases, the container can comprise glass, plastic, metal, or a combination thereof.

[0254] Exemplary embodiments

[0255] Many compositions and methods are disclosed herein. Specific exemplary embodiments of these compositions and methods are disclosed below. The following embodiments describe non-limiting arrangements of the combinations of features disclosed herein. Other arrangements of feature combinations are also contemplated. Specifically, each of these numbered embodiments is considered to be dependent on or related to each previous or subsequent numbered embodiment, regardless of their order of listing.

[0256] Embodiment 1. A method of inducing weight loss or fat loss in a subject, the method comprising administering to the subject a ferroptosis-inducing agent in an amount sufficient to reduce adipocyte size or adipocyte number in the subject.

[0257] Embodiment 2. The method of embodiment 1, wherein the subject is a human.

[0258] Embodiment 3. The method of embodiment 1, wherein the ferroptosis-inducing agent is administered orally.

[0259] Embodiment 4. The method of embodiment 1, wherein the administration is discontinuous and the administration occurs daily for about 5 days, 10 days, 15 days, 20 days, 25 days, 30 days, two months, three months, four months, five months, six months, or one year.

[0260] Embodiment 5. The method of embodiment 1, wherein the administration results in a reduction in the subject's adipocyte size or adipocyte number, wherein the reduction is determined by measuring the adipocyte size or adipocyte number before and after administration of a therapeutic amount of a ferroptosis modulator, optionally using microscopic evaluation of a biopsy sample.

[0261] Embodiment 6. The method of embodiment 1, wherein the method further comprises reducing epididymal white adipose tissue in the subject.

[0262] Embodiment 7. The method of embodiment 6, wherein the subject is overweight or obese.

[0263] Embodiment 8. The method of embodiment 6, wherein the subject is obese and has a BMI of 30.0 or higher.

[0264] Embodiment 9. The method of embodiment 6, wherein the subject is overweight and has a BMI of 25.0 to <30.

[0265] Embodiment 10. A method for inducing, regulating or modulating weight loss, body composition or fat loss or fat loss in a subject, the method comprising administering to the subject a therapeutically effective amount of a ferroptosis regulator, a glutamate-cysteine ​​ligase (GCL) regulator, an agent that binds to GCL, an agent that inhibits GCL, or any combination thereof, thereby inducing, regulating or modulating weight loss, body composition or fat loss or fat loss in the subject.

[0266] Embodiment 11. The method of embodiment 10, wherein the subject is a human.

[0267] Embodiment 12. The method of embodiment 11, wherein the subject is male.

[0268] Embodiment 13. The method of any preceding embodiment, wherein the agent is delivered orally as a solution having a concentration of at least about 5 mg of agent / mL of solution.

[0269] Embodiment 14. The method of embodiment 1, wherein the administering is discontinuous and the administering occurs daily for about 5 days, 10 days, 15 days, 20 days, 25 days, 30 days, two months, three months, four months, five months, six months, or one year.

[0270] Embodiment 15. The method of embodiment 1, wherein the agent is selected from: (1S,3R)-RSL3, ML-162, ML-210, JKE-1674, JKE-1716, erastin, jacaric acid, trigonelline, glutamate, sulfasalazine, auranofin, brucea bruceiol, sorafenib, sorafenib-d3, sorafenib tosylate, trigonelline, FIN56, FINO2, CIL56, dihydroisotanshinone I, GPX4-IN-3, sulfoximine, BSO and salts of any of these.

[0271] Embodiment 16. The method of embodiment 1, wherein the administration results in a decrease in the size of adipocytes in the subject, wherein the decrease is determined by measuring adipocyte size before and after administration of a therapeutic amount of a ferroptosis modulator, optionally using microscopic evaluation of a biopsy sample.

[0272] Embodiment 17. The method of embodiment 1, wherein the administration results in a reduction in the number of adipocytes in the subject, wherein the reduction is determined by measuring the number of adipocytes before and after administration of a therapeutic amount of a ferroptosis modulator, optionally using microscopic evaluation of a biopsy sample.

[0273] Embodiment 18. The method of embodiment 16 or embodiment 17, wherein the method comprises reducing epididymal white adipose tissue in the subject.

[0274] Embodiment 19. The method of embodiment 18, wherein the subject is overweight or obese.

[0275] Embodiment 20. The method of embodiment 19, wherein the subject is obese and has a BMI of 30.0 or higher.

[0276] Embodiment 21. The method of embodiment 19, wherein the subject is overweight and has a BMI of 25.0 to <30.

[0277] Embodiment 22. The method of any preceding embodiment, wherein an additional agent or therapy is administered to the subject concurrently or consecutively.

[0278] Embodiment 23. The method of embodiment 22, wherein the additional agent or therapy is selected from: a GLP-1 agonist, a gastric inhibitory polypeptide analog, naltrexone-bupropion or a salt thereof, orlistat or a salt thereof, phentermine or a salt thereof, phentermine-topiramate or a salt thereof, selenium, exercise therapy, surgery, dietary modification, and any combination thereof.

[0279] Embodiment 24. The method of embodiment 23, wherein the additional agent or therapy comprises administration of a GLP-1 agonist comprising dulaglutide, exenatide, semaglutide, liraglutide, lixisenatide, or a salt of any of these.

[0280] Embodiment 25. The method of embodiment 23, wherein the additional agent or therapy comprises administration of a gastric inhibitory polypeptide comprising telpotide or a derivative thereof.

[0281] Embodiment 26. The method of embodiment 23, wherein the additional agent or therapy comprises a surgical procedure comprising a gastric sleeve, a gastric bypass, an adjustable gastric band, a balloon placed in the stomach, or any combination thereof.

[0282] Embodiment 27. The method of any preceding embodiment, wherein the subject has not been diagnosed with cancer.

[0283] Embodiment 28. The method of any preceding embodiment, wherein the subject is a subject in need thereof.

[0284] Embodiment 29. The method of any preceding embodiment, wherein the subject has Type I diabetes, Type II diabetes, Type IIIa diabetes, or metabolic syndrome.

[0285] Embodiment 30. The method of any preceding embodiment, wherein the medicament is in the form of a pharmaceutical composition further comprising a pharmaceutically acceptable: carrier, excipient, or diluent.

[0286] Embodiment 31. The method of embodiment 30, wherein the pharmaceutical composition is in unit dosage form.

[0287] Embodiment 32. A composition comprising i) a ferroptosis regulator, a glutamate-cysteine ​​ligase (GCL) regulator, an agent that binds to GCL, an agent that inhibits GCL; and ii) a GLP-1 agonist or a salt thereof, a gastric inhibitory polypeptide analog, naltrexone-bupropion or a salt thereof, orlistat or a salt thereof, phentermine or a salt thereof, and phentermine-topiramate or a salt thereof, or selenium or a salt thereof.

[0288] Embodiment 33. A composition comprising i) a glutamate-cysteine ​​ligase (GCL) modulator, and ii) a GLP-1 agonist or a salt thereof, a gastric inhibitory polypeptide analog, naltrexone-bupropion or a salt thereof, orlistat or a salt thereof, phentermine or a salt thereof, and phentermine-topiramate or a salt thereof, or selenium or a salt thereof.

[0289] Embodiment 34. The composition of embodiment 32 or embodiment 33, which is a pharmaceutical composition.

[0290] Embodiment 35. The pharmaceutical composition of embodiment 34, which is in unit dosage form.

[0291] Embodiment 36. The composition of any one of embodiments 32-35 for use in treating a disease or disorder.

[0292] Embodiment 37. The composition of any one of embodiments 32-35 for use in treating a kidney disease or disorder.

[0293] Embodiment 38. The composition of any one of embodiments 32-35 for use in treating symptoms associated with a kidney disease or disorder.

[0294] Embodiment 39. The composition for use according to embodiment 36, wherein the disease or condition is weight loss, weight maintenance, a disease or condition related to iron metabolism, or a disease or condition related to high, low or abnormal cholesterol.

[0295] Embodiment 40. A method for regulating iron metabolism in a subject, the method comprising administering to the subject a therapeutically effective amount or an effective amount of a ferroptosis inducer, a glutamate-cysteine ​​ligase (GCL) regulator, an agent that binds to GCL, an agent that inhibits GCL, or any combination thereof, thereby regulating iron metabolism in the subject and thereby inducing ferroptosis in the subject.

[0296] Embodiment 41. A method of regulating iron metabolism in a subject, the method comprising administering to the subject a therapeutically effective amount or an effective amount of a ferroptosis-inducing agent, thereby regulating iron metabolism in the subject.

[0297] Embodiment 42. A method for treating, inducing, regulating or adjusting iron metabolism or a disease or condition related to iron metabolism in a subject, the method comprising administering to the subject a therapeutically effective amount or an effective amount of a ferroptosis regulator, a glutamate-cysteine ​​ligase (GCL) regulator, an agent that binds to GCL, an agent that inhibits GCL, or any combination thereof, thereby treating, inducing, regulating or adjusting iron metabolism or a disease or condition related to iron metabolism in the subject.

[0298] Embodiment 43. A method for regulating cholesterol metabolism in a subject, the method comprising administering to the subject a therapeutically effective amount or an effective amount of a ferroptosis regulator, a glutamate-cysteine ​​ligase (GCL) regulator, an agent that binds to GCL, an agent that inhibits GCL, or any combination thereof, thereby regulating cholesterol metabolism in the subject.

[0299] Embodiment 44. A method of regulating cholesterol metabolism in a subject, the method comprising administering to the subject a therapeutically effective amount or an effective amount of a ferroptosis-inducing agent, thereby regulating cholesterol metabolism in the subject.

[0300] Embodiment 45. A method for treating, inducing, regulating, adjusting or reducing cholesterol metabolism or a disease related to cholesterol metabolism in a subject, the method comprising administering to the subject a therapeutically effective amount or an effective amount of a ferroptosis regulator, a glutamate-cysteine ​​ligase (GCL) regulator, an agent that binds to GCL, an agent that inhibits GCL, or any combination thereof, thereby treating, inducing, regulating, adjusting or reducing cholesterol metabolism or a disease or condition related to cholesterol metabolism in the subject.

[0301] Embodiment 46. A method for treating, inducing, regulating, modulating, reducing or increasing reactive oxygen species or a disease or condition associated with reactive oxygen species in a subject, the method comprising administering to the subject a therapeutically effective amount or an effective amount of a ferroptosis modulator, a glutamate-cysteine ​​ligase (GCL) modulator, an agent that binds to GCL, an agent that inhibits GCL, or any combination thereof, thereby treating, inducing, regulating, modulating, reducing or increasing reactive oxygen species or a disease or condition associated with reactive oxygen species in the subject.

[0302] Embodiment 47. A method for increasing reactive oxygen species in a subject, the method comprising administering to the subject a therapeutically effective amount or an effective amount of a ferroptosis regulator, a glutamate-cysteine ​​ligase (GCL) regulator, an agent that binds to GCL, an agent that inhibits GCL, or any combination thereof, thereby increasing reactive oxygen species in the subject.

[0303] Embodiment 48. A method for promoting at least one of the following: treating, overcoming, regulating or reducing acute stress; a chronic stress response; an immune hypersensitivity reaction; cachexia; kidney disease; neurodegeneration; cardiotoxicity; or cardiotoxicity caused at least in part by chemotherapy in a subject; the method comprising administering to the subject a therapeutically effective amount or an effective amount of a ferroptosis regulator, a glutamate-cysteine ​​ligase (GCL) regulator, an agent that binds to GCL, an agent that inhibits GCL, or any combination thereof, thereby treating, inducing, regulating, modulating, reducing or increasing acute stress; a chronic stress response; an immune hypersensitivity reaction; cachexia; kidney disease; neurodegeneration; cardiotoxicity; or cardiotoxicity caused at least in part by chemotherapy in the subject.

[0304] Embodiment 49. A method for promoting at least one of the following: treating kidney disease in a subject, the method comprising administering to the subject a therapeutically effective amount or an effective amount of a ferroptosis regulator, a glutamate-cysteine ​​ligase (GCL) regulator, an agent that binds to GCL, an agent that inhibits GCL, or any combination thereof, thereby treating the kidney disease in the subject.

[0305] Embodiment 50. A method of promoting at least one of the following: treating a kidney disease in a subject, the method comprising administering to the subject a therapeutically effective amount or an effective amount of a ferroptosis-inducing agent, thereby treating the kidney disease in the subject.

[0306] Embodiment 51. The method of any one of embodiments 40-50, wherein the method comprises reducing epididymal white adipose tissue in the subject.

[0307] Embodiment 52. The method of embodiment 51, wherein the subject is overweight or obese.

[0308] Embodiment 53. The method of embodiment 52, wherein the subject is obese and has a BMI of 30.0 or higher.

[0309] Embodiment 54. The method of embodiment 52, wherein the subject is overweight and has a BMI of 25.0 to <30.

[0310] Embodiment 55. The method of any one of Embodiments 40-54, wherein an additional agent or therapy is administered to the subject concurrently or consecutively.

[0311] Embodiment 56. The method of embodiment 55, wherein the additional agent or therapy is selected from: a GLP-1 agonist, a gastric inhibitory polypeptide analog, naltrexone-bupropion or a salt thereof, orlistat or a salt thereof, phentermine or a salt thereof, phentermine-topiramate or a salt thereof, selenium, exercise therapy, surgery, dietary modification, and any combination thereof.

[0312] Embodiment 57. The method of embodiment 56, wherein the additional agent or therapy comprises administration of a GLP-1 agonist comprising dulaglutide, exenatide, semaglutide, liraglutide, lixisenatide, or a salt of any of these.

[0313] Embodiment 58. The method of embodiment 56, wherein the additional agent or therapy comprises administration of a gastric inhibitory polypeptide comprising telpotide or a derivative thereof.

[0314] Embodiment 59. The method of embodiment 56, wherein the additional agent or therapy comprises a surgical procedure comprising a gastric sleeve, a gastric bypass, an adjustable gastric band, a balloon placed in the stomach, or any combination thereof.

[0315] Embodiment 60. The method of any one of Embodiments 40-59, wherein the subject has not been diagnosed with cancer.

[0316] Embodiment 61. The method of any one of Embodiments 40-59, wherein the subject is a subject in need thereof.

[0317] Embodiment 62. The method of any one of embodiments 40-59, wherein the subject has type I diabetes, type II diabetes, type IIIa diabetes, or metabolic syndrome.

[0318] Embodiment 63. The method of any one of embodiments 40-59, wherein the medicament is in the form of a pharmaceutical composition further comprising a pharmaceutically acceptable: carrier, excipient, or diluent.

[0319] Embodiment 64. The method of embodiment 63, wherein the pharmaceutical composition is in unit dosage form.

[0320] Embodiment 65. The method of any preceding embodiment, wherein the agent is a compound in Table 1 or a salt thereof.

[0321] Embodiment 66. The method of any preceding embodiment, wherein the medicament is in the form of a pharmaceutical composition further comprising a pharmaceutically acceptable: carrier, excipient, or diluent.

[0322] Embodiment 67. The method of embodiment 46, wherein the pharmaceutical composition is in unit dosage form.

[0323] Embodiment 68. A composition comprising i) a ferroptosis regulator, a glutamate-cysteine ​​ligase (GCL) regulator, an agent that binds to GCL, an agent that inhibits GCL; and ii) a GLP-1 agonist or a salt thereof, a gastric inhibitory polypeptide analog, naltrexone-bupropion or a salt thereof, orlistat or a salt thereof, phentermine or a salt thereof, and phentermine-topiramate or a salt thereof, or selenium or a salt thereof.

[0324] Embodiment 69. The composition of embodiment 68, which is a pharmaceutical composition.

[0325] Embodiment 70. The pharmaceutical composition of embodiment 69, which is in unit dosage form.

[0326] Embodiment 71. The composition of any one of embodiments 68-70 for use in treating a disease or disorder.

[0327] Embodiment 72. The composition of any one of embodiments 68-70 for use in treating a kidney disease or disorder.

[0328] Embodiment 73. The composition of any one of embodiments 68-70 for use in treating symptoms associated with a kidney disease or disorder.

[0329] Embodiment 74. The composition for use according to embodiment 71, wherein the disease or condition is weight loss, weight maintenance, a disease or condition related to iron metabolism, or a disease or condition related to high, low or abnormal cholesterol.

[0330] Embodiment 75. A composition for inducing weight loss, fat loss, or both in a subject, the composition for use comprising an inducing agent in an amount sufficient to induce weight loss, fat loss, or both.

[0331] Embodiment 76. The composition for use according to embodiment 75, for inducing weight loss.

[0332] Embodiment 77. The composition for use according to embodiment 75, which is used for inducing fat loss.

[0333] Embodiment 78. The composition for use according to embodiment 75, for inducing weight loss and fat loss.

[0334] Embodiment 79. The composition for use of Embodiment 75, further comprising administering a trigger to the subject before administering the ferroptosis inducer to the subject.

[0335] Embodiment 80. The composition for use of embodiment 75, wherein after administering the ferroptosis-inducing agent to the subject, the adipose tissue in the subject is reduced in weight, volume, or both relative to the weight, volume, or both of the adipose tissue in the subject before administration.

[0336] Embodiment 81. The composition for use of embodiment 80, wherein after administration of the ferroptosis-inducing agent to the subject, the adipose tissue in the subject is reduced in volume relative to the volume of adipose tissue before administration, as determined by a spring clip test, by a dual x-ray absorptiometry (DEXA) scan, waist circumference reduction measurement, a computer axial tomography (CAT) test, or any combination thereof.

[0337] Embodiment 82. The composition for use of embodiment 80, wherein after administration of the ferroptosis-inducing agent to the subject, the adipose tissue in the subject is reduced in weight relative to the weight of the adipose tissue before administration.

[0338] Embodiment 83. The composition for use of embodiment 82, wherein the weight loss is determined as follows: the total weight of the subject is reduced after administration of the ferroptosis inducing agent compared to the total weight of the subject before administration of the ferroptosis inducing agent.

[0339] Embodiment 84. The composition for use of Embodiment 82, wherein the weight loss is determined by comparing DEXA scans of the subject obtained before and after administration of the ferroptosis-inducing agent to the subject.

[0340] Embodiment 85. The composition for use of Embodiment 75 further comprises a decrease in the cell size, cell volume, or both of multiple cells in the adipose tissue of the subject after administering the ferroptosis inducer to the subject, compared to the cell size, cell volume, or both of multiple cells in the adipose tissue of the subject before administration.

[0341] Embodiment 86. The composition for use according to embodiment 75, wherein the subject is a human.

[0342] Embodiment 87. The composition for use according to Embodiment 75, wherein the ferroptosis inducing agent is administered orally.

[0343] Embodiment 88. The composition for use of embodiment 75, wherein said administration occurs for about: 1 day, 2 days, 3 days, 4 days, 5 days, 1 week, 10 days, 2 weeks, 15 days, 20 days, 3 weeks, 25 days, 30 days, one month, two months, three months, four months, five months, six months, one year, two years, three years, four years, five years, lifetime, or as needed.

[0344] Embodiment 89. The composition for use of Embodiment 87, wherein the ferroptosis inducing agent is administered in an amount ranging from about 0.1 ng to about 25,000 mg.

[0345] Embodiment 90. The composition for use of embodiment 87, wherein the ferroptosis inducing agent is administered in an amount of about 1 ng, 10 ng, 100 ng, 1 microgram, 10 micrograms, 100 micrograms, 1 mg, 10 mg, 100 mg, 1 g, 2 g, 3 g, 4 g, 5 g, 6 g, 7 g, 8 g, 9 g, 10 g, 15 g, 20 g or 25 g.

[0346] Embodiment 91. The composition for use of embodiment 87, wherein the ferroptosis inducing agent is administered in an amount ranging from about 1 ng / kg to 1,000 mg / kg, wherein mg is mg of the ferroptosis inducing agent and kg is kg of the subject's body weight.

[0347] Embodiment 92. The composition for use according to Embodiment 75, wherein the ferroptosis inducing agent is administered orally.

[0348] Embodiment 93. The composition for use according to embodiment 75, wherein the ferroptosis inducing agent is administered by one of the following routes: oral administration, subcutaneous, intradermal, intramuscular, inhalation, intravenous, intraperitoneal, intracranial, intrathecal, transdermal, rectal, or direct administration to adipose tissue.

[0349] Embodiment 94. The composition for use of embodiment 75, which treats at least one of: obesity, metabolic syndrome, elevated blood sugar, diabetes, type 2 diabetes, type 3 diabetes, insulin resistance, hypertension, cardiovascular disease, coronary artery disease, cerebrovascular disease, stroke, rheumatic heart disease, arteriosclerosis, atherosclerosis, liver disease, fatty liver disease, non-alcoholic fatty liver disease (NAFLD), non-alcoholic steatohepatitis (NASH), or any combination thereof.

[0350] Embodiment 95. The composition for use according to embodiment 75, wherein the subject is a human being.

[0351] Embodiment 96. The composition for use according to embodiment 75, wherein the subject is a woman.

[0352] Embodiment 97. The composition for use of embodiment 75, wherein the subject is in the range of about 1 year old to about 18 years old.

[0353] Embodiment 98. The composition for use of embodiment 75, wherein the subject is in the range of about 18 to about 120 years old.

[0354] Embodiment 99. The composition for use according to embodiment 75, wherein the subject is a subject in need thereof.

[0355] Embodiment 100. The composition for use according to Embodiment 75, wherein the ferroptosis inducer is contained in a pharmaceutical composition comprising a pharmaceutically acceptable excipient, carrier or diluent.

[0356] Embodiment 101. The composition for use according to embodiment 100, wherein the pharmaceutical composition is in unit dosage form.

[0357] Embodiment 102. The composition for use of Embodiment 75, further comprising administering another therapeutic agent to the subject.

[0358] Embodiment 103. The composition for use according to embodiment 102, wherein the other therapeutic agent is administered concurrently with the ferroptosis-inducing agent.

[0359] Embodiment 104. The composition for use according to embodiment 102, wherein the other therapeutic agent is administered consecutively with the ferroptosis inducing agent.

[0360] Embodiment 105. The composition for use according to embodiment 75, wherein the ferroptosis inducing agent is administered directly to adipose tissue.

[0361] Embodiment 106. The composition for use according to embodiment 105, wherein the administration is continuous administration.

[0362] Embodiment 107. The composition for use of embodiment 105, wherein the administration lasts for about: 1 / 2 hour, 1 hour, 2 hours, 3 hours, 4 hours, 5 hours, 6 hours, 7 hours, 8 hours, 9 hours, 10 hours, 11 hours, 12 hours, 13 hours, 14 hours, 15 hours, 16 hours, 17 hours, 18 hours, 19 hours, 20 hours, 21 hours, 22 hours, 23 hours, 24 hours, 2 days, 3 days, 4 days, 5 days, 6 days or 1 week.

[0363] Embodiment 108. The composition for use of embodiment 105, wherein the adipose tissue is heated to a temperature of about 23, 24, 25, 26, 27, 28, 29, 30, 31, 32 or 33 degrees Celsius.

[0364] Embodiment 109. The composition for use according to embodiment 105, wherein the ferroptosis inducing agent is administered in the form of an aqueous solution.

[0365] Embodiment 110. The composition for use according to embodiment 105, wherein the concentration of the ferroptosis inducing agent in the aqueous solution is about 0.1 nM to about 500 μM.

[0366] Embodiment 111. The composition for use of embodiment 75, wherein the administration results in a reduction in the size of fat cells or the number of fat cells in the subject, wherein the reduction is determined by measuring the size of fat cells or the number of fat cells before and after administration of a therapeutic amount of a ferroptosis regulator, optionally using microscopic evaluation of a biopsy sample.

[0367] Embodiment 112. The composition for use of Embodiment 75, wherein the method further comprises reducing epididymal white adipose tissue in the subject.

[0368] Embodiment 113. The composition for use according to embodiment 75, wherein the subject is overweight or obese.

[0369] Embodiment 114. The composition for use according to embodiment 75, wherein the subject is obese and has a BMI of about 30.0 or higher.

[0370] Embodiment 115. The composition for use according to embodiment 75, wherein the subject is overweight and has a BMI of about 25.0 to about 30.

[0371] Embodiment 116. The composition for use according to embodiment 102, wherein the other therapeutic agent has glucagon-like peptide-1 (GLP-1) receptor agonist activity.

[0372] Embodiment 117. The composition for use according to embodiment 116, wherein the other therapeutic agent comprises semaglutide, dulaglutide, liraglutide, exenatide, telpotide or any combination thereof. Example

[0373] The following examples are set forth to more clearly illustrate to those skilled in the art the principles and practices of the embodiments disclosed herein, but should not be construed as limiting the scope of any claimed embodiments.Unless otherwise stated, all parts and percentages are based on weight.

[0374] Example 1: Cell lines and culture conditions

[0375] Human cancer cell lines were cultured in Ham's F12 medium supplemented with 10% (v / v) fetal bovine serum (FBS), penicillin (100 U / mL) and streptomycin (100 μg / mL). Human cancer cells were cultured in RPMI medium supplemented with 10% FBS, penicillin (100 U / mL) and streptomycin (100 μg / mL). Cells were grown in a humidified incubator at 37° C., 5% carbon dioxide, and split every 3-4 days using a trypsin / EDTA solution.

[0376] Method for adding exogenous PUFA or MUFA: Exogenous fatty acids were dissolved in DMSO and added to the cell culture medium 24 hours after seeding the cells.

[0377] Example 2: Determination of PUFA status

[0378] Lipidomics was performed using gas chromatography-mass spectrometry (GC-MS) or direct infusion mass spectrometry.

[0379] For GC-MS evaluation of cellular PUFA status, membrane lipids were transesterified with 500 μL methanolic HCl, 250 μL n-hexane, and 500 μL internal standard (0.8 mg di-C17-phosphatidylcholine in 1 mL methanol, 0.2% butylated hydroxytoluene as antioxidant). After cooling, 500 μL n-hexane and 1 mL Aqua Dest were added. The upper hexane phase was evaporated with nitrogen. Fatty acid methyl esters (FAME) were dissolved in 60 μL n-hexane. An aliquot of 1 μL was injected into a column of a Varian CP 3800 gas chromatograph (Varian, Darmstadt, Germany) equipped with an Omegawax TM 320 column (0.32 mm inner diameter, 30 m length) (Supelco, Bellefonte, USA). The column temperature was 200 ° C.

[0380] For direct infusion MS analysis, lipids were extracted using a two-step chloroform / methanol procedure. Samples were spiked with an in-house lipid standard mixture containing the following components: cardiolipin 16:1 / 15:0 / 15:0 / 15:0 (CL), ceramide 18:1; 2 / 17:0 (Cer), diacylglycerol 17:0 / 17:0 (DAG), hexosylceramide 18:1; 2 / 12:0 (HexCer), lysophosphatidic acid 17:0 (LPA), lysophosphatidylcholine 12:0 (LPC), lysophosphatidylethanolamine 17:1 (LPE), lysophosphatidylglycerol 17:1 (LPG), lysophosphatidylinositol 17:1 (LPI), lysophosphatidylserine 17:1 (LPS), phosphatidic acid 17:0 / 17:0 (PA), phosphatidylcholine 17:0 / 17:0 (PC), phosphatidylethanolamine 17:0 / 17:0 (PE), phosphatidylglycerol 17:0 / 17:0 (PG), phosphatidylinositol 16:0 / 16:0 (PI), phosphatidylserine 17:0 / 17:0 (PS), cholesterol ester 20:0 (CE), sphingomyelin 18:1;2 / 12:0;0 (SM), triacylglycerol 17:0 / 17:0 / 17:0 (TAG). After extraction, the organic phase was transferred to an infusion plate and dried in a high-speed vacuum concentrator. The dried extract was resuspended in 7.5 mM ammonium acetate in chloroform / methanol / propanol (1:2:4, V:V:V), and the second dried extract was resuspended in a 33% ethanol solution of methylamine in chloroform / methanol (0.003:5:1; V:V:V). Samples were analyzed by direct infusion on a QExactive mass spectrometer (ThermoFisher Scientific) equipped with a TriVersaNanoMate ion source (Advion Biosciences). Samples were analyzed in positive and negative ion modes with a resolution of Rm / z=200=280000 for MS in a single acquisition and Rm / z=200=17500 for tandem mass spectrometry (MS-MS). MS-MS was triggered by an inclusion list covering the corresponding MS mass range scanned in 1 Da increments.

[0381] Example 3: Cell line profiling using ferroptosis inducers with and without rescue agents

[0382] Cell viability assays were performed by seeding 1,000 cells per well (30 μL volume) in opaque white 384-well plates (Corning). Cells were allowed to adhere for 24 hours and then exposed to the compound for 72 hours. The DMSO stock solution of the compound was added to the cells using a CyBioWell Vario liquid dispenser (Analytik Jena AG). CellTiter-Glo (Promega) was used to measure cellular ATP levels as a surrogate for viability. Rescue assays were performed using rescue agents selected from Table 2, which were referred to as anti-ferroptosis rescue agents (N) (N, 1.5 μM), anti-ferroptosis rescue agents (M) (M, 1 μM), anti-ferroptosis rescue agents (P) (P, 50 μM) and other ferroptosis inhibitors in the assay, and the rescue agents were added to the cells when added to the assay plate.

[0383] Target knockdown using genetic reagents ± anti-ferroptosis rescue agents (N)

[0384] For lentiviral shRNA production, 293-T cells were seeded in 6-well plates (280,000 cells / well) in antibiotic-free medium. The next day, cells were transfected with the appropriate shRNA encoding plasmid (450 ng), viral packaging plasmid (p-Delta8.9, 400 ng), and viral envelope plasmid (p-VSV-G, 45 ng) using FuGENE. After 24 hours, the medium was removed and replaced with fresh medium. Three viral supernatants were collected and combined for each shRNA within 36 hours. The combined supernatants were centrifuged, aliquoted, and stored at -80°C until viral infection.

[0385] Lentiviral infection was performed by seeding cells for 12 hours and replacing the medium with medium supplemented with polybrene (8 μg / mL) and aliquots of viral supernatant. Plates were incubated for 48 hours, then the medium was replaced with medium containing 1.5 μg / mL puromycin and incubated at 37°C for 48 hours. Knockdown was assessed by immunoblotting and RT-qPCR.

[0386] Knockout of target using genetic reagents ± anti-ferroptosis rescue agents (N)

[0387] To generate knockout cell lines, lentivirus was generated by overnight polyethylenimine transfection of Leni-X 293T cells with target lentiviral plasmids and packaging plasmids pCMV-dR8.2 dvpr and pCMV-VSV-G in DMEM supplemented with 10% FBS. The next day, the medium was changed to fresh DMEM containing 10% FBS. After 24 and 48 hours, the virus-containing medium was collected and filtered with a 0.45 μm polyethersulfone filter, pooled, and stored at -80°C until viral infection.

[0388] Cells were transduced with pLenti-CRISPR-V2 encoding the appropriate sgRNA of the target gene using 2 μg / mL polybrene, followed by puromycin selection (1 μg / mL) in the presence of ferrostatin-1 (1 μM) for 4 days. Protein knockout was verified by immunoblotting.

[0389] Example 4: Using C11-BODIPY to display lipid peroxidation as an indicator of ferroptosis

[0390] Imaging assay: Human cancer cells were seeded at 5,000 cells / well in 150 μl RPMI medium containing 10% FBS in CellCarrier Ultra 96-well plates (Perkin-Elmer). Cells were incubated at 37°C for 24 hours and then treated with the indicated compounds or DMSO (90 minutes, 37°C). In the last 30 minutes of incubation, 60 nM DRAQ7 (Abcam), 1 μg ml -1 Hoechst 33342 (ThermoFisher) and 1 μM BODIPY 581 / 591C11 (ThermoFisher) dyes were used. Cells were imaged using an Opera Phenix high-volume screening system (Perkin-Elmer) equipped with 405, 488, 560, and 647 nm lasers. Image analysis was performed using Harmony high-volume imaging and analysis software (Perkin-Elmer).

[0391] Lipid peroxidation using C11-BODIPY (flow cytometry assay)

[0392] Human cancer cells were seeded in 96-well plates at 15,000 cells per well in RPMI medium containing 10% FBS. After 48 hours, the medium was replaced with 200 μl of medium containing DMSO or the indicated inhibitor (10 μM) and 1 μM anti-ferroptosis rescue agent (if indicated). The culture was incubated at 37°C for 2 hours. 10 μM ODIPY 581 / 591C11 (Molecular Probes no. C10445) was added to the cells 30 minutes before the end of the incubation period. The cells were aggregated in 200 μl PBS + 0.1% BSA and analyzed by flow cytometry (BD FACSCanto II).

[0393] Example 5: Administration of microdoses of ferroptosis-inducing agents to tumors.

[0394] For allograft studies, cancer cell line-derived tumor cells were injected into the flanks of male C57BL6 / J mice. The assay was initiated when tumors were approximately 6-7 mm in diameter.

[0395] Microdose drug delivery was performed for the assays described herein. The compounds in Table 3 were loaded into the device reservoir using a tapered metal needle. The reservoir was loaded for initial release of the anti-ferroptosis rescue agent (M) (if included), followed by a 4-6 hour delayed release of the ferroptosis inducer. The device was prepared for administration of the dose into mouse tumors. The device delivered the ferroptosis inducer to the tissue for 24 to 72 hours. The tumor was then excised and the tissue was rapidly frozen with liquid nitrogen. The tissue was sectioned using a standard cryostat and 20 μm thick tissue sections were collected from each reservoir for analysis by immunoassays, transcriptomics, and metabolomics assays.

[0396] Table 3. Microdosing of ferroptosis inducers and conditions

[0397]

[0398] Example 6: System for in vivo delivery of ferroptosis-inducing agents

[0399] Ferroptosis inducers and / or triggers are systemically administered to mammals by injection to establish local pharmacokinetics of the drug. Representative drugs tested include: ferroptosis inducers (A), ferroptosis inducers (C), ferroptosis inducers (B), and anti-ferroptosis rescue agents (M). Representative animal models that can be used include, for example, those carrying tumors in a flammable membrane state.

[0400] Each micropore of the drug delivery system with micropores is loaded with about 1.5 micrograms of ferroptosis inducer (crystalline powder). Based on the results of the systemic test, the system is loaded with the same drug. Each drug is loaded alone and at more than one concentration, as well as in combination. After 10, 12, 18, 24, 36 or 48 hours, the device is removed and the histology of the tissue is examined to determine the effect of the ferroptosis inducer on tumor cells adjacent to each hole. The effect of the compound eluted from the micropore is evaluated by different techniques. The tissue excised with the device is measured by standard histopathological techniques, including immunohistochemistry and immunofluorescence. The tissue / device sections are stained by standard immunohistochemistry (IHC) techniques, including hematoxylin and eosin (H&E) staining or any nuclear cell staining such as DAPI, to observe the ingrowth of the tissue, ranging from 20 to about 300 microns. Local biomarkers (e.g., mesenchymal cell state markers or PUFA concentrations) indicating the effect of ferroptosis inducers are measured using mass spectrometry. Analysis of apoptosis, necrosis, mitotic cell death and proliferation is performed. Local microdose responses are determined and used to determine appropriate cancer treatment regimens.

[0401] Several methods have been developed for controlling the release / diffusion of ferroptosis-inducing agents into tissues, including precise placement of micropores along the device housing; the geometry and size of the micropores; and the formulation of the releasing agent. The device micropores through which the ferroptosis-inducing agent is diffused are designed to expose only the tissue area directly adjacent to the micropore opening to the agent being released. This creates different local areas in the tissue in which the effect of the compound is evaluated without interference from other compounds released from different micropores. The creation of discrete drug areas can be used to evaluate the efficacy, and / or dosage and / or release time (sustained, pulsed, delayed, sustained after bolus, etc.) of different agents or combinations thereof.

[0402] The agent is released upward and diffuses into a larger area, or is released downward into a relatively small area of ​​the target tissue. By precisely controlling the transport time as a function of the distance from the micropores, local concentrations of the first agent as a function of the distance from the micropores can be provided at multiple time points after implantation in vivo.

[0403] The concentration gradient region is defined as the concentration of the administered agent decreasing with increasing distance from the microwell. Cleaved caspase 3-positive cells (expressed as the percentage of area stained with 3,3′-diaminobenzidine (DAB) as a function of distance from the microwell) is an example of a functional readout from an implantable drug delivery system. The agent concentration gradient forms at approximately 100-250 μm from the microwell, with the tissue concentration being greatest in the region closest to the microwell.

[0404] The system is used to deliver microdoses of ferroptosis-inducing agents to tissues in vivo. The system is also used to deliver a trigger (e.g., an anti-ferroptosis rescue agent (M)) to tissues in vivo, followed by delivery of a ferroptosis-inducing agent (e.g., a ferroptosis-inducing agent (C)) to induce targeted cell death in the tissue. The system is also implanted directly into tumors of about 6 millimeters (mm) to about 7 mm in diameter to achieve about 10 ng / mm at the microporous site. 2 The minimum amount of medication lasts at least 4 hours.

[0405] Example 7: Ferroptosis regulatory perturbations affect adipose lipid accumulation

[0406] Specific knockout of ferroptosis regulators that lead to ferroptosis was performed in mice. Figure 2As shown, compared with WT mice, the fat in mutant mice is significantly reduced. Compared with knockout mice, the amount of epididymal / gonadal white adipose tissue in WT mice is also significantly reduced. This reduction of adipose tissue can also be seen by pharmacological regulation. The disturbance is safe and tolerable. When mice are fed a high-fat diet (HFD), partial loss of function leads to a thin phenotype. It is known that constitutive activation of this pathway causes fatty liver and hepatomegaly in mice.

[0407] Example 8: Treatment with ferroptosis agents recapitulates weight loss phenotype

[0408] Male C57Bl / 6 mice (5-6 weeks old) were treated with a ferroptosis agent (5 mg / mL) together with a glutamate-cysteine ​​ligase inhibitor to show weight loss. Glutathione (GSH) is synthesized by the sequential action of the rate-limiting glutamate-cysteine ​​ligase (GCL) and glutathione synthase. Figures 4A-4C The results showed that mice treated with ferroptosis agents had reduced weight change (percentage) as well as reduced epididymal white adipose tissue (eWAT) weight (g) and the ratio of eWAT weight to body weight (g / g). Figure 4B Shown is the difference in water intake between the control and ferroptosis-treated groups, as well as the reduction in hepatic GSH levels in ferroptosis-treated mice compared to the control group that drank only water ( Figure 4D ). The data suggest that inhibition of glutamate-cysteine ​​ligase can lead to weight loss.

[0409] Similarly, Figure 5 Shown are histological samples of male C57Bl / 6 mice (4-5 weeks old) treated with 5 mg / mL ferroptosis agent for 7 days, and histological samples of control mice treated with water. In the treated samples, reduced fat was observed in the histological samples.

[0410] Example 9: Ferroptosis Treatment of Mice on a High Fat Diet (HFD) Male C57Bl / 6 mice and diet-induced obese (DIO) mice were treated with 5 mg / mL of ferroptosis to show weight loss in mice on a high fat diet. The mice were 8-week-old male mice. The mice were acclimated to the facility for one week. Ferroptosis treatment was started 2 days prior to HFD diet supplementation. Figures 6A-6C It was shown that mice treated with ferroptosis agents (C57Bl / 6 and DIO mice) had reduced weight change (percentage) and reduced epididymal white adipose tissue (eWAT) weight (g). In addition, Figure 6D-6E The results showed that triglyceride levels were reduced in DIO-treated mice and LDL cholesterol levels were increased in C57Bl / 6 mice treated with ferroptosis agents. The data showed that mice receiving a HFD diet did not gain significant weight when treated with ferroptosis agents.

Claims

1. A method of inducing weight loss, fat loss, or both in a subject, the method comprising administering to the subject an amount of a ferroptosis-inducing agent sufficient to induce weight loss, fat loss, or both.

2. The method of claim 1, wherein the method is a method of inducing weight loss.

3. The method of claim 1, wherein the method is a method for inducing fat loss.

4. The method of claim 1, wherein the method is a method of inducing weight loss and fat loss.

5. The method of claim 1, further comprising administering a trigger to the subject before administering the ferroptosis-inducing agent to the subject.

6. The method of claim 1, wherein after administering the ferroptosis-inducing agent to the subject, the adipose tissue in the subject is reduced in weight, volume, or both relative to the weight, volume, or both of the adipose tissue in the subject before administration.

7. The method of claim 6, wherein after administering the ferroptosis-inducing agent to the subject, the adipose tissue in the subject is reduced in volume relative to the volume of adipose tissue before administration, as determined by a spring clip test, by a dual x-ray absorptiometry (DEXA) scan, waist circumference reduction measurement, a computer axial tomography (CAT) test, or any combination thereof.

8. The method of claim 6, wherein after administering the ferroptosis-inducing agent to the subject, the adipose tissue in the subject is reduced in weight relative to the weight of the adipose tissue before administration.

9. The method of claim 8, wherein the weight loss is determined as follows: the subject's total body weight is reduced after administration of the ferroptosis-inducing agent compared to the subject's total body weight before administration of the ferroptosis-inducing agent.

10. The method of claim 8, wherein the weight loss is determined by comparing DEXA scans of the subject obtained before and after administration of a ferroptosis-inducing agent to the subject.

11. The method of claim 1, further comprising reducing the cell size, cell volume, or both of multiple cells in the adipose tissue of the subject after administering a ferroptosis-inducing agent to the subject, compared to the cell size, cell volume, or both of the multiple cells in the adipose tissue of the subject before administration.

12. The method of claim 1, wherein the subject is a human.

13. The method of claim 1, wherein the ferroptosis-inducing agent is administered orally.

14. The method of claim 1, wherein the administering occurs for about 1 day, 2 days, 3 days, 4 days, 5 days, 1 week, 10 days, 2 weeks, 15 days, 20 days, 3 weeks, 25 days, 30 days, one month, two months, three months, four months, five months, six months, one year, two years, three years, four years, five years, lifetime, or as needed.

15. The method of claim 14, wherein the ferroptosis-inducing agent is administered in an amount ranging from about 0.1 ng to about 25,000 mg.

16. The method of claim 14, wherein the ferroptosis inducing agent is administered in an amount of about 1 ng, 10 ng, 100 ng, 1 microgram, 10 micrograms, 100 micrograms, 1 mg, 10 mg, 100 mg, 1 g, 2 g, 3 g, 4 g, 5 g, 6 g, 7 g, 8 g, 9 g, 10 g, 15 g, 20 g, or 25 g.

17. The method of claim 14, wherein the ferroptosis-inducing agent is administered in an amount ranging from about 1 ng / kg to 1,000 mg / kg, wherein mg is mg of the ferroptosis-inducing agent and kg is kg of the subject's body weight.

18. The method of claim 1, wherein the ferroptosis-inducing agent is administered orally.

19. The method of claim 1, wherein the ferroptosis inducing agent is administered by one of the following routes: oral administration, subcutaneous, intradermal, intramuscular, inhalation, intravenous, intraperitoneal, intracranial, intrathecal, transdermal, rectal, or direct administration to adipose tissue.

20. The method of claim 1, treating at least one of obesity, metabolic syndrome, elevated blood sugar, diabetes, type 2 diabetes, type 3 diabetes, insulin resistance, hypertension, cardiovascular disease, coronary artery disease, cerebrovascular disease, stroke, rheumatic heart disease, arteriosclerosis, atherosclerosis, liver disease, fatty liver disease, non-alcoholic fatty liver disease (NAFLD), non-alcoholic steatohepatitis (NASH), or any combination thereof.

21. The method of claim 1, wherein the subject is a human male.

22. The method of claim 1, wherein the subject is a female.

23. The method of claim 1, wherein the subject is in the range of about 1 year old to about 18 years old.

24. The method of claim 1, wherein the subject is in the range of about 18 to about 120 years old.

25. The method of claim 1, wherein the subject is a subject in need thereof.

26. The method of claim 1, wherein the ferroptosis inducing agent is contained in a pharmaceutical composition comprising a pharmaceutically acceptable excipient, carrier or diluent.

27. The method of claim 26, wherein the pharmaceutical composition is in unit dosage form.

28. The method of claim 1, further comprising administering to the subject another therapeutic agent.

29. The method of claim 28, wherein the other therapeutic agent is administered concurrently with the ferroptosis-inducing agent.

30. The method of claim 28, wherein the other therapeutic agent is administered concomitantly with the ferroptosis-inducing agent.

31. The method of claim 1, wherein the ferroptosis-inducing agent is administered directly to adipose tissue.

32. The method of claim 31, wherein the administering is continuous administration.

33. The method of claim 32, wherein the administration continues for about: 1 / 2 hours, 1 hour, 2 hours, 3 hours, 4 hours, 5 hours, 6 hours, 7 hours, 8 hours, 9 hours, 10 hours, 11 hours, 12 hours, 13 hours, 14 hours, 15 hours, 16 hours, 17 hours, 18 hours, 19 hours, 20 hours, 21 hours, 22 hours, 23 hours, 24 hours, 2 days, 3 days, 4 days, 5 days, 6 days or 1 week.

34. The method of claim 31, wherein the adipose tissue is heated to a temperature of about 23, 24, 25, 26, 27, 28, 29, 30, 31, 32 or 33 degrees Celsius.

35. The method of claim 31, wherein the ferroptosis-inducing agent is administered in the form of an aqueous solution.

36. The method of claim 35, wherein the concentration of the ferroptosis inducing agent in the aqueous solution is about 0.1 nM to about 500 μM.

37. The method of claim 1, wherein the administration results in a decrease in adipocyte size or adipocyte number in the subject, wherein the decrease is determined by measuring adipocyte size or adipocyte number before and after administration of a therapeutic amount of a ferroptosis modulator, optionally using microscopic evaluation of a biopsy sample.

38. The method of claim 1, wherein the method further comprises reducing epididymal white adipose tissue in the subject.

39. The method of claim 1, wherein the subject is overweight or obese.

40. The method of claim 1, wherein the subject is obese and has a BMI of about 30.0 or greater.

41. The method of claim 1, wherein the subject is overweight and has a BMI of about 25.0 to about 30.

42. The method of claim 28, wherein the second therapeutic agent has glucagon-like peptide-1 (GLP-1) receptor agonist activity.

43. The method of claim 42, wherein the second therapeutic agent comprises semaglutide, dulaglutide, liraglutide, exenatide, telportide, or any combination thereof.

44. A method for inducing, regulating or modulating weight loss, body composition or fat loss or fat loss in a subject, the method comprising administering to the subject a therapeutically effective amount of a ferroptosis modulator, a glutamate-cysteine ​​ligase (GCL) modulator, an agent that binds GCL, an agent that inhibits GCL, or any combination thereof, thereby inducing, regulating or modulating weight loss, body composition or fat loss or fat loss in the subject.

45. The method of claim 44, wherein the subject is a human.

46. ​​The method of claim 45, wherein the subject is male.

47. The method of claim 44, wherein the ferroptosis modulator, the glutamate-cysteine ​​ligase (GCL) modulator, the agent that binds to GCL, the agent that inhibits GCL, or any combination thereof is orally delivered as a solution having a concentration of at least about 5 mg of agent / mL of solution.

48. The method of claim 44, wherein the administering is discontinuous and the administering occurs daily for about 5 days, 10 days, 15 days, 20 days, 25 days, 30 days, two months, three months, four months, five months, six months, or one year.

49. The method of claim 1, wherein the ferroptosis inducing agent is selected from the group consisting of: (1S, 3R)-RSL3, ML-162, ML-210, JKE-1674, JKE-1716, erastin, jacaric acid, trigonelline, glutamate, sulfasalazine, auranofin, brucea bruceiol, sorafenib, sorafenib-d3, sorafenib tosylate, trigonelline, FIN56, FINO2, CIL56, dihydroisotanshinone I, GPX4-IN-3, sulfoximine, BSO, and salts of any of these.

50. The method of claim 1, wherein the administration results in a decrease in adipocyte size in the subject, wherein the decrease is determined by measuring adipocyte size before and after administration of the ferroptosis-inducing agent, optionally using microscopic evaluation of a biopsy sample.

51. The method of claim 1, wherein the administration results in a reduction in the number of adipocytes in the subject, wherein the reduction is determined by measuring adipocyte number before and after administration of a therapeutic amount of a ferroptosis modulator, optionally using microscopic evaluation of a biopsy sample.

52. The method of claim 50 or claim 51, wherein the method comprises reducing epididymal white adipose tissue in the subject.

53. The method of claim 52, wherein the subject is overweight or obese.

54. The method of claim 53, wherein the subject is obese and has a BMI of about 30.0 or greater.

55. The method of claim 53, wherein the subject is overweight and has a BMI of about 25.0 to about 30.

56. The method of any preceding claim, wherein an additional agent or therapy is administered to the subject concurrently or consecutively.

57. The method of claim 56, wherein the additional agent or therapy is selected from the group consisting of a GLP-1 agonist, a gastric inhibitory polypeptide analog, naltrexone-bupropion or a salt thereof, orlistat or a salt thereof, phentermine or a salt thereof, phentermine-topiramate or a salt thereof, selenium, exercise therapy, surgery, dietary modification, and any combination thereof.

58. The method of claim 57, wherein the additional agent or therapy comprises administration of a GLP-1 agonist comprising dulaglutide, exenatide, semaglutide, liraglutide, lixisenatide, or a salt of any of these.

59. The method of claim 57, wherein the additional agent or therapy comprises administration of a gastric inhibitory polypeptide comprising telportide or a derivative thereof.

60. The method of claim 57, wherein the additional agent or therapy comprises a surgical procedure comprising a gastric sleeve, a gastric bypass, an adjustable gastric band, a balloon placed in the stomach, or any combination thereof.

61. The method of any preceding claim, wherein the subject has not been diagnosed with cancer.

62. The method of any preceding claim, wherein the subject is a subject in need thereof.

63. The method of any preceding claim, wherein the subject has Type I diabetes, Type II diabetes, Type IIIa diabetes, or metabolic syndrome.

64. The method of claim 44, wherein the ferroptosis regulator, the glutamate-cysteine ​​ligase (GCL) regulator, the agent that binds to GCL, the agent that inhibits GCL, or any combination thereof is in the form of a pharmaceutical composition, and the pharmaceutical composition further comprises a pharmaceutically acceptable carrier, excipient, or diluent.

65. The method of claim 64, wherein the pharmaceutical composition is in unit dosage form.

66. A composition comprising i) a ferroptosis regulator, a glutamate-cysteine ​​ligase (GCL) regulator, an agent that binds to GCL, an agent that inhibits GCL; and ii) a GLP-1 agonist or a salt thereof, a gastric inhibitory polypeptide analog, naltrexone-bupropion or a salt thereof, orlistat or a salt thereof, phentermine or a salt thereof, and phentermine-topiramate or a salt thereof, or selenium or a salt thereof.

67. A composition comprising i) a glutamate-cysteine ​​ligase (GCL) modulator, and ii) a GLP-1 agonist or a salt thereof, a gastric inhibitory polypeptide analog, naltrexone-bupropion or a salt thereof, orlistat or a salt thereof, phentermine or a salt thereof, and phentermine-topiramate or a salt thereof, or selenium or a salt thereof.

68. The composition of claim 66 or claim 67 which is a pharmaceutical composition.

69. The pharmaceutical composition of claim 68 in unit dosage form.

70. The composition of any one of claims 66-69 for use in treating a disease or condition.

71. A composition according to any one of claims 66-69 for use in treating a kidney disease or disorder.

72. A composition according to any one of claims 66-69 for use in treating symptoms associated with a kidney disease or disorder.

73. The composition for use of claim 70, wherein the disease or condition is weight loss, weight maintenance, a disease or condition related to iron metabolism, or a disease or condition associated with high, low or abnormal cholesterol.

74. A method for regulating iron metabolism in multiple cells in a subject, the method comprising administering to the subject a therapeutically effective amount or an effective amount of a ferroptosis inducer, a glutamate-cysteine ​​ligase (GCL) regulator, an agent that binds to GCL, an agent that inhibits GCL, or any combination thereof, thereby regulating iron metabolism in the subject and thereby inducing ferroptosis in the subject.

75. The method of claim 74, wherein immediately following the modulation, a majority of the cells in the plurality of cells remain viable.

76. A method of regulating iron metabolism in a plurality of cells in a subject, the method comprising administering to the subject a therapeutically effective amount or an effective amount of a ferroptosis-inducing agent, thereby regulating iron metabolism in the subject.

77. The method of claim 76, wherein immediately following the modulation, a majority of the cells in the plurality of cells remain viable.

78. A method for treating, inducing, regulating or adjusting iron metabolism or a disease or condition associated with iron metabolism in a subject, the method comprising administering to the subject a therapeutically effective amount or an effective amount of a ferroptosis regulator, a glutamate-cysteine ​​ligase (GCL) regulator, an agent that binds to GCL, an agent that inhibits GCL, or any combination thereof, thereby treating, inducing, regulating or adjusting iron metabolism or a disease or condition associated with iron metabolism in the subject.

79. The method of claim 78, wherein the subject is a human.

80. The method of claim 78, wherein the subject is a subject in need thereof.

81. The method of claim 78, wherein a therapeutically effective amount of a ferroptosis modulator is added.

82. A method for regulating cholesterol metabolism in a subject, the method comprising administering to the subject a therapeutically effective amount or an effective amount of a ferroptosis regulator, a glutamate-cysteine ​​ligase (GCL) regulator, an agent that binds to GCL, an agent that inhibits GCL, or any combination thereof, thereby regulating cholesterol metabolism in the subject.

83. The method of claim 82, wherein the subject is a human.

84. The method of claim 82, wherein the subject is a subject in need thereof.

85. The method of claim 84, wherein the ferroptosis-inducing agent is administered.

86. A method of regulating cholesterol metabolism in a subject, the method comprising administering to the subject a therapeutically effective amount or an effective amount of a ferroptosis-inducing agent, thereby regulating cholesterol metabolism in the subject.

87. The method of claim 86, wherein the subject is a human.

88. The method of claim 86, wherein the subject is a subject in need thereof.

89. The method of claim 86, wherein the ferroptosis-inducing agent is administered.

90. A method for treating, inducing, regulating, adjusting or reducing cholesterol metabolism or a disease related to cholesterol metabolism in a subject, the method comprising administering to the subject a therapeutically effective amount or an effective amount of a ferroptosis regulator, a glutamate-cysteine ​​ligase (GCL) regulator, an agent that binds to GCL, an agent that inhibits GCL, or any combination thereof, thereby treating, inducing, regulating, adjusting or reducing cholesterol metabolism or a disease or condition related to cholesterol metabolism in the subject.

91. The method of claim 90, wherein the subject is a human.

92. The method of claim 90, wherein the subject is a subject in need thereof.

93. The method of claim 90, wherein the ferroptosis modulator is administered.

94. A method for treating, inducing, regulating, adjusting, reducing or increasing reactive oxygen species or a disease or condition associated with reactive oxygen species in a subject, the method comprising administering to the subject a therapeutically effective amount or an effective amount of a ferroptosis regulator, a glutamate-cysteine ​​ligase (GCL) regulator, an agent that binds to GCL, an agent that inhibits GCL, or any combination thereof, thereby treating, inducing, regulating, adjusting, reducing or increasing reactive oxygen species or a disease or condition associated with reactive oxygen species in the subject.

95. The method of claim 94, wherein the subject is a human.

96. The method of claim 94, wherein the subject is a subject in need thereof.

97. The method of claim 94, wherein the ferroptosis modulator is administered.

98. A method for increasing reactive oxygen species in a subject, the method comprising administering to the subject a therapeutically effective amount or an effective amount of a ferroptosis regulator, a glutamate-cysteine ​​ligase (GCL) regulator, an agent that binds to GCL, an agent that inhibits GCL, or any combination thereof, thereby increasing reactive oxygen species in the subject.

99. The method of claim 98, wherein the subject is a human.

100. The method of claim 98, wherein the subject is a subject in need thereof.

101. The method of claim 98, wherein the ferroptosis modulator is administered.

102. A method for promoting at least one of the following: treating, overcoming, regulating or reducing acute stress; a chronic stress response; an immune hypersensitivity reaction; cachexia; kidney disease; neurodegeneration; cardiotoxicity; or cardiotoxicity caused at least in part by chemotherapy in a subject; the method comprising administering to the subject a therapeutically effective amount or an effective amount of a ferroptosis regulator, a glutamate-cysteine ​​ligase (GCL) regulator, an agent that binds to GCL, an agent that inhibits GCL, or any combination thereof, thereby treating, inducing, regulating, adjusting, reducing or increasing acute stress; a chronic stress response; an immune hypersensitivity reaction; cachexia; kidney disease; neurodegeneration; cardiotoxicity; or cardiotoxicity caused at least in part by chemotherapy in the subject.

103. The method of claim 102, wherein the subject is a human.

104. The method of claim 102, wherein the subject is a subject in need thereof.

105. The method of claim 102, wherein the ferroptosis modulator is administered.

106. A method for promoting at least one of the following: treating kidney disease in a subject, the method comprising administering to the subject a therapeutically effective amount or an effective amount of a ferroptosis regulator, a glutamate-cysteine ​​ligase (GCL) regulator, an agent that binds GCL, an agent that inhibits GCL, or any combination thereof, thereby treating the kidney disease in the subject.

107. The method of claim 106, wherein the subject is a human.

108. The method of claim 106, wherein the subject is a subject in need thereof.

109. The method of claim 106, wherein the ferroptosis modulator is administered.

110. A method of promoting at least one of the following: treating kidney disease in a subject, the method comprising administering to the subject a therapeutically effective amount or an effective amount of a ferroptosis-inducing agent, thereby treating the kidney disease in the subject.

111. The method of claim 110, wherein the subject is a human.

112. The method of claim 110, wherein the subject is a subject in need thereof.

113. The method of claim 110, wherein the ferroptosis-inducing agent is administered.

114. The method of any one of claims 74-110, wherein the method comprises reducing epididymal white adipose tissue in the subject.

115. The method of claim 114, wherein the subject is overweight or obese.

116. The method of claim 115, wherein the subject is obese and has a BMI of 30.0 or greater.

117. The method of claim 115, wherein the subject is overweight and has a BMI of 25.0 to <30.

118. The method of any one of claims 74-117, wherein an additional agent or therapy is administered to the subject concurrently or consecutively.

119. The method of claim 118, wherein the additional agent or therapy is selected from the group consisting of a GLP-1 receptor agonist, a gastric inhibitory polypeptide analog, naltrexone-bupropion or a salt thereof, orlistat or a salt thereof, phentermine or a salt thereof, phentermine-topiramate or a salt thereof, selenium, exercise therapy, surgery, dietary modification, and any combination thereof.

120. The method of claim 119, wherein the additional agent or therapy comprises administration of a GLP-1 receptor agonist comprising dulaglutide, exenatide, semaglutide, liraglutide, lixisenatide, or a salt of any of these.

121. The method of claim 119, wherein the additional agent or therapy comprises administration of a gastric inhibitory polypeptide comprising telportide or a derivative thereof.

122. The method of claim 119, wherein the additional agent or therapy comprises a surgical procedure comprising a gastric sleeve, a gastric bypass, an adjustable gastric band, a balloon placed in the stomach, or any combination thereof.

123. The method of any one of claims 74-122, wherein the subject has not been diagnosed with cancer.

124. The method of any one of claims 74-122, wherein the subject is a subject in need thereof.

125. The method of any one of claims 74-122, wherein the subject has Type I diabetes, Type II diabetes, Type IIIa diabetes, or metabolic syndrome.

126. The method of any one of claims 74-122, wherein the agent is in the form of a pharmaceutical composition further comprising a pharmaceutically acceptable: carrier, excipient, or diluent.

127. The method of claim 126, wherein the pharmaceutical composition is in unit dosage form.

128. The method of any preceding claim, wherein the agent is a compound in Table 1 or a salt thereof.

129. The method of any preceding claim, wherein the agent is in the form of a pharmaceutical composition further comprising a pharmaceutically acceptable: carrier, excipient, or diluent.

130. The method of claim 46, wherein the pharmaceutical composition is in unit dosage form.

131. A composition comprising i) a ferroptosis regulator, a glutamate-cysteine ​​ligase (GCL) regulator, an agent that binds to GCL, an agent that inhibits GCL; and ii) a GLP-1 agonist or a salt thereof, a gastric inhibitory polypeptide analog, naltrexone-bupropion or a salt thereof, orlistat or a salt thereof, phentermine or a salt thereof, and phentermine-topiramate or a salt thereof, or selenium or a salt thereof.

132. The composition of claim 131, which is a pharmaceutical composition.

133. The pharmaceutical composition of claim 132 in unit dosage form.

134. The composition of any one of claims 131-133 for use in treating a disease or condition.

135. The composition of any one of claims 131-133 for use in treating a kidney disease or disorder.

136. The composition of any one of claims 131-133 for use in treating symptoms associated with a kidney disease or disorder.

137. The composition for use of claim 134, wherein the disease or condition is weight loss, weight maintenance, a disease or condition related to iron metabolism, or a disease or condition associated with high, low or abnormal cholesterol.

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