How to protect your heart

Abstract

The present invention relates to a method for preventing or reducing drug-induced cardiotoxicity in a subject caused by a cardiotoxic agent, and a composition and a kit therefor, comprising administering to the subject an effective amount of a cardioprotectant comprising bisantrene or a derivative thereof, or a pharma- ceutically acceptable salt of bisantrene or a derivative thereof. Also provided are such methods, compositions and kits for the treatment of cancer, and the use of such compositions for the manufacture of a medicament with reduced cardiotoxicity.

Description

[Technical Field]

[0001] The present disclosure relates to methods for preventing or reducing cardiotoxicity in a subject, and pharmaceutical compositions and kits for preventing or reducing cardiotoxicity in a subject. [Background technology]

[0002] Cardiotoxicity is damage to the cells of the heart muscle caused by cardiotoxic agents, such as cardiotoxic chemotherapy agents, and can result in symptoms such as blood pressure changes, thrombosis, electrocardiogram changes, arrhythmias, myocarditis, pericarditis, myocardial infarction, cardiomyopathy, heart failure (left ventricular failure), and congestive heart failure.

[0003] Many chemotherapy drugs for the treatment of cancer and other conditions also exhibit significant cardiotoxicity.

[0004] The drugs most associated with cardiotoxicity include anthracyclines, some tyrosine kinase inhibitors (TKIs), and some biologics based on monoclonal antibodies.

[0005] Anthracyclines are chemotherapy drugs widely used in the treatment of cancer. Anthracyclines are among the most effective classes of anticancer drugs developed to date, and are more effective against more types of cancer than many other classes of chemotherapy drugs. Cancers treated with anthracyclines include leukemia, lymphoma, breast cancer, gastric cancer, uterine cancer, ovarian cancer, bladder cancer, and lung cancer. Anthracycline drugs include, for example, doxorubicin, daunorubicin, epirubicin, valrubicin, mitoxantrone, and idarubicin.

[0006] Despite their effectiveness, the use of anthracyclines as chemotherapeutic agents is limited by their cardiotoxicity. Anthracyclines exhibit dose-dependent and cumulative cardiotoxicity, and as a result, the maximum recommended cumulative dose must be set to prevent the development of adverse cardiac events, such as congestive heart failure. For example, Non-Patent Document 1 reports that patients should receive 400 mg / m 2, 550 mg / m 2 and 700 mg / m 2 reported that the incidence of congestive heart failure was 4.7%, 26%, and 48%, respectively, when patients received 100 mg / m² of doxorubicin. Therefore, to reduce the incidence of congestive heart failure to less than 5%, a lifetime cumulative doxorubicin exposure of 400–450 mg / m² should be considered. 2 Although doxorubicin is limited to 20 mg / kg / day, there is inter-individual variability in tolerance to doxorubicin (Non-Patent Document 2). A similar situation exists with daunorubicin, although cardiac toxicity is slightly lower. The need to limit cumulative exposure to anthracyclines may limit the full potential of these drugs for treating cancer. Furthermore, even at current limited doses, cardiac damage may occur in some patients.

[0007] In addition to anthracyclines, some tyrosine kinase inhibitors (TKIs) and antibody-based therapeutics have also been reported to be cardiotoxic. TKIs such as imatinib mesylate (Gleevec), nilotinib (Tasigna), sorafenib (Nexavar), sunitinib (Sutent), and dasatinib (Sprycel) have been reported to cause cardiotoxicity (Non-Patent Literatures 3-5). The antibody-based therapeutic trastuzumab is an example of a monoclonal antibody that can cause cardiotoxicity.

[0008] Proteasome inhibitors are also known to have cardiotoxic side effects. For example, carfilzomib is a tetrapeptide epoxyketone and epoxomicin analogue that acts as a selective irreversible proteasome inhibitor and is used as an anticancer drug for multiple myeloma. However, carfilzomib may cause cardiotoxicity in up to 10% of patients, and due to this cardiotoxicity, it is not used in solid tumors such as breast cancer. Several reports have also suggested the cardiotoxicity of bortezomib (Velcade®).

[0009] Risk factors can predispose a patient to cardiotoxicity, including, for example, cumulative dose of cardiotoxic agent, course of administration of cardiotoxic agent, rate of administration of cardiotoxic agent, concomitant administration of cardiotoxic agent, previous treatment with cardiotoxic agent, and history of cardiovascular disorder or pre-existing cardiovascular disorder.

[0010] Several cardioprotective strategies have been investigated, including the use of modified anthracyclines, statins, cardioselective beta-blockers, and angiotensin antagonists, however, results have been mixed in humans.

[0011] It would be advantageous to develop improved cardioprotective strategies to reduce or prevent cardiotoxicity caused by cardiotoxic agents. [Prior art documents] [Non-patent literature]

[0012] [Non-Patent Document 1] McGowan et al. (2017) Cardiovascular Drugs and Therapy, 31(1):63~75 [Non-patent document 2] Ewer & Ewer(2015), Nature Reviews Cardiology.12(9): 547-558 [Non-patent document 3] Chu et al., Lancet(2007) 370:2011-2019 [Non-patent document 4] Xu et al.,Hematol Rev.(2009) Mar 1; 1(1): e4 [Non-patent document 5] Kerketla et al., Nature Medicine (2006) 12:908-916 Summary of the Invention

[0013] Surprisingly, this study showed that bisantrene, an anthracycline-like anthracene previously known to have anti-tumorigenic properties, is not only toxic to cancer cells alone or at least additively, if not synergistically, when combined with other chemotherapeutic agents, but also reduces the toxicity to human cardiomyocytes of cardiotoxic agents including anthracyclines, the anti-tumor monoclonal antibody trastuzumab, and the proteasome inhibitors carfilzomib and bortezomib, even though it is mildly toxic to human cardiomyocytes itself.

[0014] These properties of bisantrene allow for improved efficacy in treating subjects with cardiotoxic therapeutic agents with reduced cardiotoxicity, and for reduced administration of cardiotoxic therapeutic agents while achieving substantially the same therapeutic effect as achieved with full administration (but without bisantrene). These benefits are particularly important with respect to the use of drugs such as anthracyclines, which have lifetime dosing limits. The enhanced antitumor activity and avoidance of cardiotoxicity offered by bisantrene should also enable more widespread use of chemotherapy drugs such as carfilzomib, which have historically been too toxic for use in cancers other than multiple myeloma.

[0015] Accordingly, a first aspect of the present invention provides a method for preventing or reducing drug-induced cardiotoxicity in a subject caused by a cardiotoxic agent, the method comprising the step of administering to the subject an effective amount of a cardioprotective agent comprising bisantrene or a derivative thereof, or a pharmaceutically acceptable salt of bisantrene or a derivative thereof.

[0016] An alternative first aspect provides a cardioprotective agent comprising bisantrene or a derivative thereof, or a pharmaceutically acceptable salt of bisantrene or a derivative thereof, for use in preventing or reducing drug-induced cardiotoxicity in a subject caused by a cardiotoxic agent; or use of a cardioprotective agent comprising bisantrene or a derivative thereof, or a pharmaceutically acceptable salt of bisantrene or a derivative thereof, in the manufacture of a medicament for preventing or reducing drug-induced cardiotoxicity in a subject caused by a cardiotoxic agent.

[0017] A second aspect provides a method for preventing or reducing cardiotoxicity induced or exacerbated by a cardiotoxic agent in a subject, the method comprising administering to the subject an effective amount of a cardioprotective agent comprising bisantrene or a derivative thereof, or a pharmaceutically acceptable salt of bisantrene or a derivative thereof, before, simultaneously with, or after administration of the cardiotoxic agent.

[0018] An alternative second aspect provides a cardioprotective agent comprising bisantrene or a derivative thereof, or a pharmaceutically acceptable salt of bisantrene or a derivative thereof, for use in preventing or reducing cardiotoxicity induced or exacerbated by a cardiotoxic agent in a subject; or the use of a cardioprotective agent comprising bisantrene or a derivative thereof, or a pharmaceutically acceptable salt of bisantrene or a derivative thereof, in the manufacture of a medicament for preventing or reducing cardiotoxicity induced or exacerbated by a cardiotoxic agent in a subject.

[0019] A third aspect provides a method for treating cancer in a subject, comprising: a. a cardiotoxic chemotherapeutic agent in an amount therapeutically effective against cancer, and b. A cardioprotective agent comprising a cardioprotective amount of bisantrene or a derivative thereof, or a pharmaceutically acceptable salt of bisantrene or a derivative thereof. The method includes administering

[0020] An alternative third aspect provides a cardioprotective agent and a cardiotoxic chemotherapeutic agent comprising bisantrene or a derivative thereof, or a pharmaceutically acceptable salt of bisantrene or a derivative thereof, for use in treating cancer in a subject; or the use of a cardioprotective agent and a cardiotoxic chemotherapeutic agent comprising bisantrene or a derivative thereof, or a pharmaceutically acceptable salt of bisantrene or a derivative thereof, in the manufacture of a medicament for treating cancer in a subject.

[0021] A fourth aspect provides a method for preventing or reducing cardiotoxicity in a subject being treated with a cardiotoxic chemotherapeutic agent, the method comprising administering to the subject an effective amount of a cardioprotective agent comprising bisantrene or a derivative thereof, or a pharmaceutically acceptable salt of bisantrene or a derivative thereof.

[0022] An alternative fourth aspect provides a cardioprotective agent comprising bisantrene or a derivative thereof, or a pharmaceutically acceptable salt of bisantrene or a derivative thereof, for use in preventing or reducing cardiotoxicity in a subject being treated with a cardiotoxic chemotherapeutic agent; or the use of a cardioprotective agent comprising bisantrene or a derivative thereof, or a pharmaceutically acceptable salt of bisantrene or a derivative thereof, in the manufacture of a medicament for preventing or reducing cardiotoxicity in a subject being treated with a cardiotoxic chemotherapeutic agent.

[0023] A fifth aspect provides a pharmaceutical composition comprising a cardioprotective agent comprising bisantrene or a derivative thereof, or a pharmaceutically acceptable salt of bisantrene or a derivative thereof, and a cardiotoxic therapeutic agent, wherein the composition has reduced cardiotoxicity compared to a composition comprising the cardiotoxic therapeutic agent but not the cardioprotective agent.

[0024] A sixth aspect provides a pharmaceutical composition comprising: (a) a cardioprotective agent comprising bisantrene or a derivative thereof, or a pharmaceutically acceptable salt of bisantrene or a derivative thereof; and (b) a cardiotoxic chemotherapeutic agent.

[0025] A seventh aspect provides a kit for preventing or reducing drug-induced cardiotoxicity in a subject, the kit comprising: (a) a cardioprotective agent comprising bisantrene or a derivative thereof, or a pharmaceutically acceptable salt of bisantrene or a derivative thereof; and (b) a cardiotoxicity therapeutic agent.

[0026] An eighth aspect provides a kit for treating cancer, comprising: (a) a cardioprotective agent comprising bisantrene or a derivative thereof, or a pharmaceutically acceptable salt of bisantrene or a derivative thereof; and (b) a cardiotoxic chemotherapeutic agent. [Brief explanation of the drawings]

[0027] [Figure 1] Viability of primary human cardiomyocytes cultured in the presence of 0-5 μM bisantrene alone or in the presence of 1 μM doxorubicin or 1 μM carfilzomib for (A) 24 hours, (B) 48 hours, and (C) 72 hours. [Figure 2] 1 shows the viability of primary human cardiomyocytes when cultured in the presence of 1 μM doxorubicin alone; 1 μM doxorubicin and 500 nM trastuzumab; 1 μM doxorubicin, 500 nM trastuzumab and 1 μM bisantrene; 1 μM doxorubicin and 1 μM bisantrene; 1 μM doxorubicin, 500 nM trastuzumab and 5 μM bisantrene; and 1 μM doxorubicin, 500 nM trastuzumab and 5 μM bisantrene. [Figure 3] 1 shows the viability of MCF-7 breast cancer cells when cultured in the presence of 0-10 μM bisantrene alone, or in the presence of 1 μM doxorubicin or 1 μM carfilzomib over a 72 hour period. [Figure 4] 1 shows the viability of H929 multiple myeloma cells when cultured in the presence of 0-5 μM bisantrene alone or in addition to 1 μM carfilzomib over a 72 hour period. [Figure 5]1 shows the viability of primary human cardiomyocytes when cultured in the presence of 1 μM bisantrene, 1 μM daunorubicin, or 1 μM bisantrene and 1 μM daunorubicin for 72 hours. [Figure 6] 1 shows the viability of primary human cardiomyocytes when cultured in the presence of 1 μM doxorubicin, 1 μM doxorubicin and 1 μM bisantrene, 1 μM epirubicin, or 1 μM epirubicin and 1 μM bisantrene for 72 hours. [Figure 7] Figure 1 shows the viability of primary human cardiomyocytes when cultured in the presence of approved proteasome inhibitors or in addition to 1 μM bisantrene at the concentrations listed over a 72-hour period. [Figure 8] (A) Expected additive effects of bisantrene and doxorubicin at concentrations of 0-2 μM bisantrene alone, 0-1 μM doxorubicin alone, and 0-2 μM bisantrene in combination with 0-1 μM doxorubicin; (B) Expected additive effects of bisantrene and doxorubicin at concentrations of 0-2 μM bisantrene alone, 0-500 nM doxorubicin alone, and 0-2 μM bisantrene in combination with 0-500 nM doxorubicin; and (C) Expected additive effects of bisantrene and doxorubicin at concentrations of 0-1 μM bisantrene alone, 0-1 μM doxorubicin alone, and 0-1 μM bisantrene in combination with 0-1 μM doxorubicin. [Figure 9](A) Expected additive effects of bisantrene and epirubicin at concentrations of 0-2 μM bisantrene alone, 0-1 μM epirubicin alone, and 0-2 μM bisantrene in combination with 0-1 μM epirubicin, respectively; (B) Expected additive effects of bisantrene and epirubicin at concentrations of 0-2 μM bisantrene alone, 0-500 nM epirubicin alone, and 0-2 μM bisantrene in combination with 0-500 nM epirubicin, respectively; and (C) Expected additive effects of bisantrene and epirubicin at concentrations of 0-1 μM bisantrene alone, 0-1 μM epirubicin alone, and 0-1 μM bisantrene in combination with 0-1 μM epirubicin, respectively. [Figure 10] (A) 0–250 nM bisantrene alone, 0–250 nM bisantrene in the presence of 3.9 nM doxorubicin, 0–250 nM bisantrene in the presence of 7.8 nM doxorubicin, 0–250 nM bisantrene in the presence of 15 nM doxorubicin, 0–250 nM bisantrene in the presence of 31.25 nM doxorubicin, 0–250 nM bisantrene in the presence of 62.5 nM doxorubicin; (B) 0–250 nM bisantrene alone, 0–250 nM bisantrene in the presence of 3.9 nM epirubicin, 0–250 nM bisantrene in the presence of 7.8 nM epirubicin, 0–250 nM bisantrene in the presence of 15 nM epirubicin. (C) MDA-MB-231 breast cancer cell viability when cultured in the presence of 0–250 nM bisantrene, 0–250 nM bisantrene in the presence of 31.25 nM epirubicin, and 0–250 nM bisantrene in the presence of 62.5 nM epirubicin; (D) 0–250 nM bisantrene alone, 0–250 nM bisantrene in the presence of 0.625 nM carfilzomib, 0–250 nM bisantrene in the presence of 1.25 nM carfilzomib, 0–250 nM bisantrene in the presence of 2.5 nM carfilzomib, 0–250 nM bisantrene in the presence of 5 nM carfilzomib, and 0–250 nM bisantrene in the presence of 10 nM carfilzomib. [Figure 11]Figure 1 shows the viability of primary human cardiomyocytes cultured in the presence of 0-10 μM of the FTO inhibitor FB23 / 2 alone or in the presence of 1 μM doxorubicin or 1 μM carfilzomib over 72 hours. [Figure 12] Primary human cardiomyocyte viability is shown when cultured in the presence of 0-10 μM of the FTO inhibitor brequinar alone or in the presence of 1 μM doxorubicin or 1 μM carfilzomib over (A) 24 hours; (B) 48 hours; and (C) 72 hours. [Figure 13] Figure 1 shows MCF-7 breast cancer cell viability when cultured in the presence of FTO inhibitors FB23 / 2 (A) or brequinar (B), FTO inhibitors alone, or in the presence of 1 μM doxorubicin over a 72-hour period. [Figure 14] (A) The effect of bisantrene on left ventricular fractional shortening in mice when administered alone or in combination with doxorubicin. (B) The effect of bisantrene on cardiac E / A ratio in mice when administered alone or in combination with doxorubicin. Vehicle alone; bisantrene alone at 7.33 mg / kg once weekly for 4 weeks; doxorubicin alone at 5 mg / kg once weekly for 4 weeks (■); doxorubicin at 5 mg / kg:3.67 mg / kg once weekly for 4 weeks (Dox+Bis 1:1); doxorubicin at 5 mg / kg:7.33 mg / kg once weekly for 4 weeks (Dox+Bis 1:2); doxorubicin at 4 mg / kg:5.65 mg / kg once weekly for 4 weeks (Dox+Bis 80:20). *: p<0.001, Dox+vehicle vs. vehicle on days 21 and 42. #: p<0.05, Dox+Bis 1:1 vs. Dox+vehicle on days 21 and 42. &: p<0.01, Dox+Bis 1:2 vs. Dox+vehicle on day 42 in (A), or p<0.05, Dox+Bis 1:2 vs. Dox+vehicle on day 42 in (B). [Figure 15](A) The effect of bisantrene on left ventricular diastolic function in mice when administered with or without doxorubicin; (B) The effect of bisantrene on left ventricular diastolic function in mice when administered with or without doxorubicin; (C) The effect of bisantrene on cardiac output in mice when administered with or without doxorubicin. Vehicle alone; bisantrene alone at 7.33 mg / kg once weekly for 4 weeks; doxorubicin alone at 5 mg / kg once weekly for 4 weeks (■); doxorubicin at 5 mg / kg once weekly for 4 weeks: bisantrene at 3.67 mg / kg (Dox+Bis 1:1); doxorubicin at 5 mg / kg once weekly for 4 weeks: bisantrene at 7.33 mg / kg (Dox+Bis 1:2); doxorubicin at 4 mg / kg once weekly for 4 weeks: bisantrene at 5.65 mg / kg (Dox+Bis 80:20). *: p<0.01, Dox+vehicle vs. vehicle on day 42. #: p<0.05 in (A) and (B), Dox+Bis 1:1 vs. Dox+vehicle on day 42; p<0.01 in (C), Dox+Bis 1:1 vs. Dox+vehicle on day 42. &: p<0.09 in (A), Dox+Bis 1:2 vs. Dox+vehicle on day 42; p<0.19 in (B), Dox+Bis 1:2 vs. Dox+vehicle on day 42; p<0.01 in (B), Dox+Bis 1:2 vs. Dox+vehicle on day 42. [Figure 16](A) Effect of bisantrene on plasma triglyceride levels in mice when administered with doxorubicin alone or in combination with doxorubicin. *: p<0.05, Dox+Vehicle vs. Vehicle on days 21 and 42; #: p<0.05, Dox+Bis 1:1 vs. Dox+Vehicle on days 21 and 42; &: p<0.05, Dox+Bis 1:2 vs. Dox+Vehicle on day 42; (B) Effect of bisantrene on blood creatine kinase-MB (CK-MB) in mice when administered with doxorubicin alone or in combination with doxorubicin. *: p<0.01, Dox+Vehicle vs. Vehicle on day 21; #: p<0.05, Dox+Bis 1:1 vs. Dox+Vehicle on days 21 and 42; &: p<0.16, Dox+Bis 1:2 vs. Dox+Vehicle on day 21. (C) Effect of bisantrene on plasma brain natriuretic peptide (BNP) in mice when administered with doxorubicin alone or in combination with doxorubicin. *: p<0.05, Dox+Vehicle vs. Vehicle on days 21 and 42. #: p<0.05, Dox+Bis 1:1 vs. Dox+Vehicle on days 21 and 42. [Figure 17] (A) Effect of bisantrene on histological cardiac fibrosis in mice when administered with doxorubicin alone or in combination with doxorubicin. *: p<0.01, Dox+Vehicle vs. Vehicle on day 21; #: p<0.05, Dox+Vehicle 1:1 vs. Dox+Bis on day 21. DETAILED DESCRIPTION OF THE INVENTION

[0028] (definition) As used herein, "treating" means affecting a subject, tissue, or cell to obtain a desired pharmacological and / or physiological effect, including inhibiting a condition, i.e., slowing or arresting its development, or reducing or ameliorating the effects of a condition, i.e., causing a reversal or regression of the effects of a condition.

[0029] As used herein, "preventing" means preventing a condition from occurring in a cell, tissue, or subject that may be at risk of having the condition, but does not necessarily mean that the condition will not eventually develop or that the subject will not eventually develop the condition. Preventing includes delaying the onset of the condition in a cell, tissue, or subject.

[0030] As used herein, "reducing cardiotoxicity" means reducing cardiotoxicity in a subject compared to cardiotoxicity in a subject not treated to reduce cardiotoxicity as described herein. Reducing cardiotoxicity can include, for example, reducing the severity or number of symptoms presented compared to the severity or number of symptoms presented in an untreated response.

[0031] As used herein, "cardiotoxicity" refers to damage to cardiac tissue, such as damage to heart muscle cells. Typically, cardiotoxicity is induced or exacerbated by a cardiotoxic agent.

[0032] An agent is "cardioprotective" if it prevents or reduces cardiotoxicity. In the context of bisantrene derivatives and pharmaceutically acceptable salts thereof, it is not expected that all derivatives of bisantrene or its salts are cardioprotective. Rather, when reference is made to a "cardioprotectant comprising bisantrene or a derivative thereof, or a pharmaceutically acceptable salt of bisantrene or a derivative thereof," only derivatives or pharmaceutically acceptable salts thereof that are cardioprotective are contemplated by the present invention.

[0033] As used herein, the term "subject" refers to a mammal. A mammal can be a human or a non-human. Examples of non-humans include primates, livestock animals (e.g., sheep, cows, horses, donkeys, pigs), companion animals (e.g., dogs, cats), laboratory test animals (e.g., mice, rabbits, rats, guinea pigs, hamsters), and captive wild animals (e.g., foxes, deer). Typically, the mammal is a human or a non-human primate. More typically, the mammal is a human.

[0034] The term "composition" encompasses compositions and formulations that include an active pharmaceutical ingredient ("bisantrene or a cardioprotective derivative of bisantrene, or a pharmaceutically acceptable salt of bisantrene or a cardioprotective derivative of bisantrene") together with an excipient or carrier, as well as compositions and formulations that include a carrier. In a pharmaceutical composition, the excipient or carrier is "pharmaceutically acceptable," meaning that it is not biologically or otherwise undesirable; i.e., the material can be incorporated into a pharmaceutical composition administered to a patient without causing any undesirable biological effects or interacting in a deleterious manner with any of the other components of the composition in which it is included. Supplementary active ingredients can also be incorporated into the composition.

[0035] "Pharmaceutically acceptable," as in the description of a "pharmaceutically acceptable salt" or a "pharmaceutically acceptable excipient or carrier," as used herein means a substance that is not biologically or otherwise undesirable, i.e., the substance can be incorporated into a pharmaceutical composition administered to a patient without causing any undesired biological effects or interacting in a deleterious manner with any of the other components of the composition in which it is included.

[0036] The term "effective amount" or "therapeutically effective amount" means an amount of an active pharmaceutical ingredient sufficient to bring about the desired therapeutic response. The specific effective amount or therapeutically effective amount will vary depending on factors such as the particular condition being treated, the age, weight, general health, physical condition, sex, and diet of the subject, the duration of treatment, the nature of concomitant therapy (if any), and the severity of the particular condition.

[0037] As used herein, "carrier" includes any and all solvents, dispersion media, vehicles, coatings, diluents, fillers, carrier solutions, suspensions, colloids, and formers and binders, any or all of which may include other pharmaceutical excipients known in the art, including lubricants, antibacterial and antifungal agents, isotonic and absorption delaying agents, buffers, antioxidants, other stabilizers including physical stabilizers such as thickeners and viscosity enhancers, colorants, flavorings, and sweeteners. The use of such media and agents for pharmaceutically active substances is well known in the art. Except insofar as any conventional media or agent is incompatible with the active ingredient, its use in the therapeutic compositions is contemplated.

[0038] As used herein, "administration" or "administer" or "administering" refers to dispensing, applying, or tendering one or more agents to a subject. Administration can be accomplished using any of several methods known in the art. For example, as used herein, "administration" means via injection (intravenous administration (iv)), parenteral administration. "Parenteral" means intravenous, subcutaneous, and intramuscular administration.

[0039] (Detailed explanation) One aspect provides a method for preventing or reducing drug-induced cardiotoxicity in a subject caused by a cardiotoxic agent, the method comprising administering to the subject an effective amount of bisantrene, a cardioprotective derivative of bisantrene, or a pharmaceutically acceptable salt of bisantrene or a cardioprotective derivative of bisantrene.

[0040] (cardiotoxicity) Cardiotoxicity is toxicity to cardiac cells, typically myocardial cells. Symptoms are usually chest pain and / or palpitations due to myopericarditis, sinus tachycardia, paroxysmal nonsustained supraventricular tachycardia, and premature atrial and ventricular beats. Electrocardiograms may reveal nonspecific ST-T changes, left axis deviation, and decreased amplitude of the QRS complex. Cardiotoxicity manifests in symptoms ranging in severity from overt clinical symptoms (e.g., heart failure) to asymptomatic detectable changes in cardiac function (e.g., electrophysiological dysfunction, contractile dysfunction, and cardiomyocyte apoptosis).

[0041] Typically, electrophysiological dysfunction includes QT prolongation and systolic dysfunction includes reduced cardiac ejection fraction (EF) or fractional shortening (FS).

[0042] Cardiotoxicity can be measured using several approaches.

[0043] One current measure of cardiotoxicity is the decrease in left ventricular ejection fraction (LVEF) volume relative to baseline. In this regard, the current standard definition of cardiotoxicity is defined by the Cardiac Review and Evaluation Committee (CREC) and ESMO Clinical Practice Guidelines for Trastuzumab-Related Cardiotoxicity, and is defined as a decrease in echocardiographic left ventricular ejection fraction (LVEF) of more than 10% to less than 50% from baseline. Methods for echocardiographic measurement of LVEF are known in the art and are described, for example, in Chatterjee, K., Zhang, J., Honbo, N. & Karliner, JS Doxorubicin Cardiomyopathy. Cardiology 115, 155-162 (2010).

[0044] Another measure of cardiotoxicity is an increase in the expression of cardiac marker proteins, such as cardiac troponin. Methods for measuring cardiac troponin levels are known in the art and are described, for example, in Dong, J. & Chen, H. Cardiotoxicity of Anticancer Therapeutics. Frontiers Cardiovasc Medicine 5, 9 (2018).

[0045] Elevated or abnormal expression levels of several biomarkers can be used as indicators for screening and assessing risk factors for future cardiotoxic complications. Interleukin-6 (IL-6), a cytokine produced by adipose tissue, increases blood pressure and induces inflammation. Overexpression of IL-6 inhibits cell apoptosis, stimulates angiogenesis, and plays a role in drug resistance (Guo Y, Xu F, Lu T, Duan Z, Zhang Z. Interleukin-6 signaling pathway in targeted therapy for Cancer. Cancer Treat Rev (2012) 38:904-10). There is also promising data exploring biomarkers for type I and type II cardiac events. Increased brain-type natriuretic peptide (BNP) and N-terminal pro-BNP have all been associated with reduced LVEF (Jain D, Russell RR, Schwartz RG, Panjrath GS, Aronow W. Cardiac complications of cancer therapy: pathophysiology, identification, prevention, treatment, and future direction. Curr Cardiol Rep (2017) 19:36). Plasma myeloperoxidase also predicts decline in cardiac function (Tromp, J., Steggink, L., Veldhuisen, DV, Gietema, J. & Meer, P. van der. Cardio-Oncology: Progress in Diagnosis and Treatment of Cardiac Dysfunction. Clin Pharmacol Ther 101, 481–490 (2017)). MicroRNAs have emerged as potential markers for the early onset of heart failure.miR-1, miR-133b, and miR-146a were all overexpressed after doxorubicin chemotherapy (Cappetta, D. et al. Doxorubicin targets multiple players: A new view of an old problem. Pharmacol Res 127, 4-14(2018)).

[0046] Typically, cardiotoxicity is induced or exacerbated by a cardiotoxic agent. Typically, cardiotoxicity is induced or exacerbated by treating a subject with a cardiotoxic substance, usually a therapeutic agent. Many chemotherapeutic agents, particularly anthracyclines, and many other therapeutic agents, such as carfilzomib, bortezomib, and trastuzumab, are cardiotoxic.

[0047] Thus, one aspect provides a method for preventing or reducing drug-induced cardiotoxicity induced or exacerbated by a cardiotoxic agent in a subject, the method comprising administering an effective amount of a cardioprotective agent comprising bisantrene or a derivative thereof, or a pharmaceutically acceptable salt of bisantrene or a derivative thereof.

[0048] (cardiotoxic agents) The cardioprotective properties of bisantrene, a cardioprotective derivative of bisantrene, or a pharmaceutically acceptable salt thereof make them useful as cardioprotective agents for subjects treated with cardiotoxic agents, particularly cardiotoxic chemotherapeutic agents, typically for the treatment of cancer.

[0049] In one embodiment, the cardiotoxic chemotherapeutic agent is an anthracycline or a pharmaceutically acceptable salt thereof. Examples of anthracyclines include daunorubicin, doxorubicin, epirubicin, idarubicin, mitoxantrone, and valrubicin.

[0050] In one embodiment, the anthracycline is doxorubicin.

[0051] In one embodiment, the anthracycline is daunorubicin.

[0052] In one embodiment, the anthracycline is epirubicin.

[0053] In one embodiment, the anthracycline is idarubicin.

[0054] In one embodiment, the anthracycline is mitoxantrone.

[0055] In one embodiment, the anthracycline is valrubicin.

[0056] In one embodiment, a method is provided for preventing or reducing cardiotoxicity induced or exacerbated by an anthracycline or a pharmaceutically acceptable salt thereof in a subject, the method comprising administering an effective amount of a cardioprotectant comprising bisantrene or a derivative thereof, or a pharmaceutically acceptable salt of bisantrene or a derivative thereof. The cardioprotectant can be administered before, simultaneously with, or after administration of the anthracycline or a pharmaceutically acceptable salt thereof.

[0057] In another embodiment, the cardiotoxic chemotherapeutic agent is a thymidine kinase inhibitor. Examples of cardiotoxic tyrosine kinase inhibitors include imatinib mesylate, dasatinib, nilotinib, sunitinib, sorafenib, and lapatinib.

[0058] In another embodiment, the cardiotoxic chemotherapeutic agent is an antibody-based chemotherapeutic agent. An example of a cardiotoxic antibody-based chemotherapeutic agent is trastuzumab (Herceptin®).

[0059] In another embodiment, the cardiotoxic chemotherapeutic agent is a proteasome inhibitor. All clinically approved proteasome inhibitors are associated with cardiotoxic effects. One specific example of a cardiotoxic proteasome inhibitor chemotherapeutic agent is carfilzomib (Kyprolis®), which has been reported to cause cardiotoxic effects in up to 10% of patients. Another example is bortezomib (Velcade®), for which there have been multiple case reports suggesting potential cardiotoxicity.

[0060] (Bisantrene) Bisantrene is a compound with direct cytotoxicity, as well as genomic and immunological mechanisms of action, including as an inhibitor (IC50 142nM) of fat mass and obesity-associated protein (FTO), an RNA N6-methyladenosine (m6A) demethylase (Su, R., Dong, L., Li, Y., Gao, M., Han, L., Wunderlich, M., et al. (2020). Targeting FTO Suppresses Cancer Stem Cell Maintenance and Immune Evasion. Cancer Cell, 38(1), 79-96.e11).

[0061] The chemical name of bisantrene is 9,10-anthracenedicarboxaldehyde bis[(4,5-dihydro-1H-imidazol-2-yl)hydrazone]. Typically, bisantrene is administered as a pharmaceutically acceptable salt. Typically, the pharmaceutically acceptable salt of bisantrene is bisantrene dihydrochloride.

[0062] The structure of bisantrene dihydrochloride is represented by formula (I): [ka] Shown below.

[0063] Bisantrene dihydrochloride is a tricyclic aromatic compound with the chemical name 9,10-anthracenedicarboxaldehyde bis[(4,5-dihydro-1H-imidazol-2-yl)hydrazone] dihydrochloride. The molecular formula of bisantrene hydrochloride is C22H22N8·2HCl and the molecular weight is 471.4 gmol. -1 The alkylimidazole side chain is highly basic and positively charged at physiological pH, which is thought to facilitate electrostatic attraction to the negatively charged ribose phosphate groups in DNA.

[0064] ] Bisantrene has a planar structure based on a resonant aromatic ring structure that intercalates within the helix of DNA and disrupts various functions, including replication, likely due to its potent inhibitory effect on the enzyme topoisomerase II.

[0065] Prior to this disclosure, bisantrene was known to exhibit anticancer activity. It was found to kill tumor cells in clonogenic assays, intercalate with DNA, and inhibit both DNA and RNA synthesis. Bisantrene's primary chemotherapeutic mechanism is its preferential binding to AT-rich regions, where it affects supercoiling and associates with DNA-associated proteins to initiate strand breaks. This is due to inhibition of the enzyme topoisomerase II, which relaxes DNA coiling during replication. It was found to be effective in cancer models using colon 26, Lewis lung, Ridgway osteosarcoma, B16, Lieberman plasma cell, P388, or L1210 cancer cells. Activity in clonogenic assays from 684 patients was observed in breast, small cell lung, large cell lung, squamous lung, ovarian, pancreatic, kidney, adrenal, head and neck, sarcoma, gastric, lymphoma, and melanoma tumor cells, but not in colorectal cancer.

[0066] Bisantrene possesses immunological properties that may be responsible for some of its anticancer activity. Following treatment with bisantrene, macrophages isolated from peritoneal exudates possessed cytostatic and antiproliferative functions in cultures of P815 (mastocytoma) tumor cells. Furthermore, supernatants from bisantrene-activated macrophages also possessed protective cytostatic effects in tumor cell cultures. Further studies revealed that macrophages activated with bisantrene and adoptively transferred into mice bearing EL-4 lymphoma more than doubled their median survival time, curing 7 out of 10 mice in the group. Multiple administrations of activated macrophages were more effective than a single administration. Recent studies have identified that bisantrene suppresses immune checkpoint gene expression and immune evasion through the enzymatic inhibition of FTO RNA demethylase, as described in R. Su. et al., “Targeting FTO Suppresses Cancer Stem Cell Maintenance and Immune Evasion”. Cancer Cell, 38(1), 79-96.e11 (2020).

[0067] Bisantrene has also been found to have non-immunological telomere-modifying effects. Bisantrene binds to DNA at sites known as G-quadruplexes, where four guanines are folded together. Stabilizing the G-quadruplexes disrupts telomere-telomerase interactions and can therefore inhibit telomerase activity in various ways, including by displacing telomerase-binding proteins. Because the level of topoisomerase II inhibition does not always correlate with cytotoxicity, alternative mechanisms may play a role in the action of bisantrene.

[0068] Bisantrene also has other mechanisms that may contribute to its anticancer activity, including immune enhancement. These mechanisms are described, for example, in the following: (i) N.R. West et al., "Tumor-Infiltrating Lymphocytes Predict Response to Anthracycline-Based Chemotherapy in Estrogen-Resistant Breast Cancer," Breast Canc. Res. 13: R126 (2011), concluded that the level of tumor-infiltrating lymphocytes correlates with the response to anthracycline drug administration, and markers associated with tumor-infiltrating lymphocytes (TILs) include CD19, CD3D, CD48, GZMB, LCK, MS4A1, PRF1, and SELL. (ii) L. Zitvogel et al., “Immunological Aspects of Cancer Chemotherapy,” Nature Rev. Immunol. 8: 59-73 (2008) states that DNA damage, such as that caused by intercalating agents such as bisantrene, induces the expression of NKG2D ligands on tumor cells in an ATM-dependent and CHK1-dependent (but p53-independent) manner; NKG2D is an activating receptor involved in tumor immune surveillance by NK cells, NKT cells, γδT cells, and resting (in mice) and / or activated (in humans) CD8 T cells; and that anthracyclines, particularly in combination with IL-12, can act as immunostimulants, and that such agents also promote HMGB1 release and activate T cells. (iii) DV Krysko et al., “TLR2 and TLR9 Are Sensors of Apoptosis in a Mouse Model of Doxorubicin-Induced Acute Inflammation,” Cell Death Different. 18: 1316-1325 (2011) states that anthracycline antibiotics induce an immunogenic form of apoptosis with immunostimulatory properties mediated by MyD88, TLR2, and TLR9.(iv) C. Ferraro et al., “Anthracyclines Trigger Apoptosis of Both G0-G1 and Cycling Peripheral Blood Lymphocytes and Induce Massive Deletion of Mature T and B Cells,” Cancer Res. 60: 1901-1907 (2000) reported that anthracyclines induce apoptosis and ceramide production, and activate caspase-3 in resting and cycling cells, and that the induced apoptosis is independent of CD95-L / CD95 and TNF / TNF-R. (v) K. Lee et al., "Anthracycline Chemotherapy Inhibits HIF-1 Transcriptional Activity and Tumor-Induced Mobilization of Circulating Angiogenic Cells," Proc. Natl. Acad. Sci. USA 106: 2353-2358 (2009), provides another antitumor mechanism of anthracycline antibiotics: inhibition of HIF-1-mediated gene transcription, which in turn inhibits the transcription of VEGF, which is required for angiogenesis; HIF-1 also activates the transcription of genes encoding the glucose transporter GLUT1 and hexokinases HK1 and HK2, which are required for the high levels of glucose uptake and phosphorylation observed in metastatic cancer cells, and pyruvate dehydrogenase kinase 1 (PDK1), which shunts pyruvate from mitochondria, thereby increasing lactate production. It was suggested that patients with HIF-1α overexpression based on immunohistochemistry results are good candidates for treatment with anthracycline antibiotics. (iv) R. Su et al., “Targeting FTO Suppresses Cancer Stem Cell Maintenance and Immune Evasion.” Cancer Cell, 38(1), 79-96.e11 (2020) identified a new mechanism of action of bisantrene via inhibition of the FTO enzyme.Inhibition of FTO was found to enhance immunotherapy responses through suppression of immune checkpoint gene expression.

[0069] (Bisantrene derivative) Derivatives of bisantrene are described in U.S. Patent Application Publication No. 2016 / 0166546 by Garner et al. Derivatives of bisantrene include, for example, derivatives of bisantrene having the following structure: [ka] [ka] [ka] [ka] [ka] The compounds include:

[0070] Further examples of derivatives of bisantrene include those having the following structures: [ka] The compounds include: wherein R1 and R3 are the same or different and are selected from hydrogen, C1-C6 alkyl, -C(O)-R5, where R5 is hydrogen, C1-C6 alkyl, phenyl, monosubstituted phenyl (wherein the substituents can be ortho, meta, or para and are fluoro, nitro, C1-C6 alkyl, C1-C3 alkoxy, or cyano), pentafluorophenyl, naphthyl, furanyl, [ka] -SO3H and wherein only one of R1 and R3 can be hydrogen or C1-C6 alkyl, and R2 and R4 are the same or different and are selected from hydrogen, C1-C4 alkyl, or —C(O)—R6; wherein R6 is hydrogen, C1-C6 alkyl, phenyl, monosubstituted phenyl (wherein the substituents can be in the ortho, meta, or para positions and are fluoro, nitro, C1-C6 alkyl, C1-C3 alkoxy, or cyano), pentafluorophenyl, naphthyl, furanyl, or -CHOCH3; The compounds can have the general structure B(Q)n, where B is a residue formed by removing a hydrogen atom from one or more basic nitrogen atoms of an amine-, amidine-, guanidine-, isourea-, isothiourea-, or biguanide-containing pharmaceutically active compound, Q is hydrogen or A, and A is [ka] and Thus, R' and R" are the same or different and are R, where R is C1-C6 alkyl, aryl, arylalkyl, heteroalkyl, NC-CH2CH2-, [ka] Cl3C-C-, or R7OCH2CH2-, where R7 is hydrogen or C1-C6 alkyl, hydrogen, or a pharmaceutically acceptable cation, or R' and R" are linked to form a -CH2CH2- group or [ka] group, and n is an integer representing the number of primary or secondary basic nitrogen atoms in the compound such that at least one Q is A. Further examples of derivatives of bisantrene include the following compounds: 9,10-bis[(2-hydroxyethyl)iminomethyl]anthracene; 9,10-bis{[2-(-2-hydroxyethylamino)ethyl]iminomethyl}anthracene; 9,10-bis{[2-(-2-hydroxyethylamino)ethyl]iminomethyl}anthracene; 9,10-bis{[2-(morpholin-4-yl)ethyl]iminomethyl}anthracene; 9,10-bis[(2-hydroxyethyl)aminomethyl]anthracene; 9,10-bis{[2-(2-hydroxyethylamino)ethyl]aminomethyl}anthracene tetrahydrochloride; 9,10-bis{[2-(piperazin-1-yl)ethyl]aminomethyl}anthracene hexahydrochloride; 9,10-bis{[2-(morpholin-4-yl)ethyl]aminomethyl}anthracene tetrahydrochloride; N,N'-bis[2-(dimethylamino)ethyl]-9,10-anthracene-bis(methylamine); and N,N'-bis(1-ethyl-3-piperidinyl)-9,10-anthracene-bis(methylamine) Includes:

[0071] Derivatives of bisantrene that are cardioprotective can be readily determined using the methods described herein.

[0072] (Treatment method) In one aspect, the present invention provides a method for preventing or reducing drug-induced cardiotoxicity in a subject, the method comprising administering to the subject an effective amount of a cardioprotectant comprising bisantrene or a derivative thereof, or a pharmaceutically acceptable salt of bisantrene or a derivative thereof.

[0073] Drug-induced cardiotoxicity can be caused by any cardiotoxic agent to which a subject is exposed or which is endogenously produced, but typically cardiotoxicity is caused by exposure to or administration to a subject of a cardiotoxic therapeutic agent, particularly a cardiotoxic chemotherapeutic agent such as an anthracycline, a proteasome inhibitor, or a tyrosine kinase inhibitor.

[0074] Thus, in certain aspects, the present invention provides a method for preventing or reducing cardiotoxicity in a subject being treated with a cardiotoxic chemotherapeutic agent, the method comprising administering to the subject an effective amount of a cardioprotective agent comprising bisantrene or a derivative thereof, or a pharmaceutically acceptable salt of bisantrene or a derivative thereof.

[0075] In an even more particular embodiment, the present invention provides a method of treating cancer in a subject, the method comprising: a. a cardiotoxic chemotherapeutic agent in an amount therapeutically effective against cancer, and b. A cardioprotective agent comprising a cardioprotective amount of bisantrene or a derivative thereof, or a pharmaceutically acceptable salt of bisantrene or a derivative thereof. The method includes administering

[0076] The cancer may be any cancer that responds to treatment with a cardiotoxic chemotherapy agent. For example, anthracyclines have been shown to be effective in treating a wide range of cancers, including breast cancer, leukemia, lymphoma, gastric cancer, uterine cancer, ovarian cancer, bladder cancer, lung cancer, melanoma, and myeloma. In one embodiment, the cancer is selected from the group consisting of bladder cancer, bone cancer, brain cancer, breast cancer, esophageal cancer, colorectal cancer, leukemia, liver cancer, lung cancer, lymphoma, myeloma, ovarian cancer, prostate cancer, sarcoma, gastric cancer, and thyroid cancer. According to one embodiment, the cancer is renal clear cell carcinoma. According to another embodiment, the cancer is melanoma. According to another embodiment, the cancer is multiple myeloma. According to another embodiment, the cancer is acute myeloid leukemia.

[0077] Pharmaceutical compositions and medicaments of the present invention may be administered to a subject by standard enteral or parenteral routes (including, but not limited to, injection (intravenous, subcutaneous, intramuscular, bolus, etc.)), or by, for example, topical, oral, sublingual, nasal, pulmonary, otic, rectal, or vaginal routes of administration. In some embodiments, pharmaceutical compositions according to the present invention may be administered to a subject alone or in combination with other pharmaceutical compositions. When combined with other pharmaceutical compositions, administration may be simultaneous or sequential, or the administration of the pharmaceutical compositions may be independent of one another.

[0078] In certain embodiments, bisantrene is administered intravenously, either centrally or peripherally, including intramuscularly, subcutaneously, and / or intradermally.

[0079] Generally, the pharmaceutical compositions and medicaments of the present invention can be administered in a manner compatible with the route of administration and the physical characteristics (including health status) of the subject, and so as to induce the desired effect (i.e., therapeutically effective and / or prophylactic). For example, the appropriate dosage may depend on various factors, including, but not limited to, the physical characteristics of the subject (e.g., age, weight, sex), whether the composition or medicament is used as a single agent, and the progression of the disease or disorder being treated (i.e., pathological state), as well as other factors readily apparent to those skilled in the art.

[0080] The appropriate dosage, frequency, duration, and route of administration of a chemotherapeutic agent are known in the art. Bisantrene, a cardioprotective derivative of bisantrene, or a pharmaceutically acceptable salt of bisantrene or a cardioprotective derivative of bisantrene can be administered in the same pharmaceutical composition as the cardiotoxic chemotherapeutic agent, in a separate composition from the cardiotoxic chemotherapeutic agent, but at the same time as the cardiotoxic chemotherapeutic agent, or at a different time. When bisantrene, a cardioprotective derivative of bisantrene, or a pharmaceutically acceptable salt of bisantrene or a cardioprotective derivative of bisantrene is administered at a different time from the cardiotoxic chemotherapeutic agent, it can be administered either before or after the cardiotoxic chemotherapeutic agent and / or at a different timing, and according to a different timing and / or frequency regimen. Those skilled in the art can determine an appropriate administration schedule based on variables such as the subject's age, weight, and sex, the subject's susceptibility to cardiotoxicity, genetic markers, the dose of the cardiotoxic agent, the subject's medical history of previous cardiotoxic agents, and pharmacokinetic parameters such as cardiac function.

[0081] The methods and compositions provided herein allow subjects to receive treatment more frequently without having to significantly alter their dosing regimen due to the risk of cardiotoxicity.A daily dose of a cardiotoxic chemotherapy agent and bisantrene, a cardioprotective derivative of bisantrene, or a pharmaceutically acceptable salt of bisantrene or a cardioprotective derivative of bisantrene can be administered to a subject in one or more doses per day.In some cases, the daily dose of a chemotherapy agent can be administered in a single dose together with bisantrene, a cardioprotective derivative of bisantrene, a pharmaceutically acceptable salt of bisantrene or a cardioprotective derivative of bisantrene.

[0082] The pharmaceutical compositions described herein can be administered to a patient one or more times a day. In some cases, the pharmaceutical composition can be administered to a patient once a day. In some cases, the pharmaceutical composition can be administered to a patient at least two, three, four, five, or six times a day. For example, the pharmaceutical composition can be administered to a patient three times a day.

[0083] In the methods described herein, the appropriate dosage of bisantrene (or a cardioprotective derivative of bisantrene) can be determined by one skilled in the art. The selected dosage level will depend on various pharmacokinetic factors, including the amount of cardiotoxic chemotherapeutic agent administered, the route of administration, the time of administration, the rate of excretion of the particular compound used, the severity of the condition, other health considerations affecting the subject, and the state of the subject's liver and kidney function. The selected dosage level will also depend on the duration of treatment, other drugs, compounds, and / or materials used in combination with the particular therapeutic agent used, as well as the age, weight, condition, general health, and previous medical history of the subject being treated, and similar factors. Methods for determining optimal dosages can be found in the art, for example, in "Remington: The Science and Practice of Pharmacy," Mack Publishing Co., 2012. thed., 2000, and Gilman et al., (Eds), (1990), "Goodman and Gilman's: The Pharmacological Bases of Therapeutics", Pergamon Press. Optimal dosages for a given set of conditions can be ascertained by one skilled in the art using conventional dosage-determining tests in view of the experimental data for the agent.

[0084] According to certain embodiments, the administration of bisantrene is about 0.1 mg / m 2 / day~about 100mg / m 2 / day, e.g., about 0.2 mg / m 2 / day~about 50mg / m 2 / day, about 0.5mg / m 2 / day~about 20mg / m 2 / day, approximately 1.0 mg / m 2 / day~about 10mg / m 2 / day, approximately 1.0 mg / m 2 / day~about 8mg / m 2 / day, approximately 1mg / m 2 / day, about 2mg / m 2 / day, about 3mg / m 2 / day, approximately 4mg / m 2 / day, about 5mg / m 2 / day, approximately 6mg / m 2 / day, approximately 7mg / m 2 / day, or approximately 10 mg / m 2 In some embodiments, bisantrene is administered daily or weekly, once every two weeks, once every three weeks, or once every four weeks for a period of time, e.g., 1 day, 2 days, 3 days, 4 days, 5 days, 6 days, 7 days, 8 days, 9 days, 10 days, 11 days, 12 days, 13 days, 14 days, 21 days, 28 days, 35 days, 42 days, 49 days, 56 days, or 63 days. In certain embodiments, bisantrene is administered at a dose of, for example, about 20-100 mg / m 2 / 28 day dosage, one or more times over a 28 day period, optionally daily or once or more times weekly, once every two weeks, once every three weeks, or once every four weeks. For example, in certain embodiments, bisantrene is administered at a dose of about 0.5 mg / m once weekly for four weeks. 2 / week~about 50mg / m 2 / week, about 1mg / m 2 / week~about 40mg / m 2 / week, about 2mg / m 2 / week~about 30mg / m 2 / week, about 5mg / m 2 / week~about 25mg / m 2 / week, about 5mg / m 2 / week, about 10mg / m 2 / week, about 15mg / m 2 / week, about 20mg / m 2 / week, or approximately 25 mg / m 2 The patient may be administered a 100mg / week dosing regimen. The administration of a pharmaceutically acceptable salt of bisantrene or a cardioprotective derivative thereof or a pharmaceutically acceptable salt thereof may be at a similar dosing rate adjusted for molar equivalents.

[0085] According to certain embodiments, the biological activity observed when a cardioprotectant is administered with a cardiotoxic agent may provide at least an additive, if not a synergistic, result, thereby enabling a more effective therapeutic outcome for a given administration rate of the cardiotoxic agent. This biological activity of the cardioprotectant may also enable the use or extended use of cardiotoxic therapeutic agents, such as proteasome inhibitors like carfilzomib or bortezomib, which cannot be safely administered to patients or whose administration to patients is limited due to their cardiotoxicity.

[0086] Alternatively, when administered in combination with a cardioprotectant, a cardiotoxic agent can be administered to a patient at lower doses and for longer periods than would normally be administered while maintaining comparable ongoing treatment outcomes. This is particularly important for cardiotoxic agents, such as many anthracyclines, which have lifetime cumulative dose limits. Thus, in certain embodiments of the methods of the present invention, when administered in combination with a cardioprotectant, the cardiotoxic agent can be administered at a rate significantly lower than the recommended administration rate for that cardiotoxic agent when administered alone, e.g., at least 10%, 20%, 30%, 40%, 50%, 60%, or 70% lower than the administration rate for that cardiotoxic agent when administered alone. For example, a typical administration rate for doxorubicin is about 50-75 mg / m 2 / 28 days, but when co-administered with bisantrene to a patient undergoing cancer treatment by the method according to the present invention, doxorubicin is administered at about 10 mg / m 2 / 28 days ~ approx. 60mg / m 2 / 28 days, e.g., about 15 mg / m 2 / 28 days ~ approx. 50mg / m 2 / 28 days, about 20mg / m 2 / 28 days ~ approx. 50mg / m 2 / 28 days, or approximately 25 mg / m 2 / 28 days ~ approx. 45mg / m 2 / 28 days, and bisantrene is administered at a similar rate, e.g., about 30-40 mg / m 2 Bisantrene at a dose rate of approximately 30-40 mg / m² / 28 days 2 Co-administration with an anthracycline such as doxorubicin, daunorubicin, or epirubicin at a dosing rate of 1 / 28 days is contemplated for cancer treatment methods according to the present invention.

[0087] In certain embodiments of the treatment methods according to the present invention, the doses of the cardiotoxic agent and the cardioprotective agent can be in a molar ratio of about 1:10 to about 10:1, e.g., about 1:5 to about 5:1, about 1:4 to 4:1, about 1:3 to 3:1, about 1:2 to 2:1, about 1:1.5 to 1.5:1, or about 1:1. In alternative embodiments, the dose of the cardiotoxic agent and the dose of the cardioprotective agent are independent. For example, if the cardioprotective agent is bisantrene, it can be administered to the patient at the rates discussed above, regardless of the dosing regimen of the cardiotoxic agent, and thus, for example, about 0.5 mg / m once weekly for 4 weeks, regardless of the timing or rate of administration of the cardioprotective agent. 2 / week~about 50mg / m 2 / week, about 1mg / m 2 / week~about 40mg / m 2 / week, about 2mg / m 2 / week~about 30mg / m 2 / week, about 5mg / m 2 / week~about 25mg / m 2 / week, about 5mg / m 2 / week, about 10mg / m 2 / week, about 15mg / m 2 / week, about 20mg / m 2 / week, or approximately 25 mg / m 2 The patient may be administered a 100 mg / week dosing regimen.

[0088] (composition) Cardioprotective agents are typically administered in pharmaceutical compositions.

[0089] The cardioprotective agent may be administered to a subject separately from any other therapeutic agent or in combination with a cardiotoxic agent, such as a cardiotoxic chemotherapeutic agent. Thus, according to one embodiment, a pharmaceutical composition according to the present invention may comprise a cardioprotective agent comprising bisantrene or a derivative thereof, or a pharmaceutically acceptable salt of bisantrene or a derivative thereof. According to another embodiment of the present invention, the composition may also comprise a cardiotoxic agent, such as a cardiotoxic chemotherapeutic agent.

[0090] In one embodiment, the cardiotoxic chemotherapeutic agent is an anthracycline or a pharmaceutically acceptable salt thereof.

[0091] In one embodiment, the anthracycline is selected from daunorubicin, doxorubicin, epirubicin, idarubicin, mitoxantrone, and valrubicin.

[0092] In another embodiment, the cardiotoxic chemotherapeutic agent is a proteasome inhibitor such as carfilzomib or bortezomib.

[0093] In other embodiments, the cardiotoxic chemotherapeutic agent is a tyrosine kinase inhibitor.

[0094] In other embodiments, the cardiotoxic chemotherapeutic agent is a monoclonal antibody, such as trastuzumab.

[0095] The compositions according to the invention can be administered by any route in a form suitable for that route, as is known in the art. Thus, the compositions according to the invention can be adapted for administration by enteral or parenteral routes, including injection (intravenous, subcutaneous, intramuscular, bolus, etc.), or by, for example, topical, oral, sublingual, nasal, pulmonary, aural, rectal or vaginal routes of administration.

[0096] Typically, the pharmaceutical compositions described herein contain at least one pharmaceutically acceptable carrier or excipient and / or diluent. For preparing pharmaceutical compositions and medicaments, inert pharmaceutically acceptable carriers may be solid or liquid. Liquid form preparations include solutions, suspensions, and emulsions, such as water or water-propylene glycol solutions for parenteral injection. Solid form preparations, such as tablets, or amorphous or crystalline powders, including lyophilized preparations, are also included, which are intended to be converted immediately before use into liquid form preparations for either oral or injectable administration. Such liquid forms include solutions, suspensions, and emulsions. Examples of pharmaceutically acceptable carriers and manufacturing methods for various compositions can be found, for example, in "Remington: The Science and Practice of Pharmacy," Mack Publishing Co., 2009. thed., 2000, and Gilman et al., (Eds), (1990), "Goodman and Gilman's: The Pharmacological Bases of Therapeutics", Pergamon Press.

[0097] Pharmaceutically acceptable carriers and excipients include: (i) a liquid carrier; (ii) isotonic agents; (iii) a wetting or emulsifying agent; (iv) preservatives; (v) buffering agents; (vi) acidifying agent; (vii) antioxidants; (viii) alkalizing agents; (ix) a carrier; (x) chelating agents, (xi) coloring agents; (xii) a complexing agent; (xiii) a solvent, (xiv) suspending and / or thickening agents; (xv) oil; (xvi) penetration enhancers; (xvii) polymers, (xviii) a curing agent; (xix) proteins, (xx) carbohydrates, (xxi) bulking agents, and (xxii) Lubricants Includes:

[0098] Other pharmaceutically acceptable carriers and excipients known in the art may also be used.

[0099] Carriers, diluents, excipients and adjuvants must be "acceptable" in the sense of being compatible with the other ingredients of the composition or medicament, and generally not harmful to the subject. Non-limiting examples of pharmaceutically acceptable carriers or diluents include demineralized or distilled water; saline; vegetable oils such as peanut oil, safflower oil, olive oil, cottonseed oil, corn oil, etc.; peanut oil, safflower oil, olive oil, cottonseed oil, corn oil, sesame oil such as sesame oil, peanut oil or coconut oil; silicone oils including polysiloxanes such as methylpolysiloxane, phenylpolysiloxane and methylphenylpolysorboxane; volatile silicones; mineral oils such as liquid paraffin, soft paraffin or squalane; methylcellulose, ethylcellulose, carboxymethyl ... Examples of suitable carriers include cellulose derivatives such as sodium ethyl cellulose or hydroxypropylmethylcellulose; lower alkanols such as ethanol or isopropanol; lower aralkanols; lower polyalkylene glycols or lower alkylene glycols such as polyethylene glycol, polypropylene glycol, ethylene glycol, propylene glycol, 1,3-butylene glycol, or glycerin; fatty acid esters such as isopropyl palmitate, isopropyl myristate, or ethyl oleate; polyvinylpyrrolidone; agar; gum tragacanth or gum acacia, and petrolatum. Typically, the carrier forms about 10% to about 99.9% by weight of the composition, vaccine, or medicament.

[0100] For administration as an injectable solution or suspension, non-toxic parenterally acceptable diluents or carriers may include Ringer's solution, isotonic saline, phosphate buffered saline, ethanol, and 1,2 propylene glycol. Methods for preparing parenterally administrable pharmaceutical compositions and medicaments are apparent to those skilled in the art, and are described, for example, in "Remington: The Science and Practice of Pharmacy," Mack Publishing Co., 2002. thed., 2000, and Gilman et al., (Eds), (1990), "Goodman and Gilman's: The Pharmacological Bases of Therapeutics", Pergamon Press.

[0101] For oral administration, some examples of suitable carriers, diluents, excipients and adjuvants include peanut oil, liquid paraffin, sodium carboxymethylcellulose, methylcellulose, sodium alginate, gum acacia, gum tragacanth, dextrose, sucrose, sorbitol, mannitol, gelatin and lecithin.In addition, these oral preparations can contain suitable flavoring and coloring agents.When used in capsule form, capsules can be coated with compounds such as glyceryl monostearate or glyceryl stearate to delay disintegration.

[0102] Solid forms for oral administration may contain binders, sweeteners, disintegrants, diluents, flavorings, coatings, preservatives, lubricants, and / or time-delay agents acceptable in human and veterinary medicine. Suitable binders include gum acacia, gelatin, corn starch, tragacanth, sodium alginate, carboxymethylcellulose, or polyethylene glycol. Suitable sweeteners include sucrose, lactose, glucose, aspartame, or saccharin. Suitable disintegrants include corn starch, methylcellulose, polyvinylpyrrolidone, guar gum, xanthan gum, bentonite, alginic acid, or agar. Suitable diluents include lactose, sorbitol, mannitol, dextrose, kaolin, cellulose, calcium carbonate, calcium silicate, or dicalcium phosphate. Suitable flavoring agents include peppermint oil, oil of wintergreen, cherry, orange, or raspberry flavoring. Suitable coating agents include polymers or copolymers of acrylic acid and / or methacrylic acid and / or their esters, waxes, fatty alcohols, zein, shellac, or gluten. Suitable preservatives include sodium benzoate, vitamin E, α-tocopherol, ascorbic acid, methylparaben, propylparaben, or sodium bisulfite. Suitable lubricants include magnesium stearate, stearic acid, sodium oleate, sodium chloride, or talc. Suitable time delay agents include glyceryl monostearate or glyceryl distearate.

[0103] Liquid forms for oral administration may contain, in addition to the above-mentioned agents, a liquid carrier. Suitable liquid carriers include water, oils such as olive oil, peanut oil, sesame oil, sunflower oil, safflower oil, peanut oil, coconut oil, liquid paraffin, ethylene glycol, propylene glycol, polyethylene glycol, ethanol, propanol, isopropanol, glycerol, fatty alcohols, triglycerides or mixtures thereof.

[0104] Suspensions for oral administration may further contain a dispersing agent and / or a suspending agent. Suitable suspending agents include sodium carboxymethylcellulose, methylcellulose, hydroxypropylmethylcellulose, polyvinylpyrrolidone, sodium alginate, or acetyl alcohol. Suitable dispersing agents include lecithin, polyoxyethylene esters of fatty acids, such as stearic acid, polyoxyethylene sorbitol mono- or di-oleate, -stearate, or -laurate, polyoxyethylene sorbitan mono- or di-oleate, -stearate, or -laurate, etc.

[0105] In certain embodiments, the pharmaceutical composition may contain liposomes. A suitable liposome formulation for bisantrene or its cardioprotective derivatives comprises small unilamellar or multilamellar liposomes ranging in size from 0.01 to 100 μM and approximately 50 to 95% of the total liposomes, composed of hydrogenated soybean phosphatidylcholine, distearoylphosphatidylglycerol, and cholesterol from lipids of natural or synthetic origin, in an aqueous solution that can be reconstituted from lyophilized form into an injectable liposomal suspension. The composition is prepared by reconstituting the lyophilized bisantrene / liposome composition into a liposome concentrate and then diluting the concentrate for parenteral administration for the treatment of melanoma.

[0106] In yet another embodiment, the pharmaceutical composition may include a complex with β-cyclodextrin. Suitable liposome formulations for bisantrene, a cardioprotective derivative of bisantrene, or a pharmaceutically acceptable salt or derivative of bisantrene include a complex formed in aqueous solution that can be reconstituted from a lyophilized form into an injectable suspension. One such composition is prepared by reconstituting a lyophilized bisantrene / β-cyclodextrin composition into a concentrate and then diluting the concentrate for parenteral administration. β-cyclodextrin complexes and methods for preparing such complexes are known in the art and are described, for example, in WO 2019 / 073296 (Rothman).

[0107] Various formulations suitable for use in administering bisantrene, cardioprotective derivatives of bisantrene, or pharmaceutically acceptable salts of bisantrene or cardioprotective derivatives of bisantrene are known in the art. U.S. Patent No. 4,784,845 (Desai et al.) discloses a composition for delivering a hydrophobic drug (i.e., bisantrene or its cardioprotective derivative), which comprises (i) the hydrophobic drug, (ii) an oily vehicle or oil phase substantially free of butylated hydroxyanisole (BHA) or butylated hydroxytoluene (BHT), (iii) a co-surfactant or emulsifier, (iv) a co-surfactant or co-emulsifier, and (v) benzyl alcohol as a co-solvent. U.S. Pat. No. 4,816,247 (Desai et al.) discloses a composition for delivering a hydrophobic drug (e.g., bisantrene or its cardioprotective derivatives or analogs) via intravenous, intramuscular, or intraarticular routes, the composition comprising: (i) the hydrophobic drug; (ii) a pharmaceutically acceptable oily vehicle or oil selected from the group consisting of (a) naturally occurring vegetable oils and (b) semi-synthetic mono-, di-, and triglycerides, wherein the oily vehicle or oil does not contain BHT or BHA; (iii) a surfactant or emulsifier; (iv) a co-surfactant or emulsifier; (v) an ion-pairing agent selected from C6-C20 saturated or unsaturated aliphatic acids when the hydrophobic drug is basic, and a pharmaceutically acceptable aromatic amine when the hydrophobic drug is acidic; and (vi) water.

[0108] According to a particular embodiment, the composition according to the invention has a concentration of about 0.1 mg / m 2 / day~about 100mg / m 2 / day, e.g., about 0.2 mg / m 2 / day~about 50mg / m 2 / day, about 0.5mg / m 2 / day~about 20mg / m 2 / day, approximately 1.0 mg / m 2 / day~about 10mg / m 2 / day, approximately 1.0 mg / m 2 / day~about 8mg / m 2 / day, approximately 1mg / m 2 / day, about 2mg / m 2 / day, about 3mg / m 2 / day, approximately 4mg / m 2 / day, about 5mg / m 2 / day, approximately 6mg / m 2 / day, approximately 7mg / m 2 / day, or approximately 10 mg / m 2 In some embodiments, the compositions according to the invention can be adapted for administration of bisantrene at a dosage of about 20-50 mg / m², optionally once or more times daily or weekly, once every two weeks, once every three weeks, or once every four weeks, over a period of time, e.g., 1 day, 2 days, 3 days, 4 days, 5 days, 6 days, 7 days, 8 days, 9 days, 10 days, 11 days, 12 days, 13 days, 14 days, 21 days, 28 days, 35 days, 42 days, 49 days, 56 days, or 63 days. In certain embodiments, the compositions according to the invention can be adapted for administration of bisantrene at a dosage of about 20-50 mg / m², optionally once or more times daily or weekly, once every two weeks, once every three weeks, or once every four weeks, over a period of 28 days. 2 The compositions according to the present invention comprising a pharmaceutically acceptable salt of bisantrene or a cardioprotective derivative thereof or a pharmaceutically acceptable salt thereof can be adapted to deliver similar administration rates adjusted for molar equivalents.

[0109] In certain embodiments, compositions according to the present invention include both a cardioprotective agent and a cardiotoxic therapeutic agent, and the composition is adapted to deliver the cardioprotective agent at the dosages described above and the cardiotoxic therapeutic agent at a desired administration rate according to the recommended advice of the cardioprotective agent or according to the specific protocol designed for the treatment regimen being adopted.

[0110] In certain embodiments of the compositions according to the invention, the dose of the cardiotoxic agent and the cardioprotective agent can be in a molar ratio of about 1:10 to about 10:1, e.g., about 1:5 to about 5:1, about 1:4 to 4:1, about 1:3 to 3:1, about 1:2 to 2:1, about 1:1.5 to 1.5:1, or about 1:1. When the cardioprotective agent is bisantrene and the cardiotoxic agent is an anthracycline, the compositions according to the invention can be, for example, 20 to 60 mg / m 2 / 28-day dose delivers bisantrene, approximately 20-60 mg / m 2 Carfilzomib, on the other hand, has a significantly higher specific activity than doxorubicin and must be present in the composition at a significantly lower ratio than bisantrene or other cardioprotective agents.

[0111] (kit) The present invention also provides kits for reducing, preventing, or eliminating drug-induced cardiotoxicity in a subject.

[0112] One aspect provides a kit for preventing or reducing cardiotoxicity in a subject, the kit comprising bisantrene, a cardioprotective derivative of a cardioprotectant comprising bisantrene or a derivative thereof, or a pharmaceutically acceptable salt of bisantrene or a derivative thereof.

[0113] In one embodiment, the kit further comprises a cardiotoxic agent.

[0114] In one embodiment, the cardiotoxic agent is a cardiotoxic chemotherapeutic agent.

[0115] In one embodiment, the cardiotoxic chemotherapeutic agent is an anthracycline or a pharmaceutically acceptable salt thereof.

[0116] In one embodiment, the anthracycline is selected from daunorubicin, doxorubicin, epirubicin, idarubicin, mitoxantrone, and valrubicin.

[0117] In one embodiment, the anthracycline is doxorubicin, daunorubicin, or epirubicin.

[0118] In another embodiment, the cardiotoxic chemotherapeutic agent is a tyrosine kinase inhibitor.

[0119] In another embodiment, the cardiotoxic chemotherapeutic agent is a monoclonal antibody, such as, for example, trastuzumab.

[0120] In another embodiment, the cardiotoxic chemotherapeutic agent is a proteasome inhibitor, such as, for example, carfilzomib or bortezomib.

[0121] In some cases, the kit may also include vials, tubes, needles, packaging, or other materials.

[0122] Kits containing one or more unit doses of the compounds described herein are provided, usually in injectable doses. Such kits may include a container containing the unit dose, an information package insert describing the use and associated benefits of the drug in treating disease, and optionally an instrument or device for delivering the composition.

[0123] The kit may further comprise any device suitable for administering the composition, for example, the kit may comprise a needle suitable for intravenous administration.

[0124] In some cases, the kit may be provided with instructions. The instructions may be provided in the kit or may be accessed electronically. The instructions may provide information on how to use the compositions of the present disclosure. The instructions may further provide information on how to use the devices of the present disclosure. The instructions may provide information on how to practice the methods of the present disclosure. In some cases, the instructions may provide dosing information. The instructions may provide drug information such as mechanism of action, drug formulation, adverse risks, contraindications, etc. In some cases, the kit is purchased by a physician or healthcare provider for administration in a clinic or hospital. In some cases, the kit is purchased by a laboratory and used to screen candidate compounds.

[0125] Of course, any material used in preparing the pharmaceutical compositions described herein should ideally be pharmaceutically pure and substantially non-toxic in the amounts used.

[0126] In the following claims and in the foregoing description of the invention, unless the context requires otherwise by clear words or necessary implication, the word "comprise" or variations such as "comprises" or "comprising" are used in the inclusive sense, i.e., to specify the presence of stated features but not to exclude the presence or addition of further features in various embodiments of the invention.

[0127] All publications mentioned herein are incorporated herein by reference.

[0128] Those skilled in the art will appreciate that numerous variations and / or modifications may be made to the invention as shown in the specific embodiments without departing from the spirit or scope of the invention as broadly described, and the present embodiments are, therefore, to be considered in all respects as illustrative and not restrictive.

[0129] The following non-limiting examples are provided to illustrate so that the nature of the present invention may be more clearly understood. [Example]

[0130] Example 1 - Materials and Methods (Primary human cardiomyocyte culture) Primary human cardiomyocytes (HCMs) were purchased from PromoCell® (Heidelberg, Germany) and cultured according to the manufacturer's recommendations using PromoCell® Myocyte Growth Medium (5% [v / v] fetal bovine serum, 0.5 ng / ml recombinant human epidermal growth factor, 2 ng / ml recombinant human basic fibroblast growth factor, 5 μg / ml recombinant human insulin) supplemented with 1% [v / v] penicillin-streptomycin [Sigma-Aldrich]. Briefly, 1 × 10 6HCMs were seeded and expanded in Corning® T75 flasks (Sigma-Aldrich, New South Wales, Australia) in a 37°C incubator under 5% CO2 conditions. At 80-90% confluency, HCMs were gently detached using a DetachKit (PromoCell®) and collected in Corning® 50 mL centrifuge tubes (Sigma-Aldrich) and centrifuged at 300 x g for 10 minutes at room temperature. The resulting supernatant was discarded, and the HCM pellet was resuspended in 5-10 mL of prewarmed myocyte growth medium. Total viable cells were counted using an Invitrogen™ Countess Cell Counter (Thermo Fisher Scientific, New South Wales, Australia) and collected at 1 x 10 cells / ml. 5 Live HCMs were seeded in triplicate onto Corning® 96-well white polystyrene microplates (Sigma). The seeded HCMs were incubated overnight at 37°C in a 5% CO2 incubator before treatment with bisantrene (day 1).

[0131] (MCF7 breast cancer cells) Human breast adenocarcinoma (MCF7) cells were purchased from In Vitro Technologies (Victoria, Australia) and cultured according to the manufacturer's recommendations using MCF7 growth medium, low-glucose Dulbecco's modified Eagle's medium (DMEM; Sigma-Aldrich, Australia) supplemented with 10% (v / v) fetal bovine serum (Sigma-Aldrich) and 1% (v / v) penicillin-streptomycin (Sigma-Aldrich). Briefly, 1 × 10 cells were cultured at 1 × 10 6MCF7 cells were seeded and grown in Corning® T75 flasks (Sigma-Aldrich) in an incubator at 37°C under 5% CO2. At 80-90% confluency, MCF7 cells were washed with Dulbecco's phosphate-buffered saline (Sigma-Aldrich) for 30 seconds at room temperature, followed by incubation with trypsin-EDTA (Sigma-Aldrich) for 5 minutes at 37°C under 5% CO2. The trypsin solution was then neutralized with a 1:1 volume of MCF7 growth medium. The cell suspension was then collected in a Corning® 50 mL centrifuge tube (Sigma-Aldrich) and centrifuged at 300 x g for 10 minutes at room temperature. The resulting supernatant was discarded, and the MCF7 pellet was resuspended in 5-10 mL of pre-warmed MCF7 growth medium. Total viable cells were counted using an Invitrogen™ Countess Cell Counter (Thermo Fisher Scientific, New South Wales, Australia) and 9 × 10 4 Live MCF7 cells were seeded in triplicate into Corning® 96-well white polystyrene microplates (Sigma). The seeded HCMs were incubated overnight at 37°C in a 5% CO2 incubator before treatment with bisantrene (day 1).

[0132] (MDA-MB-231 breast cancer cells) The human breast cancer cell line NDA-MB-231 was cultured in DMEM supplemented with 10% fetal bovine serum (FBS), 2 mM L-glutamine, and 2% HEPES in a humidified chamber containing 5% CO at 37°C. Cells were maintained adherent.

[0133] (H929 multiple myeloma cells) Human multiple myeloma NCI-H929 cells were cultured in RPMI-1640 growth medium (Sigma-Aldrich) supplemented with 2 mM L-glutamine (Thermo Fisher Scientific), 10% (v / v) fetal bovine serum (Sigma-Aldrich), and 0.05 mM 2-mercaptoethanol (Sigma-Aldrich). Briefly, 0.5 × 10 cells were cultured at 100°C. 6H929 cells were seeded and grown in Corning® T75 flasks (Sigma-Aldrich) in a 37°C incubator under 5% CO2 conditions. At 80-90% confluency, the cell suspension was collected in a Corning® 50 mL centrifuge tube (Sigma-Aldrich) and centrifuged at 300 x g for 10 minutes at room temperature. The resulting supernatant was discarded, and the H929 pellet was resuspended in 5-10 mL of pre-warmed H929 growth medium. Total viable cells were counted using an Invitrogen™ Countess Cell Counter (Thermo Fisher Scientific, New South Wales, Australia) and 1 x 10 cells were collected. 4 Live H929 cells were seeded in triplicate onto Corning® 96-well white polystyrene microplates (Sigma). The seeded H929 cells were incubated overnight at 37°C in a 5% CO incubator and then treated with bisantrene (day 1).

[0134] (Cell Treatment) To evaluate the therapeutic potential of bisantrene, HCM and MCF7 cells were treated with bisantrene in combination with the commercially available anticancer drugs doxorubicin hydrochloride (optionally in combination with 500 nM trastuzumab (Tz; MedChemExpress, New Jersey, USA), daunorubicin hydrochloride, epirubicin hydrochloride (Dox, Dauno, and Epi; Sigma-Aldrich), carfilzomib, or bortezomib (CFZ and BTZ; MCE®)). Bisantrene was provided by RACE Oncology Ltd (Sydney, New South Wales, Australia) or obtained from MedChemExpress (MCE®, New Jersey, USA). Both bisantrene, CFZ, and BTZ were dissolved in dimethyl sulfoxide (DMSO; Sigma-Aldrich) to stock concentrations of 500 μM and 20 mM, respectively. Dox, Dauno, and Epi were dissolved in sterile water to a stock concentration of 20 mM each, and Tz was dissolved in sterile PBS to a stock concentration of 100 μM. A series of bisantrene concentrations was prepared in HCM (0.5% v / v serum), MCF7 (10% v / v serum), or H929 growth medium. The concentrations were 0 nM, 15.62 nM, 62.5 nM, 250 nM, 500 nM, 1 μM, 5 μM, and 10 μM. Dox, Epi, or CFZ was then diluted at each bisantrene concentration to a final concentration of 1 μM. On day 0, spent medium was removed from each well, and 200 μL of bisantrene treatment (with or without anticancer drug) was added as appropriate. The cells were incubated overnight at 37°C under 5% CO2. Vehicle controls were incubated with medium supplemented with either sterile water or DMSO. After 24 hours, the spent medium was removed and replaced with 200 μL of fresh bisantrene treatment, which was repeated at 48 and 72 hours.Cell viability was then assessed at 24, 48, and 72 h, or 72 h only, using the CellTiter-Glo® Luminescent Cell Viability Assay (Promega, New South Wales, Australia) according to the manufacturer's instructions and recording luminescence using a Cytation™ 3 Cell Imaging Multi-Mode Reader (BioTek Instruments, Vermont, USA). Similar experiments were also performed using the FTO inhibitors FB23 / 2 and brequinar instead of bisantrene. Both FB23 / 2 and brequinar (MedChemExpress) were dissolved in dimethyl sulfoxide (DMSO; Sigma-Aldrich) to stock concentrations of 40 mM and 20 mM, respectively.

[0135] Example 2 - Bisantrene reduces the toxicity of cardiotoxic agents to cardiomyocytes As shown in Figure 1, both doxorubicin and carfilzomib exhibited progressive toxicity (measured as loss of cell viability) to cardiomyocytes over time (24, 48, and 72 hours), as expected. However, and surprisingly, as also shown in Figure 1, bisantrene exhibited mild progressive toxicity to cardiomyocytes over time (24, 48, and 72 hours), but also concentration-dependent protection of cardiomyocytes against the progressive toxicity of Dox and CFZ to cardiomyocytes.

[0136] Bisantrene also reduced the combined toxicity of doxorubicin and trastuzumab when cardiomyocytes were cultured in the presence of doxorubicin and trastuzumab (trastuzumab is also known to cause cardiotoxicity when administered alone). As shown in Figure 2, culturing cardiomyocytes in the presence of trastuzumab and doxorubicin resulted in decreased cardiomyocyte viability compared to culturing in the presence of doxorubicin alone. Increasing the concentration of bisantrene in the culture medium, whether containing doxorubicin alone or doxorubicin and trastuzumab, resulted in decreased viability. Although cardiomyocyte viability in the presence of trastuzumab and doxorubicin was lower than in the presence of doxorubicin alone, the relative difference in viability was no longer significant when 5 μM bisantrene was added to the culture medium compared to when 1 μM bisantrene was added to the culture medium.

[0137] Figure 5 shows that the cardiotoxicity-rescue effect of bisantrene on cardiomyocytes extended to the anthracycline, daunorubicin, when both agents were administered at 1 μM. Similarly, a trend toward statistical significance was observed for co-administration of epirubicin and bisantrene at 1 μM, as shown in Figure 6. These observations confirm that the cardioprotective effect of bisantrene is general across the anthracycline class of chemotherapeutic agents.

[0138] Figure 7 further shows that the cardioprotective effect of bisantrene is not specific to the anthracycline class, and significant rescue of myocardial cytotoxicity is observed for both the irreversible proteasome inhibitor, carfilzomib, and the reversible inhibitor, bortezomib, when each agent is co-administered with bisantrene.

[0139] Example 3 - Bisantrene is toxic to cancer cells and does not protect them against the toxicity of cardiotoxic agents The results reported in Example 2, protection of cardiomyocytes against the toxicity of cardiotoxic agents, were surprising given previous reports of bisantrene's efficacy against various cancer cells. As shown in Figures 3 and 4, these reports were confirmed, and experimental results demonstrate the concentration-dependent toxicity of bisantrene against MCF7 breast cancer cells and H929 multiple myeloma cells. Furthermore, as shown in Figures 3 and 4, these experiments demonstrated that bisantrene did not interfere with the toxicity of doxorubicin or carfilzomib, demonstrating additive toxicity of bisantrene, at least against MCF7 breast cancer cells (compared to the increased protection afforded to cardiomyocytes).

[0140] When tested against MDA-MB-231 breast cancer cells, the concentration-dependent toxic effects of bisantrene on breast cancer cells were again confirmed, and when the data were analyzed via Webb synergy analysis (see Figures 8-10), they were found to be at least additive, if not synergistic, with the effects of doxorubicin, epirubicin, or carfilzomib, with the enhanced combined activity appearing to be more pronounced at lower concentrations of both drugs.

[0141] Example 4 - Bisantrene effect may not be related to its FTO inhibitor properties Bisantrene is a compound with direct cytotoxicity as well as genomic and immunological mechanisms of action, including as an inhibitor (IC50 142nM) of fat mass and obesity-associated protein (FTO), an RNA N6-methyladenosine (m6A) demethylase (Su, R., Dong, L., Li, Y., Gao, M., Han, L., Wunderlich, M., et al. (2020). Targeting FTO Suppresses Cancer Stem Cell Maintenance and Immune Evasion. Cancer Cell, 38(1), 79-96.e11).

[0142] Several experiments were performed to investigate whether bisantrene's cardioprotective properties and its additive toxic effects on cancer cells (when combined with doxorubicin) were due to its FTO inhibitor activity.

[0143] Primary HCMs were cultured in the presence of the known FTO inhibitors brequinar and FB23 / 2, alone or in combination with doxorubicin or carfilzomib. As shown in Figures 11 and 12, neither brequinar nor FB23 / 2 exhibited appreciable toxicity to primary HCMs cultured in their presence for up to 72 hours at FTO inhibitor concentrations up to 10 μM. However, as also shown in Figures 11 and 12, unlike bisantrene, these FTO inhibitors did not reduce the toxicity of doxorubicin or carfilzomib to primary HCMs.

[0144] MCF7 breast cancer cells and H929 multiple myeloma cells were also cultured in the presence of the known FTO inhibitors brequinar and FB23 / 2, alone or in combination with doxorubicin, to determine whether these FTO inhibitors could act in combination with doxorubicin to further reduce cancer cell viability. As shown in Figure 13, neither brequinar nor FB23 / 2, at concentrations up to 10 μM, further reduced the viability of MCF7 breast cancer cells or H929 multiple myeloma cells cultured in the presence of 1 μM doxorubicin for up to 72 hours.

[0145] These results suggest that, at least for primary human cardiomyocytes, the protective effect of bisantrene (against the cardiotoxicity of doxorubicin and carfilzomib) may not be due to its FTO inhibitor activity. These results also suggest that the enhanced toxic effects (over 72 hours) between bisantrene and doxorubicin or epirubicin, at least for MCF7 or MDA-MB-231 breast cancer cells and H929 multiple myeloma cells, cannot be attributed to the FTO inhibitor activity of bisantrene.

[0146] Example 5 - Bisantrene may have an optimal dose or dose ratio for its cardioprotective effect when co-administered with doxorubicin in mice The cardioprotective effects of bisantrene were further studied in mice. Specifically, established cardiotoxicity markers, including left ventricular fractional shortening, cardiac E / A ratio, left ventricular diastolic function, left ventricular dilation, cardiac output, plasma triglyceride levels, plasma creatine kinase-MB (CK-MB), plasma LDH (lactate dehydrogenase), and plasma brain natriuretic peptide (BNP), were monitored for up to 42 days after the initiation of treatment delivered four times weekly (thus, the final treatment was on day 21).

[0147] The treatments studied included: Vehicle only Bisantrene (7.33 mg / kg) + vehicle Doxorubicin (5 mg / kg) + vehicle Vehicle + doxorubicin (5 mg / kg) + bisantrene (3.67 mg / kg) ("Dox+Bis 1:1") Vehicle + doxorubicin (5 mg / kg) + bisantrene (7.33 mg / kg) ("Dox+Bis 1:2") Vehicle + doxorubicin (4 mg / kg) + bisantrene (5.65 mg / kg) ("Dox+Bis 1:1*") Includes:

[0148] Treatments were delivered intravenously to mice on days 0, 7, 14, and 21. Mice underwent echocardiography on days 0, 21, and 42 to determine LVEF, FS, EDV, ESV, CO, E wave, A wave, E', E / A, and E / E', and blood samples were collected on days 0, 21, and 42 to assay for cardiac troponin I levels, BNP levels, CK, LDH levels, and triglycerides.

[0149] The data are shown in Figures 14 to 17.

[0150] As can be seen from Figure 14, 7.33 mg / kg (approximately 22 mg / m 2While there is suggestion of a mild (but not significant) adverse effect of bisantrene on left ventricular fractional shortening and cardiac E / A ratio at 1:1 / dose, doxorubicin had a significant cardiotoxic effect in vivo based on these parameters 21 days after treatment initiation that worsened by day 42. When bisantrene was coadministered with doxorubicin at a 1:1 or 1:2 molar ratio, doxorubicin cardiotoxicity was significantly attenuated based on these parameters, with slightly better protection afforded by Dox+Bis 1:2 treatment and lesser effects observed with Dox+Bis 80% 1:2 treatment.

[0151] As can be seen from Figure 15, 7.33 mg / kg (approximately 22 mg / m 2 Although there is some suggestion (but not significant) of a mild adverse effect of bisantrene on left ventricular diastolic function and dilation and cardiac output at 100 mg / dose, doxorubicin had a significant cardiotoxic effect in vivo based on these parameters at 42 days after treatment initiation, but not by 21 days. Interestingly, Dox+Bis 1:1 treatment appeared to attenuate the cardiotoxic effects of doxorubicin, as measured by left ventricular diastolic function and dilation, compared with both other combination treatments (which had higher bisantrene doses). Looking at cardiac output, the reduction in doxorubicin cardiotoxicity was slightly greater with Dox+Bis 1:2 treatment compared with Dox+Bis 1:1 treatment, with a lesser effect observed with Dox+Bis 80% 1:2 treatment.

[0152] Figure 16 shows the significant results resulting from the blood tests.

[0153] Figure 16A shows a slight (but not significant) increase in plasma triglycerides on day 21 for bisantrene-only treatment, but not on day 42 compared to vehicle-only controls, whereas doxorubicin treatment significantly elevated plasma triglyceride levels on days 21 and 42. Co-administration of bisantrene in the Dox+Bis 1:1 treatment resulted in significant attenuation of the doxorubicin-induced increase in plasma triglycerides on days 21 and 42 for both the Dox+Bis 1:1 and Dox+Bis 1:2 treatments, with the Dox+Bis 80% 1:2 treatment having less of an effect.

[0154] As shown in Figure 16B, a similar effect was seen on brain natriuretic peptide levels, but the results of Dox+Bis 1:2 treatment were not significant at 42 days or 21 days with respect to attenuating any effects induced by doxorubicin.

[0155] Figure 16C shows the results for creatine kinase-MB (CK-MB) levels. While doxorubicin alone resulted in a significant increase in plasma CK-MB on day 21, coadministration of bisantrene and doxorubicin, as well as both the Dox+Bis 1:1 and Dox+Bis 1:2 treatments, suppressed CK-MB levels to levels comparable to or close to those of the vehicle-only control. CK-MB levels were not significantly elevated with either treatment on day 42.

[0156] None of the co-administered treatments appeared to have any significant effect on plasma LDH levels, eosinophil, neutrophil, basophil or lymphocyte levels or viability, hematocrit or hemoglobin levels, mean corpuscular volume, platelet, red blood cell or white blood cell counts, or heart weight / body weight ratio.

[0157] As shown in Figure 17, bisantrene co-administered with doxorubicin at a 1:1 molar ratio significantly reduced histological cardiac fibrosis at day 21 compared to mice receiving the same dose of doxorubicin. This result corroborates the functional echocardiography and blood biomarker results, demonstrating that bisantrene was able to reduce accepted structural markers of anthracycline-induced cardiac tissue damage.

[0158] Overall, bisantrene appeared to increase left ventricular systolic (FS) and diastolic function (E / A, E / E') and improve cardiac function (CO, BNP) in doxorubicin-treated mice, an effect that may be caused by reducing early myocardial damage (CK-MB and cardiac fibrosis).

[0159] In this particular study, taking all factors into account, the protective effect of bisantrene was significantly greater when used in a 1:1 ratio with doxorubicin (or approximately 3.67 mg / m 2 Although the effects of Doxorubicin appeared to be greatest at doses of 1:2, the Dox+Bis combination treatment significantly reduced certain doxorubicin cardiotoxicity marker parameters compared with the Dox+Bis 1:1 treatment.

[0160] (Conclusion) This study surprisingly demonstrated that bisantrene is not only toxic to cancer cells, but can also act additively, if not synergistically, when combined with doxorubicin, epirubicin, or carfilzomib, but also reduces the toxicity of cardiotoxic therapeutic agents such as doxorubicin, daunorubicin, epirubicin, carfilzomib, and bortezomib to cardiomyocytes. This latter result potentially opens the door not only to improved treatments for various cancer types, but also to greater patient tolerance of cardiotoxic therapeutic agents or longer / greater patient exposure to such agents.

Claims

1. A composition for use in a method for preventing or reducing drug-induced cardiotoxicity in a subject caused by a cardiotoxic agent, the method comprising administering to the subject an effective amount of a cardioprotective agent comprising bisantrene or a derivative thereof, or a pharmaceutically acceptable salt of bisantrene or a derivative thereof.

2. 10. The composition of claim 1 for use in a method for treating cancer in a subject, the method comprising: a) an effective amount of a cardiotoxic chemotherapeutic agent; b) an effective amount of a cardioprotectant comprising bisantrene or a derivative thereof, or a pharmaceutically acceptable salt of bisantrene or a derivative thereof; A composition comprising the step of administering

3. 3. The composition of claim 2, wherein the cancer is selected from the group consisting of breast cancer, myeloma, acute myeloid leukemia, melanoma, and clear cell renal carcinoma.

4. A composition described in any one of claims 1 to 3, wherein the method includes a step of administering the cardioprotective agent to the subject before, simultaneously with, or after administration of a cardiotoxic agent to the subject.

5. The composition of any one of claims 1 to 3, wherein the cardiotoxic agent and the cardioprotective agent are administered simultaneously in a single composition.

6. The composition of any one of claims 1 to 3, wherein the cardiotoxic agent is an anthracycline or a pharmaceutically acceptable salt thereof.

7. 7. The composition of claim 6, wherein the anthracycline is selected from the group consisting of daunorubicin, doxorubicin, epirubicin, idarubicin, mitoxantrone, and valrubicin.

8. 8. The composition of claim 7, wherein the anthracycline is doxorubicin, daunorubicin, or epirubicin.

9. The composition of any one of claims 1 to 3, wherein the cardiotoxic agent is a proteasome inhibitor, optionally carfilzomib or bortezomib.

10. 7. The composition of claim 6, wherein the dose of the cardiotoxic agent is at least 10% lower than the required dose of the cardiotoxic agent when administered without the cardioprotective agent to achieve the same target outcome.

11. 11. The composition of claim 10, wherein the dosage of the cardiotoxic agent and the cardioprotective agent is in a molar ratio of about 1:3 to about 3:

1.

12. The composition of any one of claims 1 to 3, wherein the doses of the cardiotoxic agent and the cardioprotective agent are in a molar ratio of about 1:

1.

13. The dose of the cardioprotectant is about 5 mg / m for 4 weeks. ２ / week ~ approx. 100mg / m ２ The composition according to any one of claims 1 to 3, wherein the daily dose is 100 mg / week.

14. The composition of any one of claims 1 to 3, wherein the cardioprotectant is bisantrene or a pharmaceutically acceptable salt thereof.

15. Bisantrene at approximately 5 mg / m for 4 weeks ２ / week ~ approx. 50mg / m ２ 15. The composition of claim 14, wherein the subject is administered a weekly dose of bisantrene or a pharmaceutically acceptable salt of bisantrene administered at the same time in a molar equivalent dose, and the cardiotoxic agent is an anthracycline selected from the group consisting of daunorubicin, doxorubicin, epirubicin, idarubicin, mitoxantrone, and valrubicin.

16. A pharmaceutical composition comprising a cardioprotective agent comprising bisantrene or a derivative thereof, or a pharmaceutically acceptable salt of bisantrene or a derivative thereof, and a cardiotoxic therapeutic agent, wherein the composition has reduced cardiotoxicity compared to a composition comprising the cardiotoxic therapeutic agent but not the cardioprotective agent.

17. 17. The composition of claim 16, wherein the cardiotoxic therapeutic agent is a chemotherapeutic agent.

18. 18. The composition of claim 17, wherein the chemotherapeutic agent is an anthracycline or a pharmaceutically acceptable salt thereof.

19. 19. The composition of claim 18, wherein the anthracycline is selected from the group consisting of daunorubicin, doxorubicin, epirubicin, idarubicin, mitoxantrone, and valrubicin.

20. 20. The composition of claim 19, wherein the anthracycline is doxorubicin, daunorubicin, or epirubicin.

21. The composition of claim 17, wherein the cardiotoxic agent is a proteasome inhibitor, optionally carfilzomib or bortezomib.

22. The composition according to any one of claims 16 to 21 for the treatment of cancer.

23. 22. The composition of any one of claims 16 to 21, wherein the molar ratio of cardiotoxic agent to cardioprotective agent in the composition is from about 1:3 to about 3:

1.

24. 22. The composition of any one of claims 16 to 21, wherein the molar ratio of cardiotoxic agent to cardioprotective agent in the composition is about 1:

1.

25. Approximately 5 mg / m for 4 weeks ２ / week ~ approx. 25mg / m ２ 22. The composition of any one of claims 16 to 21, adapted to deliver a dose of a cardioprotectant per week.

26. The composition of any one of claims 16 to 21, wherein the cardioprotectant is bisantrene or a pharmaceutically acceptable salt thereof.

27. Bisantrene at approximately 5 mg / m for 4 weeks ２ / week ~ approx. 50mg / m ２ 27. The composition of claim 26, adapted to deliver to a subject a weekly dose of bisantrene or a pharmaceutically acceptable salt of bisantrene at the same time in a molar equivalent dose, wherein the cardiotoxic agent is an anthracycline selected from the group consisting of daunorubicin, doxorubicin, epirubicin, idarubicin, mitoxantrone, and valrubicin.

28. A kit for preventing or reducing drug-induced cardiotoxicity in a subject caused by a cardiotoxic therapeutic agent, the kit comprising a cardioprotective agent comprising bisantrene or a derivative thereof, or a pharmaceutically acceptable salt of bisantrene or a derivative thereof, and the cardiotoxic therapeutic agent.

29. 29. The kit according to claim 28, for treating a subject with the method according to any one of claims 1 to 3.

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