Cardiac treatment

Abstract

The present invention relates to methods and pharmaceutical compositions for reducing or preventing myocardial ischaemic-reperfusion injury, comprising administering an effective amount of a compound of formula (I), or a pharmaceutically acceptable salt thereof.

Description

[0001] CARDIAC TREATMENT FIELD OF THE INVENTION The present invention relates to methods, and pharmaceutical compositions for reducing or preventing myocardial ischaemic-reperfusion injury, including reducing or preventing inflammation, cellular dysfunction and myocardial cell death in heart tissue of a subject resulting from reperfusion of ischaemic tissue. BACKGROUND Ischaemic heart disease, secondary to atherosclerosis, is the world’s leading cause of death. Myocardial cell death or a myocardial infarction occurs as a consequence of ischemia or ischemia-reperfusion (Konstantinidis, K. et al. Arterioscler. Thromb. Vasc. Biol. 32, 1552–1562 (2012)). Reducing infarct (IF) size and protecting heart function to prevent heart failure (HF) are essential aims of treatment of acute coronary syndromes (ACS), such as myocardial infarction (MI), given the paucity of intrinsic regenerative capacity. Primary percutaneous coronary intervention (PCI) is commonly used in treatment of ACS to restore flow to the culprit vessel that provoked the ACS. Timely PCI, resulting in myocardial reperfusion of the affected area, can salvage viable myocardium, limit myocardial IF size, and prevent the onset of heart failure. Paradoxically however, reperfusion can cause further inflammation and drive up to 50% of the final IF size (Yellon, D.M. & Hausenloy, D.J. N Engl J Med 357, 1121-1135 (2007)). In this regard, sudden trauma caused by reperfusion initiates a local maelstrom involving injured and denuded endothelium, platelet activation, neutrophil and monocyte adhesion and infiltration, causing further myocardial IF formation in addition to that already caused by the ACS. This damage resulting from reperfusion is a phenomenon known as myocardial ischaemia-reperfusion (M / IR) injury. Myocardial ischaemic-reperfusion injury can lead to post-infarct left ventricular (LV) remodeling and fibrosis, a hallmark feature of heart failure (HF). LV remodelling occurs in 1 in 3 ST-elevated myocardial infarction (STEMI) patients within 6 months of PCI and HF develops in 20–30% of patients at 1 year (Sulo, G., et al. J Am Heart Assoc 5, e002667 (2016)). Globally, 26 million people suffer with HF, of which 5.6 million reside in the US. The US alone is expecting annual costs associated with HF to exceed USD70 billion by 2030 as its incidence and attributable cost increases. There is no approved cardioprotective agent that can reduce IF size in the setting of MI. Emerging anti-inflammatory drugs (viz. canakinumab, tocilizumab, colchicine, and ziltivekimab) have not demonstrated reduced occurrence of recurrent ACS after acute MI and may leave patients at risk for infection. M / IR injury causes myocardial cell death and the development of cardiac dysfunction, triggering ventricular dilation, cardiac fibrosis, impairment of contractile function and HF (Murphy, S.P., et al. J Am Coll Cardiol 75, 1324-1340 (2020)). Complex molecular and cellular processes underpin these events. These involve a range of inducible pro-inflammatory responses including transcription factor (e.g., KLF5, EGR1), matrix metalloproteinase (e.g., MMP2, MMP3) and cytokine (e.g., TNFα, IL1β, IL6) gene expression. An inflammasome (NLRP3) is activated in cardiomyocytes and circulating inflammatory cells, causing activation of caspase-1 and consequent maturation of IL1β. Release of damage-associated molecular patterns (DAMPs, e.g., S100A8, HMGB1, HSP) causes cardiomyocyte death through DAMP- sensing receptors (such as Toll-like receptors, TLR2, TLR4) and stimulating leukocyte recruitment to the IF zone. What is needed is an agent that is capable of inhibiting or reducing myocardial cell death, inflammation and cardiac dysfunction in heart tissue following reperfusion. SUMMARY The inventors have found that a compound, BT2 (a compound of formula (II)) provides cardioprotection of heart tissue from M / IR injury following reperfusion. In this regard, the inventors have found that BT2 reduces or prevents inflammation and myocardial cell death in heart tissue following reperfusion, thereby reducing IF size and reducing or preventing left ventricular remodelling, cardiac fibrosis, loss of heart function, and reduces the risk of heart failure (HF), following reperfusion. A first aspect provides a method for reducing or preventing myocardial ischaemic- perfusion (M / IR) injury in a subject, comprising administering an effective amount of a compound of formula (I), or a pharmaceutically acceptable salt thereof: wherein: R1is straight or branched C1-C6alkyl; and R2is straight or branched C1-C6alkyl, or R2is wherein q is 1, 2, 3 or 4; and R3is straight or branched C1-C6alkyl. An alternative first aspect provides a compound of formula (I), or a pharmaceutically acceptable salt thereof, for use in reducing or preventing myocardial ischaemic-perfusion injury in a subject; or use of a compound of formula (I), or a pharmaceutically acceptable salt thereof, in the manufacture of a medicament for reducing or preventing myocardial ischaemic-perfusion injury in a subject. A second aspect provides a method for reducing or preventing myocardial ischaemic- perfusion injury in a subject, comprising administering an effective amount of a compound of formula (II), or a pharmaceutically acceptable salt thereof: Formula (II) . An alternative first aspect provides a compound of formula (II), or a pharmaceutically acceptable salt thereof, for use for reducing or preventing myocardial ischaemic-perfusion injury in a subject; or use of a compound of formula (II), or a pharmaceutically acceptable salt thereof, in the manufacture of a medicament for reducing or preventing myocardial ischaemic-perfusion injury in a subject. A third aspect provides a method of reducing or preventing inflammation and / or myocardial cell death in heart tissue of a subject following reperfusion of ischaemic tissue in the heart of the subject, comprising administering an effective amount of a compound of formula (I), or a pharmaceutically acceptable salt thereof. An alternative third aspect provides a compound of formula (I), or a pharmaceutically acceptable salt thereof, for use in reducing or preventing inflammation and / or myocardial cell death in heart tissue of a subject following reperfusion of ischaemic tissue in the heart of the subject; or use of a compound of formula (I), or a pharmaceutically acceptable salt thereof, in the manufacture of a medicament for reducing or preventing inflammation and / or myocardial cell death in heart tissue of a subject following reperfusion of ischaemic tissue in the heart of the subject. A fourth aspect provides a method of reducing or preventing inflammation and / or myocardial cell death in heart tissue of a subject following reperfusion of ischaemic tissue in the heart of the subject, comprising administering an effective amount of a compound of formula (II), or a pharmaceutically acceptable salt thereof An alternative fourth aspect provides a compound of formula (II), or a pharmaceutically acceptable salt thereof, for use in reducing or preventing inflammation and / or myocardial cell death in heart tissue of a subject following reperfusion of ischaemic tissue in the heart of the subject; or use of a compound of formula (II), or a pharmaceutically acceptable salt thereof, in the manufacture of a medicament for reducing or preventing inflammation and / or myocardial cell death in heart tissue of a subject following reperfusion of ischaemic tissue in the heart of the subject. A fifth aspect provides a method of reducing or preventing left ventricular remodelling in a subject following reperfusion of ischaemic tissue in the heart of the subject, comprising administering an effective amount of a compound of formula (I), or a pharmaceutically acceptable salt thereof. An alternative fifth aspect provides a compound of formula (I), or a pharmaceutically acceptable salt thereof, for use in reducing or preventing left ventricular remodelling in a subject following reperfusion of ischaemic tissue in the heart of the subject; or use of a compound of formula (I), or a pharmaceutically acceptable salt thereof, in the manufacture of a medicament for reducing or preventing left ventricular remodelling following reperfusion of ischaemic tissue in the heart of the subject. A sixth aspect provides a method of reducing or preventing left ventricular remodelling in a subject following reperfusion of ischaemic tissue in the heart of the subject, comprising administering an effective amount of a compound of formula (II), or a pharmaceutically acceptable salt thereof. An alternative sixth aspect provides a compound of formula (II), or a pharmaceutically acceptable salt thereof, for use in reducing or preventing left ventricular remodelling in a subject following reperfusion of ischaemic tissue in the heart of the subject; or use of a compound of formula (II), or a pharmaceutically acceptable salt thereof, in the manufacture of a medicament for reducing or preventing left ventricular remodelling in a subject following reperfusion of ischaemic tissue in the heart of the subject. A seventh aspect provides a method of reducing or preventing cardiac fibrosis following reperfusion of ischaemic tissue in the heart of the subject, comprising administering an effective amount of a compound of formula (I), or a pharmaceutically acceptable salt thereof. An alternative seventh aspect provides a compound of formula (I), or a pharmaceutically acceptable salt thereof, for use in reducing or preventing cardiac fibrosis following reperfusion of ischaemic tissue in the heart of the subject; or use of a compound of formula (I), or a pharmaceutically acceptable salt thereof, in the manufacture of a medicament for reducing or preventing cardiac fibrosis following reperfusion of ischaemic tissue in the heart of the subject. An eighth aspect provides a method of reducing or preventing cardiac fibrosis following reperfusion of ischaemic tissue in the heart of the subject, comprising administering an effective amount of a compound of formula (II), or a pharmaceutically acceptable salt thereof. An alternative eighth aspect provides a compound of formula (II), or a pharmaceutically acceptable salt thereof, for use in reducing or preventing cardiac fibrosis reperfusion of ischaemic tissue in the heart of the subject; or use of a compound of formula (II), or a pharmaceutically acceptable salt thereof, in the manufacture of a medicament for reducing or preventing cardiac fibrosis following reperfusion of ischaemic tissue in the heart of the subject. A ninth aspect provides a method of reducing or preventing loss of heart function in a subject following reperfusion of ischaemic tissue in the heart of the subject, comprising administering an effective amount of a compound of formula (I), or a pharmaceutically acceptable salt thereof. An alternative ninth aspect provides a compound of formula (I), or a pharmaceutically acceptable salt thereof, for use in reducing or preventing loss of heart function in a subject following reperfusion of ischaemic tissue in the heart of the subject; or use of a compound of formula (I), or a pharmaceutically acceptable salt thereof, in the manufacture of a medicament for reducing or preventing loss of heart function in a subject following reperfusion of ischaemic tissue in the heart of the subject. A tenth aspect provides a method of reducing or preventing loss of heart function in a subject following reperfusion of ischaemic tissue in the heart of the subject, comprising administering an effective amount of a compound of formula (II), or a pharmaceutically acceptable salt thereof. An alternative tenth aspect provides a compound of formula (II), or a pharmaceutically acceptable salt thereof, for use in reducing or preventing loss of heart function in a subject following reperfusion of ischaemic tissue in the heart of the subject; or use of a compound of formula (II), or a pharmaceutically acceptable salt thereof, in the manufacture of a medicament for reducing or preventing loss of heart function in a subject following reperfusion of ischaemic tissue in the heart of the subject. An eleventh aspect provides a method of reducing the risk of heart failure following reperfusion of ischaemic tissue in the heart of the subject, comprising administering an effective amount of a compound of formula (I), or a pharmaceutically acceptable salt thereof. An alternative eleventh aspect provides a compound of formula (I), or a pharmaceutically acceptable salt thereof, for use in reducing the risk of heart failure following reperfusion of ischaemic tissue in the heart of the subject; or use of a compound of formula (I), or a pharmaceutically acceptable salt thereof, in the manufacture of a medicament for reducing the risk of heart failure following reperfusion of ischaemic tissue in the heart of the subject. A twelfth aspect provides a method of reducing the risk of heart failure following reperfusion of ischaemic tissue in the heart of the subject, comprising administering an effective amount of a compound of formula (II), or a pharmaceutically acceptable salt thereof. An alternative twelfth aspect provides a compound of formula (II), or a pharmaceutically acceptable salt thereof, for use in reducing the risk of heart failure following reperfusion of ischaemic tissue in the heart of the subject; or use of a compound of formula (II), or a pharmaceutically acceptable salt thereof, in the manufacture of a medicament for reducing the risk of heart failure following reperfusion of ischaemic tissue in the heart of the subject. A thirteenth aspect provides a method of reducing or preventing heart failure following reperfusion of ischaemic tissue in the heart of the subject, comprising administering an effective amount of a compound of formula (I), or a pharmaceutically acceptable salt thereof. An alternative thirteenth aspect provides a compound of formula (I), or a pharmaceutically acceptable salt thereof, for use in reducing or preventing heart failure following reperfusion of ischaemic tissue in the heart of the subject; or use of a compound of formula (I), or a pharmaceutically acceptable salt thereof, in the manufacture of a medicament for reducing or preventing heart failure following reperfusion of ischaemic tissue in the heart of the subject. A fourteenth aspect provides a method of reducing or preventing heart failure following reperfusion of ischaemic tissue in the heart of the subject, comprising administering an effective amount of a compound of formula (II), or a pharmaceutically acceptable salt thereof. An alternative fourteenth aspect provides a compound of formula (II), or a pharmaceutically acceptable salt thereof, for use in reducing or preventing heart failure following reperfusion of ischaemic tissue in the heart of the subject; or use of a compound of formula (II), or a pharmaceutically acceptable salt thereof, in the manufacture of a medicament for reducing or preventing heart failure following reperfusion of ischaemic tissue in the heart of the subject. A fifteenth aspect provides a method of treating a subject suffering from an ischaemic event, comprising: (a) administering a compound of Formula (I), or a pharmaceutically acceptable salt thereof; and (b) restoring blood flow to ischaemic tissue. An alternative fifteenth aspect provides a compound of Formula (I), or a pharmaceutically acceptable salt thereof, for use in treating a subject suffering from ischaemic event, wherein the compound is to be administered to the subject prior to and / or during restoration of blood flow to ischaemic tissue; or use of a compound of Formula (I), or a pharmaceutically acceptable salt thereof, in the manufacture of a medicament for treating a subject suffering from an ischaemic event, wherein the compound is to be administered to the subject prior to and / or during restoration of blood flow to the ischaemic tissue. A sixteenth aspect provides a method of treating a subject suffering from ischaemic event, comprising: (c) administering a compound of Formula (II), or a pharmaceutically acceptable salt thereof; and (d) restoring blood flow to the ischaemic tissue. An alternative sixteenth aspect provides a compound of Formula (II), or a pharmaceutically acceptable salt thereof, for use in treating a subject suffering from an ischaemic event, wherein the compound is to be administered to the subject prior to and / or during restoration of blood flow to the ischaemic tissue; or use of a compound of Formula (I), or a pharmaceutically acceptable salt thereof, in the manufacture of a medicament for treating a subject suffering from an ischaemic event, wherein the compound is to be administered to the subject prior to and / or during restoration of blood flow to the ischaemic tissue. A seventeenth aspect provides a kit for treating myocardial ischaemic perfusion injury, the kit comprising a compound of formula (I), or a pharmaceutically acceptable salt thereof. An eighteenth aspect provides a kit for treating myocardial ischaemic perfusion injury, the kit comprising a compound of formula (II), or a pharmaceutically acceptable salt thereof. A ninteenth aspect provides a kit for reducing or preventing inflammation and / or myocardial cell death in the heart of a subject following reperfusion of ischaemic tissue in the heart of the subject, the kit comprising a compound of formula (I), or a pharmaceutically acceptable salt thereof. A twentieth aspect provides a kit for reducing or preventing inflammation and / or myocardial cell death in the heart of a subject following reperfusion of ischaemic tissue in the heart of the subject, the kit comprising a compound of formula (II), or a pharmaceutically acceptable salt thereof. BRIEF DESCRIPTION OF THE FIGURES Preferred embodiments of the present invention will now be described, by way of example only, with reference to the accompanying drawings: Figure 1 is a schematic representation of a study design for treatment of M / IR injury. SD rats were subjected to 30 min left anterior descending (LAD) artery ligation followed by reperfusion for 24 h or 2 weeks. BT2 (100 mg / kg) was delivered i.p.24 h prior to and again during ischaemia. ECG Lead III representative traces are shown. Figure 2A is image of representative cross sections of TTC stained heart with the AAR and IF indicated. Figure 2B is a graph showing the IF area as a percentage of the AAR 24 h after M / IR in rats treated with BT2, BT3 or vehicle. Figure 2C is a graph showing the AAR as a percentage of the left ventricle (AAR / LV) 24 h after M / IR in rats treated with BT2, BT3 or vehicle. BT3 is an inactive structural analogue of BT2. One-way ANOVA, n=8-9 / grp. Veh denotes vehicle (saline with 0.5% v / v Tween 80, 0.01% v / v DMSO). Figure 3 is a graph showing serum cardiac troponin I levels 24 h after M / IR injury in rats treated with BT2 or vehicle. MW test, n=9 / grp. Data remains statistically significant (**, t test) even when the outlier is not included. Figure 4 is a graph showing quantitation of immunochemical staining of p-ERK in the AAR 24 hours after M / IR injury. BT2 was delivered once during ischaemia. Staining was quantified using Image-Pro Plus. Data represent mean ± SEM of n=3-4 / grp. Statistical significance was assessed by t test. Figure 5 are graphs showing (A) heart rate, (B) PR interval and (C) QTc interval, 24 h after M / IR injury in rats treated with BT2 or vehicle. PR interval was calculated from ECG Lead II data. QT interval was corrected (c) using Bazett's formula (QTc = QT / √RR). t-test or MW test, n=11-12 / grp. Figure 6 is a graph showing circulating levels of BT2 in rats treated with BT2 or vehicle 24 h after M / IR as determined by mass spectrometry. Analysis of BT2 by high resolution UPLC MS. n=11-12 / grp. Figure 7 is graphs showing the effect of BT2 and vehicle (Veh) on (A) EF and (B) FS, 24h and 2 weeks after M / IR injury as determined by ultrasound. EF=ejection fraction; FS=fractional shortening. One-way ANOVA, n=6-9 / grp. Figure 8 is graphs showing the effect of BT2 and vehicle (Veh) on LVIDs and LVIDd 24 h and 2 weeks after M / IR injury as determined by ultrasound. LVIDs=left ventricular internal diameter end systole, LVIDd=left ventricular internal diameter end diastole. One-way ANOVA, n=6-9 / grp. Figure 9 is a graph showing quantification of immunohistochemical staining with (A) CD68 antibody, (B) CD4 antibody and (C) CD4 and FoxP3 antibody, in the AAR 24 h after M / IR injury; BT2 was delivered once during ischaemia. Staining was quantified using Image- Pro Plus. Data represent mean ± SEM of n=3-4 / grp. Statistical significance was assessed by t test. Figure 10A is a graph showing ST elevation between groups in hearts treated with BT2 or vehicle. Data sourced from Lead III traces from 6-lead ECG recorded 5-10 min after left coronary artery (LCA) ligation and quantitation shows unbiased ST-elevation between groups. t-test, n=6 / grp. Figure 10B is a graph showing quantification of fibrosis in hearts treated with BT2 or vehicle and left for 2 weeks after M / IR and stained with Sirius Red. Staining was quantified using Image Pro Plus. Rats were treated with BT2 or vehicle twice prior to reperfusion and left for 2 weeks. Cross sections of heart were stained with Sirius Red and fibrosis and quantified using Image Pro Plus. t-test, n=6 / grp. Figure 11 shows (A) a schematic of a study design in which SD rats were subjected to 30 min LAD ligation followed by reperfusion for 24 h. BT2 (100 mg / kg) was delivered during ischaemia; (B) a graph showing IF / AAR assessment 24 h after M / IR in rats treated with BT2 or vehicle; (C) a graph showing neutrophil counts in the AAR 24 h after M / IR in rats treated with BT2 or vehicle. Veh denotes vehicle (saline with 0.5% v / v Tween 80, 0.01% v / v DMSO). Mann-Whitney or t test, n=3 / grp. Figure 12 is graphs showing (A) EF, (B) FS, (C) LVIDs and (D) LVIDd, 24 h after M / IR injury as determined by ultrasound. EF=ejection fraction; FS=fractional shortening. LVIDs=left ventricular internal diameter end systole, LVIDd=left ventricular internal diameter end diastole. t test, n=3 / grp. Figure 13 is graphs showing sample values (GSE129935) of differentially expressed genes as indicated in monocytes of patients with AMI (n=12) compared with stable disease (CAD). Figure 14 is graphs showing sample values (GSE974) of differentially expressed genes as indicated in LV apex from patients with AMI (n=6) compared with those from healthy individuals (n=50). Figure 15 is a graph showing sample values (GSE66360) of differentially expressed genes as indicated in circulating endothelial cells from AMI patients (n=49) compared with those from healthy individuals (n=50). Figure 16 is graphs showing sample values (GSE61145, platform GLP6884) of differentially expressed genes as indicated in peripheral blood collected from ACS patients comparing STEMI (n=7), NSTEMI (n=10) and UA (m=9) patients with normal individuals (n=7). Figure 17 is graphs showing sample values (GSE59867) of differentially expressed genes as indicated in PBMCs isolated from patients with stable CAD (n=46) or STEMI patients developed heart failure (HF). Data analysed using KW or one-way ANOVA, as appropriate. Figure 18 is a graph and Venn diagrams showing the overlap of upregulated DEG in STEMI patients who later developed HF (PBMC collected within one day of AMI, GSE59867) with DEG downregulated by BT2 in the AAR in rats 4 or 24 h after M / IR injury. (A) denotes the number of DEG upregulated in STEMI patients who developed (HF up, n=9) or did not (Non-HF up, n=8) develop HF within 6 months relative to stable CAD patients (n=46). (B) denotes the number of DEG downregulated in STEMI patients who developed (HF down, n=9) or did not (Non-HF down, n=8) develop HF within 6 months. The percentages next to the circles in the Venn diagrams indicate the proportion of “HF up” or “HF down” DEG that overlapped with down- or upregulated DEG in the 4 and 24 h BT2 groups. Expression data was fitted to a negative binomial generalized linear model before undergoing a genewise quasi-likelihood F-test to identify DEGs. Figure 19 shows a representative gating strategy for sorting single nuclei on the BD FACS AriaI II sorter. Voltage was set for FSC-A and SSC-A to bring events to within scale. P1 gate was set to gate nuclei. Single event gating was deployed via P2 (FSC-H vs FSC-A) and P3 (SSC-H vs SSC-W). Additional *Sort Nuclei gate (DAPI V450-A vs DAPI V450-W) was deployed to gate for DAPI+single events (representing single nuclei) and sorted. DETAILED DESCRIPTION The compound BT2 (compound of formula (II)) is a dibenzoxazepinone shown previously to inhibit endothelial cell proliferation and migration, angiogenesis and wound repair. As described in the Examples, the inventors have now found that administration of BT2 prior to reperfusion: 1. reduces or prevents an increase in infarct size in the AAR following reperfusion of the myocardium; 2. preserves heart function and prevents adverse remodelling following reperfusion; 3. reduces the expression of genes associated with myocardial ischaemia, inflammation and heart failure; and 4. increases expression of genes associated with reduced inflammation, in heart tissue 24 hours after reperfusion of a heart in which ischaemia has been induced. In this regard, 24 hours after reperfusion was applied to the ischaemic heart, the inventors found that BT2 reduced expression of pro-inflammatory transcription factors such as ATF3, FOS, FOSL1 and FOSL2, reduced expression of metalloproteinases such as MMP10 and MMP16, reduced expression of pro-inflammatory cytokines and their receptors such as IL6, IL6R, IL17, IL17RA, IL18, TNFSF18 and TNNFRSF12A, reduced expression of macrophage biomarkers such as CD68, reduced expression of DAMP biomarkers such as HSPB1, reduced expression of DAMP receptors such as TLR1, TLR2, TLR6, TLR7, NLRP3, TREM1, TREM3 and TRPM2, reduced expression of CXC chemokines such as CXCL1 and CXCL6, and reduced expression of chemokines such as CCL2, CCL6, CCL17, CCL22 and CCL24. The inventor also found that at 24 h following reperfusion, BT2 increased the expression of the Treg biomarker FOXP3, the autophagy regulator TRIM50, the negative regulator of inflammation CTLA2A, Treg suppressor biomarker TCAM1 and antioxidant enzyme SOD3. Thus, BT2 is effective in reducing inflammation, immune cell function and regulating transcription in heart tissue following reperfusion of ischaemic tissue in the heart of a subject. The inventors further showed that BT2 is also cardioprotective when it is administered once. BT2 administered as a single dose inhibited both IF / AAR and neutrophil infiltration (myocardial inflammation). Further, a single dose of BT2 increased both EF and FS and reduced cardiac remodelling. The inventors have shown that BT2 reduces the infarct size resulting from reperfusion injury. Thus, BT2 is effective in reducing the extent of myocardial cell death following reperfusion of ischaemic tissue in the heart of a subject. The inventors therefore envisage that the compound of formula (I) and (II), or a pharmaceutically acceptable salt thereof, will be effective for use as a cardioprotective agent to reduce inflammation and myocardial damage inflicted in heart tissue following reperfusion. One aspect therefore provides a method for reducing or preventing myocardial ischaemic-perfusion injury in a subject. The myocardial ischaemic-perfusion injury results from reperfusion of ischaemic tissue in the myocardium of the subject. The method comprises administering an effective amount of a compound of formula (I), or a pharmaceutically acceptable salt thereof: wherein: R1is straight or branched C1-C6alkyl; and R2is straight or branched C1-C6alkyl, or R2is R3qO wherein q is 1, 2, 3 or 4; and R3is straight or branched C1-C6alkyl. In one embodiment, a compound of formula (I) has the structure of formula (II): Formula (II). M / IR injury in a subject administered a compound of Formula (I) or (II), or a pharmaceutically acceptable salt thereof, is reduced relative to the M / IR injury that would have occurred had the subject not been administered the compound of Formula (I) or (II), or a pharmaceutically acceptable salt thereof. In one embodiment, the reducing or preventing myocardial ischaemic-perfusion injury reduces or prevents myocardial cell death in heart tissue following reperfusion of ischaemic tissue in the heart of a subject. Myocardial cell death in heart tissue following reperfusion of ischaemic tissue in the heart of a subject is myocardial cell death caused by reperfusion of ischaemic tissue in the heart of a subject. Myocardial cell death in heart tissue of a subject administered a compound of Formula (I) or (II), or a pharmaceutically acceptable salt thereof, is reduced relative to the myocardial cell death in the heart tissue of the subject that would have occurred had the subject not been administered the compound of Formula (I) or (II), or a pharmaceutically acceptable salt thereof. In one embodiment, the reducing or preventing myocardial ischaemic-perfusion injury reduces or prevents inflammation in heart tissue following reperfusion of ischaemic tissue in the heart of a subject. Inflammation in heart tissue of a subject administered a compound of Formula (I) or (II), or a pharmaceutically acceptable salt thereof, is reduced relative to the inflammation in the heart tissue of the subject that would have occurred had the subject not been administered the compound of Formula (I) or (II), or a pharmaceutically acceptable salt thereof. As described in the Examples, the inventor has further found that administration of a compound of Formula (II) (BT2) reduces left ventricular remodelling, reduces loss of heart function, reduces cardiac fibrosis and reduces myocardial infarct formation, in heart tissue following reperfusion of ischaemic tissue in the heart. In one embodiment, the reducing or preventing myocardial ischaemic-perfusion injury reduces or prevents left ventricular remodelling following reperfusion of ischaemic tissue in the heart of a subject. Left ventricular remodelling in a subject administered a compound of Formula (I) or (II), or a pharmaceutically acceptable salt thereof, is reduced relative to the left ventricular remodelling which would have occurred had the subject not been administered the compound of Formula (I) or (II), or a pharmaceutically acceptable salt thereof. In one embodiment, the reducing or preventing myocardial ischaemic-perfusion injury reduces or prevents cardiac fibrosis following reperfusion of ischaemic tissue in the heart of a subject. Cardiac fibrosis in a subject administered a compound of Formula (I) or (II), or a pharmaceutically acceptable salt thereof, is reduced relative to the cardiac fibrosis which would have occurred had the subject not been administered the compound of Formula (I) or (II), or a pharmaceutically acceptable salt thereof. In one embodiment, the reducing or preventing myocardial ischaemic-perfusion injury reduces or prevents loss of heart function following reperfusion of ischaemic tissue in the heart of a subject. Loss of heart function in a subject administered a compound of Formula (I) or (II), or a pharmaceutically acceptable salt thereof, is reduced relative to the loss of heart function which would have occurred had the subject not been administered the compound of Formula (I) or (II), or a pharmaceutically acceptable salt thereof. In one embodiment, the reducing or preventing myocardial ischaemic-perfusion injury reduces the risk of a subject developing heart failure following reperfusion of ischaemic tissue in the heart of the subject. The risk of heart failure in a subject administered a compound of Formula (I) or (II), or a pharmaceutically acceptable salt thereof, is reduced relative to the risk of heart failure had the subject not been administered the compound of Formula (I) or (II), or a pharmaceutically acceptable salt thereof. In one embodiment, the reducing or preventing myocardial ischaemic-perfusion injury reduces or prevents heart failure following reperfusion of ischaemic tissue in the heart of the subject. Heart failure in a subject administered a compound of Formula (I) or (II), or a pharmaceutically acceptable salt thereof, is reduced relative to heart failure which would have occurred had the subject not been administered the compound of Formula (I) or (II), or a pharmaceutically acceptable salt thereof. In one embodiment, the reducing or preventing myocardial ischaemic-perfusion injury reduces or prevents myocardial infarct formation following reperfusion of ischaemic tissue in the heart of the subject. Myocardial infarct formation following reperfusion in a subject administered a compound of Formula (I) or (II), or a pharmaceutically acceptable salt thereof, is reduced relative to the myocardial infarct formation which would have occurred following reperfusion had the subject not been administered the compound of Formula (I) or (II), or a pharmaceutically acceptable salt thereof. As used herein, “ischaemic tissue” refers to tissue to which blood supply has been reduced or restricted such that demand for oxygen by the tissue exceeds supply. Typically, ischaemic tissue is caused by an ischaemic event. As used herein, an “ischaemic event” is a circumstance in which blood flow to a tissue is restricted or reduced such that the tissue becomes ischaemic. For example, in the heart, ischaemic tissue typically occurs through as ischaemic event in which an artery which supplies blood to a portion of the myocardium is blocked or occluded. For example, during an acute myocardial infarction (MI), occlusion of a coronary artery will prevent supply of blood, and therefore prevent supply of oxygen, to that portion of the myocardium that was perfused with blood from the occluded coronary artery before the occlusion occurred. In myocardial tissue, prolonged ischaemia results in irreversible myocardial cell damage and death. As used herein, “reperfusion” refers to the restoration of blood flow to tissue after blood flow to that tissue has been restricted or reduced. The restoration of blood flow to the tissue restores oxygen supply to the tissue. Typically, the tissue is heart tissue. Heart tissue is also referred to herein as myocardium or myocardial tissue. The reperfusion may be by any means know in the art. For example, the reperfusion may be by thrombolysis or through PCI. Reperfusion by thrombolysis and PCI are known in the art as standard methods of treatment for acute coronary syndromes such as acute myocardial infarction. Ischaemic tissue in the heart of a subject is ischaemic myocardium. The ischaemic tissue may include the infarct (IF) and the area at risk (AAR). The infarct typically comprises dead myocardial cells and irreversibly damaged myocardial cells. As used herein, the AAR is the tissue (myocardial tissue), that was perfused with blood prior to the ischaemic event, and is at risk of further damage from reperfusion. Typically, the AAR was ischaemic tissue during the ischaemic event. The AAR typically comprises undamaged cells and cells that may have been reversibly damaged by the ischaemic event. The cells in the AAR are cells which are at risk of irreversible damage from M / IR injury. As used herein, reperfusion injury refers to cellular dysfunction and cell death resulting from reperfusion of ischaemic tissue. Reperfusion injury in the heart includes

[0002] cardiomyocyte dysfunction and death. Myocardial ischaemia-reperfusion (M / IR) injury refers to reperfusion injury of myocardial tissue. In one embodiment, the compound of formula (I) or (II), or a pharmaceutically acceptable salt thereof, is administered prior to reperfusion. In one embodiment, the compound of formula (I) or (II), or a pharmaceutically acceptable salt thereof, is administered after an ischaemic event but prior to reperfusion. In one embodiment, the compound of formula (I) or (II), or a pharmaceutically acceptable salt thereof, is administered prior to an ischaemic event. In one embodiment, the compound of formula (I) or (II), or a pharmaceutically acceptable salt thereof, is administered during an ischaemic event. In one embodiment, the compound of formula (I) or (II), or a pharmaceutically acceptable salt thereof, is administered prior to an ischaemic event, during an ischaemic event and / or after an ischaemic event but prior to reperfusion. In one embodiment, the compound of formula (I) or (II), or a pharmaceutically acceptable salt thereof, is administered during reperfusion. In one embodiment, the compound of formula (I) or (II), or a pharmaceutically acceptable salt thereof, is administered prior to and / or during reperfusion. In one embodiment, the compound of formula (I) or (II), or a pharmaceutically acceptable salt thereof, is administered after reperfusion. In one embodiment, the compound of formula (I) or (II), or a pharmaceutically acceptable salt thereof, is administered during and after reperfusion. In one embodiment, the compound of formula (I) or (II), or a pharmaceutically acceptable salt thereof, is administered prior to, during and / or after reperfusion. The compound of formula (I) and the compound of formula (II) may be administered as a single dose, or as multiple doses. In one embodiment, the subject has suffered a myocardial infarction. In one embodiment, the myocardial infarction is ST- elevation myocardial infarction (STEMI). In one embodiment, the myocardial ischaemic-perfusion injury is caused by reperfusion resulting from primary cutaneous intervention (PCI). In one embodiment, the myocardial ischaemic-perfusion injury is caused by reperfusion resulting from thrombolysis. In one embodiment, the myocardial ischaemic-perfusion injury is caused by reperfusion resulting from a coronary bypass. As described in the Examples, administration of BT2 prior to reperfusion results in a decrease in expression of genes predictive of heart failure. In this regard, BT2 reduced expression of NLRP3, TRPM2, FOSL2, IL1ß, CXCL1, CXCL2, CCL24, TLR6, IL1R2, HBEGF, FOS and FOSL2. Accordingly, in various embodiments, expression of one or more, 2 or more, 3 or more, 4 or more, 5 or more, 6 or more, 7 or more, 8 or more, 9 or more, 10 or more, 11 or more genes selected from NLRP3, TRPM2, FOSL2, IL1ß, CXCL1, CXCL2, CCL24, TLR6, IL1R2, HBEGF, FOS and FOSL2 is decreased in myocardium of the subject. In one embodiment, expression of the genes NLRP3, TRPM2, FOSL2, IL1ß, CXCL1, CXCL2, CCL24, TLR6, IL1R2, HBEGF, FOS and FOSL2 is decreased in myocardium of the subject. In various embodiments, the expression of one or more, 2 or more, 3 or more, 4 or more, 5 or more, 6 or more¸7 or more¸ 8 or more, 9 or more, 10 or more, 11 or more, 12 or more, 13 or more, 14 or more, 15 or more, 16 or more, 17 or more, 18 or more, 19 or more, 20 or more¸ 21 or more¸ 22 or more¸ 23 or more¸ 24 or more, 25 or more¸ 26 or more¸ 27 or more¸ 28 or more¸ 29 or more, or 30 or more, genes selected from the group consisting of ATF3, FOS, FOSL1, FOSL2, MMP10, MMP16, IL6, IL6R, IL17RA, IL18, TNFSF18, TNFRSF12A, CD68, HSPB1, TLR1, TLR2, TLR6, TLR7, NLRP3, TREM1, TREM3, TRPM2, CXCL1, CXCL6, CCL2, CCL6, CCL17, CCL22, CCL24, NPPA and NPPB is decreased in myocardium of the subject. In one embodiment, expression of the genes ATF3, FOS, FOSL1, FOSL2, MMP10, MMP16, IL6, IL6R, IL17RA, IL18, TNFSF18, TNFRSF12A, CD68, HSPB1, TLR1, TLR2, TLR6, TLR7, NLRP3, TREM1, TREM3, TRPM2, CXCL1, CXCL6, CCL2, CCL6, CCL17, CCL22, CCL24, NPPA and NPPB is decreased in myocardium of the subject. In various embodiments, expression of one or more, 2 or more, 3 or more, or 4 or more, genes selected from the group consisting of FOXP3, TRIM50, CTLA2a, TCAM1 and SOD3 is increased in myocardium of the subject. In one embodiment, expression of the genes FOXP3, TRIM50, CTLA2a, TCAM1 and SOD3 is increased in myocardium of the subject. Another aspect provides a method of treating a subject suffering from an ischaemic event, comprising: (a) administering a compound of Formula (I), typically a compound of Formula (II), or a pharmaceutically acceptable salt thereof; and (b) restoring blood flow to ischaemic tissue. It will be appreciated that an ischaemic event results in ischaemic tissue. In one embodiment, the ischaemic tissue is myocardium. In one embodiment, the ischaemic event is acute coronary syndrome, such as a myocardial infarction. In one embodiment, the myocardial infarction is an acute myocardial infarction. In one embodiment, the myocardial infarction is ST-elevation myocardial infarction (STEMI). Blood flow may be restored to the ischaemic tissue by any methods known in the art. In one embodiment, the blood flow is restored to the ischaemic tissue by administration of one or more thrombolytic agents. Examples of thrombolytic agents include streptokinase, urokinase, tissue plasminogen activator, reteplase and tenecteplase. In one embodiment, blood flow is restored to the ischaemic tissue by percutaneous intervention. An example of PCI is coronary angioplasty, typically with a stent. In another embodiment, blood flow is restored to the ischaemic tissue by a coronary artery bypass. In one embodiment, the compound of formula (I) or (II), or a pharmaceutically acceptable salt thereof, is administered prior to restoring blood flow to ischaemic tissue. In one embodiment, the compound of formula (I) or (II), or a pharmaceutically acceptable salt thereof, is administered after the ischaemic event but prior to restoring blood flow to ischaemic tissue. In one embodiment, the compound of formula (I) or (II), or a pharmaceutically acceptable salt thereof, is administered prior to the ischaemic event. In one embodiment, the compound of formula (I) or (II), or a pharmaceutically acceptable salt thereof, is administered during the ischaemic event. In one embodiment, the compound of formula (I) or (II), or a pharmaceutically acceptable salt thereof, is administered prior to the ischaemic event, during the ischaemic event and / or after the ischaemic event but prior to restoring blood flow to ischaemic tissue. In one embodiment, the compound of formula (I) or (II), or a pharmaceutically acceptable salt thereof, is administered during restoring blood flow to ischaemic tissue. In one embodiment, the compound of formula (I) or (II), or a pharmaceutically acceptable salt thereof, is administered prior to and / or during restoring blood flow to ischaemic tissue. In one embodiment, the compound of formula (I) or (II), or a pharmaceutically acceptable salt thereof, is administered after restoring blood flow to ischaemic tissue. In one embodiment, the compound of formula (I) or (II), or a pharmaceutically acceptable salt thereof, is administered during and after restoring blood flow to ischaemic tissue. In one embodiment, the compound of formula (I) or (II), or a pharmaceutically acceptable salt thereof, is administered prior to, during and / or after restoring blood flow to ischaemic tissue. wherein: R1is straight or branched C1-C6alkyl; and R2is straight or branched C1-C6alkyl, or R2is wherein q is 1, 2, 3 or 4; and R3is straight or branched C1-C6alkyl. In some embodiments of formula (I), R1is straight C1-C6alkyl or branched C1-C6alkyl. In some embodiments of formula (I), R1is -CH2CH3or -CH2CH(CH3)2. In some embodiments of formula (I), R2is straight C1-C6alkyl or branched C1-C6alkyl. In some embodiments of formula (I), R2is -CH2CH3or -CH2CH(CH3)2. R3In some embodiments of formula (I), R2isq O, wherein q is 1, 2, 3 or 4; and R3is straight C1-C6alkyl or branched C1-C6alkyl. In some embodiments of formula (I), q is 2. In some embodiments of formula (I), R3is -CH3. In some embodiments of formula (I), q is 2 and R3is -CH3. In some embodiments, the compound of formula (I) is a compound of formula (I-1): (I-1) wherein: R2is straight or branched C1-C6alkyl; or R2is: R3qO wherein q is 1, 2, 3 or 4; and R3is straight or branched C1-C6alkyl. In one embodiment, the compound of formula (I) is the compound of formula (II). The compound of formula (II) is: . (also referred to herein as BT2) In one embodiment, the compound is a pharmaceutically acceptable salt of formula (I) or (II). Examples of pharmaceutically acceptable salts include salts of pharmaceutically acceptable cations such as sodium, potassium, lithium, calcium, magnesium, ammonium and alkylammonium; acid addition salts of pharmaceutically acceptable inorganic acids such as hydrochloric, orthophosphoric, sulphuric, phosphoric, nitric, carbonic, boric, sulfamic and hydrobromic acids; or salts of pharmaceutically acceptable organic acids such as acetic, propionic, butyric, tartaric, maleic, hydroxymaleic, fumaric, citric, lactic, mucic, gluconic, benzoic, succinic, oxalic, phenylacetic, trihaloacetic (e.g. trifluoroacetic), methanesulphonic, trihalomethanesulphonic, toluenesulphonic, benzenesulphonic, salicylic, sulphanilic, aspartic, glutamic, edetic, stearic, palmitic, oleic, lauric, pantothenic, tannic, ascorbic and valeric acids. In one embodiment, the compound of Formula (I) or (II), or a pharmaceutically acceptable salt thereof, is deuterated. In one embodiment, the compound of Formula (I) or (II), or a pharmaceutically acceptable salt thereof, is an E isomer. In one embodiment, the compound of formula (I) or (II), or a pharmaceutically acceptable salt thereof, is a Z isomer. In one embodiment, the compound of formula (I) or (II), or a pharmaceutically acceptable salt thereof, is a mixture of an E isomer and a Z isomer. The compound of formula (I) or (II), or a pharmaceutically acceptable salt thereof, is typically administered as a pharmaceutical composition. The pharmaceutical composition typically comprises a pharmaceutically acceptable carrier. The methods described herein typically involve the administration of a pharmaceutical composition comprising a compound of formula (I) or (II) or a pharmaceutically acceptable salt thereof, and a pharmaceutically acceptable carrier. Described herein is a pharmaceutical composition comprising a compound of formula (I) or (II), or a pharmaceutically acceptable salt thereof, and a pharmaceutically acceptable carrier. In some embodiments, the carrier is a non-naturally occurring carrier. In some embodiments, the compounds described herein or a pharmaceutically acceptable salt thereof may be used in combination with one or more other agents. It will be understood that the combined administration of a compound described herein or a pharmaceutically acceptable salt thereof with the one or more other agents may be concurrent, sequential or separate administration. The term “composition” encompasses formulations comprising the active ingredient with conventional carriers and excipients, and also formulations with encapsulating materials as a carrier to provide a capsule in which the active ingredient (with or without other carriers) is surrounded by the encapsulation carrier. In pharmaceutical compositions, the carrier is “pharmaceutically acceptable” meaning that it is compatible with the other ingredients of the composition and is not deleterious to a subject. The pharmaceutical compositions described herein may contain other agents or further active agents as described above, and may be formulated, for example, by employing conventional solid or liquid vehicles or diluents, as well as pharmaceutical additives of a type appropriate to the mode of desired administration (for example, excipients, binders, preservatives, stabilizers, flavours, etc.) according to techniques such as those known in the art of pharmaceutical formulation (see, for example, Remington: The Science and Practice of Pharmacy, 21st Ed., 2005, Lippincott Williams & Wilkins). The pharmaceutical composition may be suitable for parenteral (including intramuscular, sub-cutaneous and intravenous), oral, rectal, nasal, topical (including dermal, buccal and sub-lingual), or vaginal administration or in a form suitable for administration by inhalation or insufflation. In some embodiments, the compounds described herein may be formulated for administration in, for example, nanoparticles or liposomes, or polymer formulations. Methods for the production of formulations comprising liposome, lipid nanoparticles and polymeric formulations are known in the art and described in, for example, Lu H. et al., Front Nutr. 8, 783831 (2021); Tenchov, R. et al. ACS Nano 15, 11, 16982–17015 (2021). Liposomes, nanoparticles or polymer formulations may comprise cationic lipids such as DOTAP, DOPE, DC-Chol / DOPE, DOTMA, and DOTMA / DOPE, polymers such as hydroxypropyl methylcellulose (HPMC), polyethylene glycol (PEG), poly(lactic acid-co-glycolic acid (PLGA), poly(lactic acid) (PLA), poly(glycolic acid) (PGA) or self-emulsifying drug delivery systems (AboulFotouh, K. et al.. Colloids Surf. B Biointerfaces 167, 82-92 (2018); Savla, R. et al. Drug Dev. Ind. Pharm.43, 1743-58 (2017)) typically comprising glyceryl monocaprylate, PEG monocaprylate, polyoxyl 35 castor oil, long chain triglyceride (corn oil), glyceryl monolinoleate, polyoxyl 35 castor oil and ethanol. The compounds described herein or a pharmaceutically acceptable salt thereof, together with a pharmaceutically acceptable carrier, may thus be placed into the form of pharmaceutical compositions and unit dosages thereof. The pharmaceutical composition may in the form of sterile injectable solutions for parenteral (including intramuscular, sub- cutaneous, intraperitoneal and intravenous) use The pharmaceutical composition may be a solid, such as a tablet or filled capsule, or a liquid such as solution, suspension, emulsion, elixir, or capsule filled with the same, for oral administration. The pharmaceutical composition may also be in the form of suppositories for rectal administration. Such pharmaceutical compositions and unit dosage forms thereof may comprise conventional ingredients in conventional proportions, with or without additional active compounds or principles, and such unit dosage forms may contain any suitable effective amount of the active ingredient commensurate with the intended daily dosage range to be employed. For preparing pharmaceutical compositions from the compounds described herein, pharmaceutically acceptable carriers can be either solid or liquid. Solid form preparations include powders, tablets, pills, capsules, cachets, lozenges (solid or chewable), suppositories, and dispensable granules. A solid carrier can be one or more substances which may also act as diluents, flavouring agents, solubilizers, lubricants, suspending agents, binders, preservatives, tablet disintegrating agents, or an encapsulating material. Suitable carriers are magnesium carbonate, magnesium stearate, talc, sugar, lactose, pectin, dextrin, starch, gelatin, tragacanth, methylcellulose, sodium carboxymethylcellulose, a low melting wax, cocoa butter, and the like. Tablets, powders, capsules, pills, cachets, and lozenges can be used as solid forms suitable for oral administration. Liquid form preparations include solutions, suspensions, and emulsions, for example, water or water-propylene glycol solutions. For example, parenteral injection liquid preparations can be formulated as solutions in aqueous polyethylene glycol solution. Sterile liquid form compositions include sterile solutions, suspensions, emulsions, syrups and elixirs. The active ingredient can be dissolved or suspended in a pharmaceutically acceptable carrier, such as sterile water, sterile organic solvent or a mixture of both. The pharmaceutical compositions according to the present invention may thus be formulated for parenteral administration (e. g. by injection, for example bolus injection or continuous infusion) and may be presented in unit dose form in ampoules, pre-filled syringes, small volume infusion or in multi-dose containers with an added preservative. The pharmaceutical compositions may take such forms as suspensions, solutions, or emulsions in oily or aqueous vehicles, and may contain formulation agents such as suspending, stabilising and / or dispersing agents. Alternatively, the active ingredient may be in powder form, obtained by aseptic isolation of sterile solid or by lyophilisation from solution, for constitution with a suitable vehicle, e.g. sterile, pyrogen-free water, before use. Pharmaceutical forms suitable for injectable use include sterile injectable solutions or dispersions, and sterile powders for the extemporaneous preparation of sterile injectable solutions. They should be stable under the conditions of manufacture and storage and may be preserved against oxidation and the contaminating action of microorganisms such as bacteria or fungi. The solvent or dispersion medium for the injectable solution or dispersion may contain any of the conventional solvent or carrier systems for injectable solutions or dispersions, and may contain, for example, water, ethanol, polyol (for example, glycerol, propylene glycol and liquid polyethylene glycol, and the like), suitable mixtures thereof, and vegetable oils. Pharmaceutical forms suitable for injectable use may be delivered by any appropriate route including intravenous, intramuscular, intracerebral, intrathecal, epidural injection or infusion. Sterile injectable solutions are prepared by incorporating the active ingredient in the required amount in the appropriate solvent with various other ingredients such as those enumerated above, as required, followed by filtered sterilization. Generally, dispersions are prepared by incorporating the various sterilised active ingredient into a sterile vehicle which contains the basic dispersion medium and the required other ingredients from those enumerated above. In the case of sterile powders for the preparation of sterile injectable solutions, preferred methods of preparation are vacuum drying or freeze-drying of a previously sterile-filtered solution of the active ingredient plus any additional desired ingredients. The formulation may also be sterilized by heat treatment (e.g. boiled) or autoclave. The compounds described herein may be formulated into compositions suitable for oral administration, for example, with an assimilable edible carrier, or enclosed in hard or soft shell gelatin capsule, or compressed into tablets, or incorporated directly with the food of the diet. For oral therapeutic administration, the active compound may be incorporated with excipients and used in the form of ingestible tablets, buccal tablets, troches, capsules, elixirs, suspensions, syrups, wafers, and the like. The amount of active compound in therapeutically useful compositions should be sufficient that a suitable dosage will be obtained. The tablets, troches, pills, capsules, lozenges, implants and the like may also contain the components as listed hereafter: a binder such as gum, acacia, corn starch or gelatin; excipients such as dicalcium phosphate; a disintegrating agent such as corn starch, potato starch, alginic acid and the like; a lubricant such as magnesium stearate; and a sweetening agent such a sucrose, lactose or saccharin may be added or a flavouring agent such as peppermint, oil of wintergreen, or cherry flavouring. When the dosage unit form is a capsule, it may contain, in addition to materials of the above type, a liquid carrier. Various other materials may be present as coatings or to otherwise modify the physical form of the dosage unit. For instance, tablets, pills, or capsules may be coated with shellac, sugar or both. A syrup or elixir may contain the active compound, sucrose as a sweetening agent, methyl and propylparabens as preservatives, a dye and flavouring such as cherry or orange flavour. Of course, any material used in preparing any dosage unit form should be pharmaceutically pure and substantially non-toxic in the amounts employed. In addition, the active ingredient(s) may be incorporated into sustained-release preparations and formulations, including those that allow specific delivery of the active ingredient to specific regions of the gut. Aqueous solutions suitable for oral use can be prepared by dissolving the active component in water and adding suitable colorants, flavours, stabilising and thickening agents, as desired. Aqueous suspensions suitable for oral use can be made by dispersing the finely divided active component in water with viscous material, such as natural or synthetic gums, resins, methylcellulose, sodium carboxymethylcellulose, or other well-known suspending agents. Pharmaceutically acceptable carriers include any and all pharmaceutically acceptable solvents, dispersion media, coatings, antibacterial and antifungal agents, isotonic and absorption delaying agents and the like. Also included are solid form preparations that are intended to be converted, shortly before use, to liquid form preparations for oral administration. Such liquid forms include solutions, suspensions, and emulsions. These preparations may contain, in addition to the active component, colorants, flavours, stabilisers, buffers, artificial and natural sweeteners, dispersants, thickeners, solubilising agents, and the like. For topical administration, the compounds described herein may be formulated as an aqueous or oily base with the addition of suitable thickening and / or gelling agents. Lotions may be formulated with an aqueous or oily base and will in general also contain one or more emulsifying agents, stabilising agents, dispersing agents, suspending agents, thickening agents, or colouring agents. Formulations suitable for topical administration in the mouth include lozenges comprising active agent in a flavoured base, usually sucrose and acacia or tragacanth; pastilles comprising the active ingredient in an inert base such as gelatin and glycerin or sucrose and acacia; and mouthwashes comprising the active ingredient in a suitable liquid carrier. Solutions or suspensions for nasal administration may be applied directly to the nasal cavity by conventional means, for example with a dropper, pipette or spray. The formulations may be provided in single or multidose form. In the case of a dropper or pipette, this may be achieved by the patient administering an appropriate, predetermined volume of the solution or suspension. In the case of a spray, this may be achieved for example by means of a metering atomising spray pump. To improve nasal delivery and retention the compounds of the invention may be encapsulated with cyclodextrins, or formulated with other agents expected to enhance delivery and retention in the nasal mucosa. Administration to the respiratory tract may also be achieved by means of an aerosol formulation in which the active ingredient is provided in a pressurised pack with a suitable propellant such as a chlorofluorocarbon (CFC) for example dichlorodifluoromethane, trichlorofluoromethane, or dichlorotetrafluoroethane, carbon dioxide, or other suitable gas. The aerosol may conveniently also contain a surfactant such as lecithin. The dose of the active ingredient may be controlled by provision of a metered valve. Alternatively the active ingredients may be provided in the form of a dry powder, for example a powder mix of the compound in a suitable powder base such as lactose, starch, starch derivatives such as hydroxypropylmethyl cellulose and polyvinylpyrrolidone (PVP). Conveniently the powder carrier will form a gel in the nasal cavity. The powder composition may be presented in unit dose form for example in capsules or cartridges of, e.g. gelatin, or blister packs from which the powder may be administered by means of an inhaler. In formulations intended for administration to the respiratory tract, including intranasal formulations, the active ingredient will generally have a small particle size for example of the order of 5 to 10 microns or less. Such a particle size may be obtained by means known in the art, for example by micronization. When desired, formulations adapted to give sustained release of the active ingredient may be employed. The pharmaceutical preparations are preferably in unit dosage forms. In such form, the preparation is subdivided into unit doses containing appropriate quantities of the active component. The unit dosage form can be a packaged preparation, the package containing discrete quantities of preparation, such as packeted tablets, capsules, and powders in vials or ampoules. Also, the unit dosage form can be a capsule, tablet, cachet, or lozenge itself, or it can be the appropriate number of any of these in packaged form. It is especially advantageous to formulate parenteral compositions in dosage unit form for ease of administration and uniformity of dosage. Parental compositions may be in the form of physically discrete units suited as unitary dosages for the subjects to be treated, each unit containing a predetermined quantity of the active ingredient calculated to produce the desired therapeutic effect in association a pharmaceutical carrier. The compounds may also be administered in the absence of carrier where the compounds are in unit dosage form. It will be understood that the specific dose level and frequency of dosage for any particular subject may be varied and will depend upon a variety of factors including the activity of the specific compound employed, the metabolic stability and length of action of that compound, the age, body weight, general health, sex and diet of the subject, the mode and time of administration, rate of excretion, drug combinations, and the severity of the particular condition. Suitable dosages of the compounds described herein or further active agents administered in combination with compounds described herein can be readily determined by a person skilled in the art having regard to the particular compound of the invention or further active agent selected. It will further be understood that when the compounds described herein are to be administered in combination with one or more agents, or other active agents, the dosage forms and levels may be formulated for either concurrent, sequential or separate administration or a combination thereof. A further aspect provides a kit for treating myocardial ischaemic perfusion injury, the kit comprising a compound of formula (I) or (II), or a pharmaceutically acceptable salt thereof, or a pharmaceutical composition comprising the compound of Formula (I) or (II), or a pharmaceutically acceptable salt thereof. The compound or composition may be contained in a suitable container, such as a sealable vial or a syringe. The kit may further comprise instructions for use. The methods of the present invention are intended for use with any subject that may experience the benefits of the methods of the invention. Thus, the term “subject” includes humans as well as non-human mammals. The subject may, for example, be a domestic animal, zoo animal or livestock. In one embodiment, the subject is a human. In one embodiment, the subject is a non-human mammal. Unless otherwise herein defined, the following terms will be understood to have the general meanings which follow. The terms referred to below have the general meanings which follow when the term is used alone and when the term is used in combination with other terms, unless otherwise indicated. Hence, for example, the definition of “alkyl” applies to “alkyl” as well as the “alkyl” portions of “haloalkyl”, “heteroalkyl”, “arylalkyl” etc. The term “alkyl” refers to a straight chain or branched chain saturated hydrocarbyl group. Unless indicated otherwise, preferred are C1-6alkyl and C1-4alkyl groups. The term “Cx-yalkyl”, where x and y are integers, refers to an alkyl group having x to y carbon atoms. For example, the term “C1-6alkyl” refers to an alkyl group having 1 to 6 carbon atoms. Examples of C1-6alkyl include methyl (Me), ethyl (Et), propyl (Pr), isopropyl (i-Pr), butyl (Bu), isobutyl (i- Bu), sec-butyl (s-Bu), tert-butyl (t-Bu), pentyl, neopentyl, hexyl and the like. Unless the context requires otherwise, the term “alkyl” also encompasses alkyl groups containing one less hydrogen atom such that the group is attached via two positions, i.e., divalent. As used herein, “treating” means affecting a subject, tissue or cell to obtain a desired pharmacological and / or physiological effect and includes inhibiting the condition, i.e., arresting its development; or relieving or ameliorating the effects of the condition i.e., cause reversal or regression of the effects of the condition. As used herein, “preventing” means preventing a condition from occurring in a cell or subject that may be at risk of having the condition, but does not necessarily mean that condition will not eventually develop, or that a subject will not eventually develop a condition. Preventing includes delaying the onset of a condition in a cell or subject. The term "effective amount" refers to the amount of the compound that will elicit the biological or medical response of a tissue, system, animal or human that is being sought by the researcher, veterinarian, medical doctor or other clinician. An effective amount of a compound described herein, or a pharmaceutically acceptable salt thereof, can be determined by a person skilled in the art having regard to the particular compound. As used herein, the word “comprise” or variations such as “comprises” or “comprising” is used in an inclusive sense, i.e. to specify the presence of the stated features but not to preclude the presence or addition of further features in various embodiments of the invention. The compounds described herein may be synthesised by methods known in the art. The compounds referred to herein as BT2 are commercially available. For example, BT2 can be purchased from Aurora Building Blocks, USA, or Life Chemicals HTS Compounds, Canada. All publications mentioned in this specification are herein incorporated by reference. It will be appreciated by persons skilled in the art 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. The present embodiments are, therefore, to be considered in all respects as illustrative and not restrictive. The present application claims priority from Australian provisional patent application no.2024900164, the entirety of which is incorporated herein by reference. The present invention is further described below by reference to the following non-limiting Examples. EXAMPLES The following experiments show the broad cardioprotective effects of BT2, a dibenzoxazepinone-based small molecule MEK / ERK inhibitor (Li, Y., et al. Science Advances 6, eaaz7815 (2020)) (Table 1), through its prevention of a range of anatomical and functional measures including M / IR injury, LV remodelling, loss of heart function, and cardiac fibrosis. Evidence from RNA-seq and bioinformatics analyses reveals that BT2 reduces transcription factor, cytokine, metalloproteinase inflammasome, DAMP, DAMP-sensing receptor and HF biomarker expression in the heart. METHODS AND MATERIALS Compounds BT2 and derivatives, and BT3, are shown in Table 1. Table 1. Compounds Comp ound Chemical name Chemical structure name (10-ethyl-11-oxo-10,11- O dihydro- H O N N BT2 dibenzo[b,f][1,4]oxazepin-2- O yl)-carbamic acid ethyl ester O (CAS 922029-50-3) 2-amino-10- O ethyldibenzo[b,f][1,4] H N N BT3 oxazepin-11 (10H)-one O (CAS 23474-61-5) N-(10-ethyl-11-oxo-10,11- O H O N BT2- dihydrodibenzo[b,f][1,4]oxazep N MeOA in-2-yl)-2-methoxyacetamide O O (CAS 922029-59-2) ethyl (10-(2-methoxyethyl)-11- O O BT2- H oxo-10,11- O N N EOMe dihydrodibenzo[b,f][1,4]oxazep O O in-2-yl)carbamate ethyl (11-oxo-10-propyl- O BT2- 10,11- H O N N Pr dihydrodibenzo[b,f][1,4]oxazep O in-2-yl)carbamate O (CAS 922029-50-3) O isobutyl (10-ethyl-11-oxo- H BT2- O N N 10,11- IC O dihydrodibenzo[b,f][1,4]oxazep O in-2-yl)carbamate O O BT2- ethyl (11-(oxetan-3- H O N N IMO ylmethoxy)dibenzo[b,f][1,4]oxa zepin-2-yl)carbamate O O ethyl (10-(oxetan-3-ylmethyl)- O BT2-HO NO11-oxo-10,11- N MO dihydrodibenzo[b,f][1,4]oxazep O O in-2-yl)carbamate O ethyl (10-ethyl(2',2',2'-d3)-11- H CD3O N BT2- oxo-10,11- N deut dihydrodibenzo[b,f][1,4]oxazep O O in-2-yl)carbamate Compound synthesis and purification. BT2 was synthesized and purified (>95%) as previously described in WO 2021 / 184059. BT3 was obtained commercially from AK Scientific Inc. Derivatives of BT2 can also be synthesized as described in WO 2021 / 184059. Experimental M / IR injury. Adult male Sprague-Dawley rats were housed for 7 days before experiments to allow acclimatisation. They consumed a standard chow diet. Males were used since estrogen may protect against M / IR injury in rats. Rats were anesthetized with isoflurane gas. Twenty-four hours before inducing myocardial ischaemia, and / or 5 min after coronary artery ligation, animals received BT2 (100 mg / kg), BT3 (100 mg / kg) (Li, Y., et al. Science Advances 6, eaaz7815 (2020)) or vehicle (saline with 0.5% v / v Tween 80, 0.01% v / v DMSO) (all 400 µl) via intraperitoneal (i.p.) injection. Six-lead ECG (eKuore Veterinary) was performed pre-operation and during ligation. Myocardial ischaemia / reperfusion (M / IR) injury was induced in rats essentially as previously described (Luo, X., et al. Arterioscler Thromb Vasc Biol 29, 1836-1842 (2009)) with minor modification. Briefly, under mechanical ventilation, a left thoracotomy was performed, and the left anterior descending (LAD) artery, was ligated 3 mm from its point of origin (distal to the junction of the pulmonary artery and left atrial appendage) with 6-0 prolene. Ischaemia was confirmed by myocardial blanching and ST segment elevation on ECG. After 30 min, the ligature was untied to allow reperfusion of the ischaemic myocardium, and the chest was closed. Using a rodent ultrasound imaging system (Vevo3100, VisualSonics), left ventricular ejection fraction (EF), fractional shortening (FS), left ventricular internal dimension at end-diastole (LVIDd) and at end-systole (LVIDs) were measured in anesthetized rats at 24 h or 2 weeks after left coronary artery (LCA) ligation. At 24 h after LCA ligation, in anesthetized rats, the femoral artery was exposed. A 0.96 mm diameter catheter was inserted into the femoral artery, and 2 ml of blood was collected into SST tubes for later use. At 4 h or 24 h after LCA ligation, the chest cavity was reopened, and the LCA was re-ligated at the same location to define the area at risk (AAR). For rats with endpoint at 4 h or 24 h after ligation, 2 ml of 2% Evans blue dye was injected directly into the right ventricle. Subsequently, 1 ml of 3 M KCl (which arrests the heart in diastole) was administered by intracardiac injection to stop the heart beating prior to heart removal. Hearts were collected at 4 h and 24 h after ligation, frozen at -30°C for 1 h and cut transversely through the LV into 5 slices (each approximately 2 mm) from the apex to the base. Slices of AAR from the 24 h time point were incubated in 4% TTC for 20 min at 37°C to determine infarct size in the AAR (IF / AAR) and AAR / LV area, which was measured using Image-Pro Plus (Cybernetics, Bethesda, MD, USA). Slices from 4 h and 24 h were stored at - 80°C for further isolation of total RNA. At 2 weeks after LCA ligation, the chest cavity of rodents in this group was reopened, 1 ml of 3 M KCl was administered by intracardiac injection prior to heart removal. Hearts were excised and fixed in 10% neutral buffered formalin for histology. Experiments were conducted with UNSW Animal Care and Ethics Committee approval. Serum troponin I detection. Cardiac troponin I levels were determined by using a Siemens Atellica High-Sensitivity Troponin I Assay on an Atellica Solution IM Analyser (Siemens Healthineers, Tarrytown, NY), a chemiluminescent-based system that has been used to measure troponin I levels in blood from multiple species including pigs and dogs. Human cardiac troponin I antibodies (Siemens) also cross-react with rat troponin I. The capture antibody epitope and detection antibody epitope in the High-Sensitivity Troponin I Assay are >97% conserved (35 / 36 residues) in human and rat cardiac troponin I. Human and rat cardiac troponin I share 93% sequence homology overall. BT2 bioavailability. Serum samples were diluted with acetone and the solution left for 14 h at 4°C. Precipitated protein was pelleted by centrifugation, and the supernatant was diluted 1:4 for MS analysis and compared against a standard curve generated with increasing amounts of BT2. Samples were run on a Thermo Gold C18column (50 x 2.1 mm) with solvent A (H2O:0.1% formic acid) and Solvent B (H2O:CH3CN 20:80, 0.1% formic acid), gradient (T=0, 1% B, T=26 min, 100% B, T=27 min, 100%, T=27.1, 1% B, T=30 min, 1% B at 0.2 ml / min) and column temperature 45oC. Mass spectrometry was performed using a QExactive HF operating in data dependent mode with MS1 scan, m / z 140-800, 3 x106ions, max IT 25 msec, resolution 120,000. top 5 MS2. Analysis of heart rate, PR and QTc interval and ST-T segment elevation. ECG was performed using a veterinary ECG monitor with a 6-lead configuration (eKuore). Parameters were set to 20 mm / mV amplitude and paper speed 25 mm / s. For each rat, 3 measurements of heart rate, PR and QT interval were taken, the mean of QT interval is corrected using Bazett's formula (QTc = QT / √RR). Elevation in ST-T segment from baseline was measured from Lead III. Rodent ultrasound imaging. Anesthetised rats were positioned in supine or slightly left lateral decubitus position to gain optimal access to the heart. Prior to imaging, ultrasound gel was applied to the rat’s epilated chest area to facilitate a good acoustic interface for optimal imaging quality. MX250s transducer was placed on the rat’s chest, taking care to adjust its position and angle for a clear view of the LV short axis, notably showing the two LV papillary muscles. Ultrasound imaging was performed using M-mode imaging and Vevo3100 system (VisualSonics). The M-mode cursor was aligned to the LV, to obtain maximum diameter in the LV short axis. M-mode recording was used to capture a continuous line of motion across the heart once the transducer and imaging parameters were properly set. The system was used to obtain LV EF, FS and LV internal dimension at end-diastole (LVIDd) and at end-systole (LVIDs). Immunohistochemical staining and analysis. Twenty-four hours after left coronary artery (LCA) ligation, hearts were excised and fixed in 10% neutral buffered formalin. The hearts were sectioned into four 3 mm slices from apex to base, processed, and embedded in paraffin. Sections from slice 2 were deparaffinized, rehydrated, and subjected to heat- induced epitope retrieval using citrate buffer (pH 6) for 5 minutes at 110°C. Sections were blocked with Dual Endogenous Enzyme Block (DAKO, S2003) for 10 minutes, followed by 2% skim milk for 20 minutes. Slides were incubated with the primary antibody (rabbit polyclonal anti-Trpm2, cat. no. PA5-119946; Thermo Fisher) at a 1:400 dilution for 60 minutes at room temperature. After rinsing with buffer, slides were incubated with the secondary antibody (goat anti-rabbit, cat. P0448, DAKO) for 30 minutes. Subsequently, diaminobenzidine (DAB) chromogen (cat. K3468, DAKO) was applied for 5 min, and the slides were counterstained with hematoxylin and Scott's blue. After counterstaining, slides were dehydrated in 100% ethanol and xylene and then coverslipped. Immunostained slides were scanned using an Olympus VS200 E26 slide scanner (Olympus, Tokyo, Japan), and images were captured using QuPath software (Bankhead et al., QuPath: Open source software for digital pathology image analysis. Sci Rep 7, 16878 (2017)). The percentage area of positive staining in 20x objective fields was quantified using Image-Pro Plus software (Cybernetics, Bethesda, MD, USA). Quantification was performed with 10 fields of view per section, with 3-4 rats per group. Multiplex immunofluorescence staining and image acquisition. Multiplex immunofluorescence staining and image acquisition was performed by the Kat Gaus Light Microscopy Facility, Mark Wainwright Analytical Centre, UNSW. Primary antibodies used were rabbit monoclonal anti-phospho-ERK (CST, cat. no.4370s), rabbit monoclonal anti- CD68 (CST, cat. no. 97778s), rabbit monoclonal anti-CD4 (CST, cat. no.25229S), rabbit monoclonal anti-Foxp3 (Abcam, cat. no. ab215206), and rabbit monoclonal anti-dystrophin (Abcam, cat. no. ab218198). Multiplex images were captured using HALO software (Indica Labs) and analysed with Image-Pro Plus (Cybernetics, USA). The necrotic zone was defined as the area with negative dystrophin staining or regions showing incomplete myocardial cells based on dystrophin staining. Regions of the border zone were defined as a 500 µm margin from the edge of the necrotic zone. Quantification of positive cells was performed using 4-5 random 20x objective view in border zone per tissue section, with 3-4 rats per group. Sirius Red and methyl green staining. Cross sections of Sirius Red and methyl green stained slides were scanned using an Aperio ScanScope XT slide scanner (Leica Biosystems, Mt Waverley, Vic, Australia) and images were captured using ImageScope software (Leica Biosystems). Red staining area and LV area were determined using Image-Pro Plus (Cybernetics, Bethesda, MD, USA). Hematoxylin and eosin (H&E) staining and neutrophil counts. Sections were deparaffinized, rehydrated, and then immersed in Harris hematoxylin solution for staining. Following differentiation and bluing steps, sections were counterstained with eosin-phloxine solution. Sections then underwent dehydration and mounted using a mounting medium. H&E-stained sections were scanned using an Olympus VS200 E26 scanner (Olympus Life Science) and digital images were captured and analysed using QuPath software (https: / / qupath.github.io / ). Neutrophils, identified as polymorphonuclear leukocytes, were counted in the border zone with displaying haemorrhage. Counting was performed under a 20x objective, using a standardized counting grid field within the software. Ten randomly selected fields per rat (n=3-4 rats) were analyzed to obtain an average neutrophil count per 20x objective field. Single nucleus RNA-seq and bioinformatics analysis. Twenty-four hours after left coronary artery ligation, the chest was reopened, and the LCA religated at the same location to define the AAR. A 2% Evans blue dye (2 ml) was injected into the RV, followed by 1 ml of 3 M KCl to arrest the heart in diastole. Hearts were harvested, frozen at -30°C for 1 h, and sectioned into four 3 mm slices from apex to base on dry ice. The AAR tissue from the left ventricle (LV) was collected and stored at -80°C for subsequent analysis. snRNA-seq data were generated from frozen AAR using the chromium single cell gene expression solution (10x Genomics) by the Garvan Genomics Platform. Briefly, frozen AAR (third slice from apex) was thawed, cut into small pieces, and homogenized in a Dounce homogenizer with 5 ml of hypotonic lysis buffer. The lysate was combined with a rinse of 1 ml buffer, gently pipetted, and centrifuged at 500 g for 5 min at 4°C. The pellet was resuspended in 5 ml washing buffer, filtered through a 40 µm cell strainer, and centrifuged. The nuclear pellet was resuspended in 0.5 ml buffer, and nuclei integrity was assessed using Trypan blue staining under a bright-field microscope. Tissue was pooled from 2 groups of rats (3 vehicle, 3 BT2 treated). Single nuclei were stained with DAPI and sorted on a Becton Dickinson FACS AriaI II, using a 70 µm nozzle, and based on the gating strategy in representative Fig. 19. Nuclei were sorted to 1.5 ml Eppendorf tube precoated and containing 100 µl collection buffer. Sorted nuclei were passed over to the Cellular & Spatial Hub for QC, capture and sequencing. Nuclei were reconcentrated to achieve an approximately 1000 nuclei / µl concentration with 5ul aliquot of nuclei enumerated using AOPI (Logos Biosystems) staining, counting on the Luna-FX7 automated cell counter (ATA Scientific). Nuclei were then pooled at an equal ratio from 3 samples per group. Of this pool, approximately 28500 nuclei were aliquoted for capture. This allows a capture for a targeted 20000 nuclei. Capture for single nuclei was processed through the 10x Genomics 3’ GEM-X assay, using the 10x Genomics Chromium-X instrument. Briefly, the processes include (a) cell coencapsulation with barcode-bearing gel beads in emulsions (GEM generation), (b) reverse- transcription and cDNA amplification, and (c) 3’ gene expression library construction. The 3’ gene expression libraries were sequenced on a 10 B flow cell on a NovaSeq X plus sequencer at the Ramaciotti Centre for Genomics. Raw sequencing data were aligned to the reference genome and gene expression matrices were generated using the Cell Ranger software (10x Genomics). For data preprocessing and quality control, cell matrices were imported and analyzed using the Seurat R package (v5) (Hao et al., Integrated analysis of multimodal single-cell data. Cell 184: 3573-3587). To ensure high-quality data, cells were filtered based on standard quality control metrics: cells with fewer than 200 or more than 5000 detected features (nFeature_RNA) and cells with >5% mitochondrial RNA content were excluded from downstream analyses. Filtered data were normalized using Seurat's NormalizeData function to scale and transform raw expression values for each gene. Highly variable features were identified using the FindVariableFeatures function. Data was then scaled to remove unwanted variation using ScaleData. Principal Component Analysis (PCA) was performed using the top 15 principal components as input for downstream analyses. The PCA embedding was used to construct a shared nearest neighbor (SNN) graph and to identify clusters. Data layers were integrated using the canonical correlation analysis (CCA) method implemented in Seurat. The integration was performed with the IntegrateLayers function, followed by layer joining using JoinLayers to create a unified dataset. Cell clusters were identified using gene lists generated from FindAllMarkers. Differentially expressed genes were identified using the MAST (Model-based Analysis of Single-cell Transcriptomics) framework, which is specifically designed for single- cell RNA-seq data. The zlm function from the MAST package was employed to fit a hurdle model to the single-cell expression data (Finak, G., et al. MAST: a flexible statistical framework for assessing transcriptional changes and characterizing heterogeneity in single- cell RNA sequencing data. Genome Biol 16, 278 (2015)). Gene set enrichment analysis (GSEA, preranked): GSEA was performed using GSEA software (v 4.3.3). For the analysis, unfiltered preranked data, including gene names and log2fold changes (log2FC), were loaded into the software. The gene set databases used were “c5.all.v2024.1.Hs.symbols.gmt” or “c5.go.v2024.1.Hs.symbols.gmt” (for monocyte analysis). The “Collapse / Remap to gene symbols” option was set to “Collapse”, and the chip platform used was “Rat_Gene_Symbol_Remapping_Human_Orthologs_MSigDB.v2024.1.Hs.chip”. All other settings were left as default. The top 10 enriched gene sets, ranked by normalized enrichment score (NES), were visualized using bar plots generated by the SRplot platform (an online tool for data visualization and graphing available at (Tang, D., et al. SRplot: A free online platform for data visualization and graphing. PLoS One 18, e0294236 (2023)). Enhanced volcano diagram and hyperbolic volcano diagram: Differentially expressed genes (DEG) were visualized using enhanced volcano and hyperbolic volcano diagrams. Unfiltered data, including gene names, log2fold changes (log2FC), and false discovery rates (FDR), were uploaded to the SRplot platform (available for visualization. For the hyperbolic volcano diagram, two hyperbolic curves with the function y=1 / x were added as reference lines to the traditional volcano plot, using the original threshold as the baseline. These curves facilitated the classification of genes, making it easier to identify genes with more significant differential expression. GO (BP) pathway enrichment bubble chart: Pathway enrichment results were visualized using bubble charts. The top 20 gene sets, including “Term Name”, “Enrichment” (ratio of intersection size to query size), “P-value”, and “Count” (intersection size), were uploaded to the SRplot platform (available at visualization. GO (BP) pathway enrichment bar plots for neutrophils: Pathway enrichment results for neutrophils were visualized using bar plots generated on the SRplot platform (available at The minimum gene set size (query size) was set to 5 genes for the analysis. Dot plot for comparison of downregulated pathway enrichment across five cell types: The comparison of downregulated pathway enrichment across five cell types was visualized using a dot plot. Selected gene sets for each cell type, including “Cell Type”, “Term Name”, “Gene Count”, and “P-value”, were uploaded to the SRplot platform (available at www.bioinformatics.com.cn) for visualization. Bulk RNA-seq and bioinformatics analysis. Ice-cold heart tissue (AAR) was transferred to labeled Precellys® Lysing CK14 tubes and TRIzol reagent (800 µl) was added. Tubes were placed into the TissueLyser adapter sets and samples were homogenized thrice for 20 s at 6500 rpm, with 3 min intervals between each round of homogenization. Samples were centrifuged for 2 min and supernatants were transferred to new 1.5 ml screw cap tubes then inverted to mix the contents, vortexed and incubated for 15 min at 22oC. Total RNA was isolated homogenized samples using the RNeasy Mini Kit (Qiagen, cat.74004) as follows. Chloroform was added to the mixture prior to microfuge centrifugation at 13000 rpm for 15 min at 4°C. The upper aqueous layer (containing total RNA) was transferred to microtubes, isopropanol was added and loaded into RNeasy columns. Columns were washed with buffers RPE and RW1. Total RNA was eluted with ribonuclease-free water. Samples were submitted to the UNSW Ramaciotti Centre for Genomics for TruSeq Stranded mRNA-seq preparation and sequencing by NextSeq 6000 to produce 75 bp single-end reads. Bioinformatics analysis performed on this sequence data was conducted by the Australian Genome Research Facility and included adapter and quality trimming with Trim Galore, alignment to the reference genome with STAR, and gene count quantification with featureCounts. Finally, DEGs were identified using EdgeR. Raw expression counts were filtered for lowly expressed genes using a minimum counts per million (CPM) threshold of 0.5, followed by normalization according to library size factors using the Trimmed Mean of M-values (TMM) method. Finally, expression data was fit to a negative binomial generalized linear model before undergoing a genewise quasi-likelihood F-test to identify DEGs. Volcano plots from RNA-seq DEG analysis. DEG data was uploaded into the Galaxy platform (https: / / usegalaxy.org.au / ) and converted to volcano plot format, which allows visualization of DEG results in scatter plot format. KEGG enrichment from RNA-seq DEG analysis: Enrichment factors were calculated using the formula: enrichment factor = (DEG in category / total genes in category) / (DEG in background / total genes in background). Enrichment bubble plots and KEGG enrichment were generated using the SRPLOT web platform (https: / / www.bioinformatics.com.cn / en?keywords=bubble). Upload the analysis results (including enrichment factor) onto the web platform. GEO dataset analysis: GEO dataset analysis was performed using the GEO2R interface Data from the GEO2R was exported in xls format and analysed for statistical significance using Graphpad PRISM v9. De-identified, open access human datasets were analysed after obtaining UNSW Human Research Ethics Committee approval. The Australian Genome Research Facility conducted an analysis where the default GEO2R parameters were used to generate two datasets of DEG in GSE59867. Firstly, samples collected from STEMI patients within one day of AMI (all underwent direct PCI) who subsequently developed HF (n=9) were compared to samples from a control group with stable CAD patients (n=46), and secondly, another comparison was made between samples from patients that did not develop HF, collected within one day of AMI (n=8) versus a control group with stable CAD (n=46). Significant DEG were isolated based on the adjusted p-value threshold of 0.05 and further subdivided into a list of upregulated and downregulated DEG based on fold-change for each dataset. The R package VennDiagram was used to detect and visualize the intersection of DEGs between these GEO2R analyzed datasets and the DEGs identified in the EdgeR analysis on BT2-regulated samples. Statistics. Statistical analysis was performed as stated using Graphpad PRISM v9, which does not draw error bars when these are shorter than the height of the symbol. If distribution was not normal, Mann-Whitney or Kruskal-Wallis was performed. Normally distributed data was analyzed by t test or one-way ANOVA. Plotted data represent mean ± SEM. Differences were considered significant when p≤0.05. Where indicated, *p≤0.05, **p<0.01, ***p<0.001, ****p<0.0001. ns denotes not significant. RESULTS BT2 inhibits IF size in the area-at-risk 24h after M / IR injury. A rodent model of M / IR injury was used in which SD rats (~350 g) were subjected to M / IR injury involving 30 min LAD ligation (confirmed by ECG) followed reperfusion (Fig.1). BT2 (100 mg / kg, delivered i.p. 24h before and prior to M / IR) inhibited IF size (relative to vehicle group) in the AAR by ~70% after 24h, despite the similar %AAR / LV between groups (Fig.2A and 2B). In contrast, BT3, an inactive structural analogue of BT2 had no such effect. The AAR / LV percentage did not differ between the groups and served as a technical control (Fig.2C). Circulating levels of cardiac troponin I, a well-established biomarker of MI, confirmed cardiac injury after 24 h and was reduced ≥50% by BT2 compared to vehicle (Fig. 3). Immunohistochemical staining showed that BT2 inhibited levels of phosphorylated ERK (p-ERK) in the AAR 24h after M / IR injury (Fig. 4). BT2 had no effect on PR or QTc interval, indicating no significant effect on conduction (Fig. 5). Mass spectrometry confirmed the serum bioavailability of i.p. delivered BT2, which reached 45.3 µg / ml 24 h after M / IR injury (Fig. 6). BT2 preserves heart function and prevents adverse remodeling. Within 24 h, both EF and FS fell in the vehicle group. In contrast, BT2 prevented this loss of cardiac function (Fig.7). Two weeks after reperfusion, while % EF and % FS remained low (and did not improve relative to % EF and % FS levels at 24 h), BT2 retained its ability to protect against M / IR - induced loss of heart function. Indeed, there was no difference in % EF or % FS at 2 weeks in the BT2 group compared with those of the sham group (i.e., surgery but no M / IR injury) 13 days earlier (Fig.7). This indicates sustained post-reperfusion cardioprotective effects of BT2 for at least 2 weeks. Ventricular remodeling and impaired contractility, like % EF and % FS, are hallmark measures of HF or dilated cardiomyopathy. In line with EF and FS data, ventricular enlargement (i.e., increased LVIDd) was evident in the vehicle group after 2 weeks, which was prevented by BT2. Importantly, there was no difference in LVIDd at 2 weeks in the BT2 group compared with the sham group (Fig. 8), indicating again, sustained cardioprotective effects of BT2. Impaired contractility evident at 24 h in the vehicle group was more profound at 2 weeks. However, this was completely inhibited by BT2 at both 24 h and 2 weeks. Like LVIDd, there was no difference in LVIDs in the BT2 group compared with the sham group at any time (Fig.8). snRNA-seq analysis. To gain insights into the mechanism of action of BT2, nuclei were isolated from the AAR of rats treated with vehicle or BT2, 24 h after M / IR injury, and single nucleus (sn)RNA-seq was performed. This technique, unlike scRNA-seq, can also facilitate the capture and assessment of gene expression in larger-sized cardiomyocytes among a range of cell types (Miranda, A.M.A., et al. Single-cell transcriptomics for the assessment of cardiac disease. Nat Rev Cardiol 20, 289-308 (2023)). To compare with bulk RNA-seq data from adjacent tissue slices, the snRNA-seq was collapsed into a pseudo-bulk dataset. Of 4442 genes in the dataset, there were 3396 significant DEG (FDR<0.05). The number of upregulated (log2FC>0 & FDR<0.05) and downregulated (log2FC<0 & FDR<0.05) DEG were 1766 and 1630, respectively. Inspection of individual DEG in the collapsed snRNA dataset revealed that extracellular matrix components (Lama2, Gpc6, Lamc1), transcription factors (Zeb1, Zfpm2, Ebf1) a cell adhesion molecule (Sdk1), glycoprotein (B3galt1), protein kinase (Prkg1), and interestingly, an effector of epithelial-to-mesenchymal transition (Rbms3) were among the 10 most downregulated DEG. Glypican-6 (Gpc6), in particular, is a known regulator of heart failure progression by controlling cardiomyocyte growth through ERK signaling (Melleby, A.O., et al. PLoS One 11, e0165079 (2016)) while zinc finger E-box binding homeobox 1 (Zeb1) regulates collagen synthesis by cardiac fibroblasts and myocardial fibrosis. Among the 10 most upregulated DEG were immune and inflammatory modulators (Slfn4, Ptprc, Dock2, Spp1, Lilrb4, Dock8), a transcription factor (Zfp710), nucleoside transporter (Slc28a2), phospholipid transporter (Atp8b4) and tumor suppressor (Rbm47). GSEA revealed that 1801 of 4803 gene sets were upregulated (578 gene sets enriched at p<0.25) whereas 3002 of 4803 gene sets were downregulated (1310 gene sets enriched at p<0.25). Among the top 10 upregulated and downregulated gene sets ranked by NES in GSEA analysis, the upregulated sets were associated with cell differentiation and immune cell receptor-mediated signaling, whereas the downregulated sets were related to the basement membrane, collagen, and extracellular matrix. Cell cluster analysis: snRNA-seq data was analyzed first without collapsing, with 18 individual cell types identified based on key gene markers (Arduini, A., et al. Transcriptional profile of the rat cardiovascular system at single cell resolution. bioRxiv (2023)). snRNA-seq revealed large changes in the proportion of certain cell types between treatments in the AAR 24 h after M / IR injury. BT2 increased the proportion of Flt1+Cyyr1+Hmcn1+Pgm5+Npr3+Vwf+endothelial cells (EC2), but not Flt1+Dach1+Cyyr1+endothelial cells (EC1) or Flt1+Dach1+Cyyr1+Hmcn1+Pgm5+Unc5c+Dnm3+Vwf+endothelial cells (EC3a). BT2 also increased the proportion of Gsn+Dcn+Col3a1+Fbn1+fibroblasts (FB1), but not Col3a1+Col1a1+Fbn1+Postn+Adam12+fibroblasts (FB3) or Col3a1+Col1a1+Fbn1+Postn+Diaph3+Adam12+Mki67+myofibroblasts. BT2 did not change the proportion of cardiomyocytes or lymphatic endothelial cells. However, BT2 reduced the proportion of inflammatory cell types including neutrophils, monocytes, macrophages and T cells. The most abundant cell types in the AAR were macrophages and EC1 cells, followed by pericytes, FB1 fibroblasts, EC3a endothelial cells and cardiomyocytes. DEG genes for each cell cluster were identified and enrichment analysis of DEG (FDR<0.05) from each cell cluster revealed that BT2 downregulated cell migration, motility and cell adhesion pathways in neutrophils, monocytes, macrophages, T cells and myofibroblasts. On the other hand, BT2 upregulated pathways associated with autophagy in neutrophils, sodium channel regulation in monocytes, immune system processes in macrophages, receptor signaling and response to endogenous stimuli in FB1 fibroblasts, and response to stress pathways in myofibroblasts. Pathway enrichment analysis in the collapsed snRNA-seq DEG dataset revealed downregulation of a range of cellular metabolic processes, and upregulation of immune processes and cellular signaling. BT2 had the most profound gene suppressive effect on a range of biological processes in myofibroblasts yet had minimal effects on FB1 fibroblasts and even less effect on neutrophils. BT2 downregulated pathway enrichment in monocytes and macrophages, with cell differentiation, intracellular signalling and morphogenesis inhibited more profoundly in macrophages than monocytes. Macrophages, monocytes and neutrophils: Inspection of individual DEG in macrophage cells revealed that LOC310926, Cblb, Lama2 and Prkg1 were among the most downregulated DEG, and that Milr1 and Slfn4 were among the most upregulated DEG. Casitas B lymphoma- b (Cblb) is a key regulator of macrophage activation while mast cell immunoglobulin-like receptor 1 (Milr1), also known as allergin-1, inhibits autoantibody production via upregulation of macrophage phagocytosis. GSEA revealed that 2020 of 4297 gene sets were upregulated (1 gene set enriched at FDR<0.25) whereas 2277 of 4297 gene sets were downregulated (24 gene sets enriched at FDR<0.25). GSEA analysis further showed that among the top 10 gene sets ranked by NES in macrophages were those involved in cholesterol efflux and sterol transport, whereas gene sets that were downregulated included those associated with cell structure and cytoskeleton. Enrichment analysis of DEG (FDR<0.05) showed that BT2 downregulated pathways in macrophages associated with cell differentiation, cell signaling, locomotion and migration. This is in line with lower CD68+macrophage staining (Fig.9A) in the AAR 24 h after M / IR injury in BT2-treated rats. In contrast, BT2 had no significant effect on CD4+T lymphocytes or Tregs (CD4+FoxP3+) (Fig. 9B and 9C). LOC310926, Prkg1, Lama2 and Ebf1 were among the most downregulated DEG in the monocyte dataset, whereas Slfn4, Zfp710 and Vps54 were among the most upregulated DEG. GSEA revealed that 983 of 2210 gene sets were upregulated (27 gene sets enriched at FDR<0.25) whereas 1227 of 2210 gene sets were downregulated (52 gene sets enriched at FDR<0.25). GSEA analysis further revealed that among the top 10 gene sets ranked by NES in monocytes. Upregulated sets were associated with STAT signaling, natural killer cell- mediated immunity and cell killing, while downregulated sets were related to cell cycle transition and collagen-containing extracellular matrix. BT2 downregulated pathways (FDR<0.05) enriched in monocytes were associated with cell motility and migration. In the neutrophil dataset, Serpinb1a, Il1r1 and Nek10 were among the most downregulated DEG whereas Usp32, Zfp710 and Jam1 were among the most upregulated, GSEA identified 1204 of 3012 gene sets upregulated (however, no gene sets were enriched at FDR<0.25) whereas 1808 of 3012 gene sets were downregulated (and again, no gene sets were enriched at FDR<0.25). BT2 enriched pathways associated with autophagy and histone deacetylase recruitment (Mad-Max), whereas gene sets that were downregulated included those associated with cell migration and motility. Myofibroblasts and fibroblasts: Heatmap analysis revealed a distinct proliferation gene signature (Mki67, Ccna2, Tk1, Top2a) that separated the myofibroblasts from the FB1 fibroblasts and FB3 fibroblasts cluster. FB3 fibroblasts, like myofibroblasts, were Postn+and Pdgfra+, but like FB1 fibroblasts, did not express the proliferation gene signature that characterized the myofibroblasts. While BT2, relative to vehicle, had no effect on proliferation marker gene expression in FB1 or FB3 fibroblasts or myofibroblasts, there were approximately 3 fold more FB1 cells in the BT2 group than the vehicle group. In contrast, BT2 did not change the proportion of cells in the FB3 and myofibroblast groups. In myofibroblasts, Fgd5, Prkg1, Plpp3 and Il4r were among the most downregulated DEG, whereas Fam111a, Actn1 and Mfap5 were upregulated. Faciogenital dysplasia 5 (Fgd5)- antisense 1 expression can reduce infarct size, enhance cardiac function and inhibit cardiac fibrosis, and protein kinase G1 (Prkg1) activity is associated with cardiac fibrosis (Schwaerzer, G.K., et al. Br J Pharmacol 179, 2413-2429 (2022)). GSEA revealed that 2457 of 5747 gene sets were upregulated (1 gene set enriched at FDR<0.25) whereas many more (3290 of 5747) gene sets were downregulated (10 gene sets enriched at FDR<0.25). GSEA analysis further revealed that among the top 10 gene sets ranked by NES upregulated in myofibroblasts were those involved in cell structure and replication, whereas gene sets that were downregulated included those associated with cell signaling and extracellular matrix production. Enrichment analysis of DEG (FDR<0.05) showed that BT2 downregulated pathways in myofibroblasts were associated with cellular development, signal transduction, cell motility and migration. In the FB1 fibroblasts Zeb1, Pcdha4 and Tmlhe were among the most downregulated DEG, whereas Aox3, Fhl2 and Zfp710 were among the most upregulated. GSEA revealed that 1789 of 4171 gene sets were upregulated (48 gene sets enriched at FDR<0.25) whereas 2382 of 4171 gene sets were downregulated (0 gene sets enriched at FDR<0.25). GSEA analysis further revealed that among the top 10 gene sets ranked by NES upregulated in FB1 fibroblasts were those involved in cell integrity and structure, whereas gene sets that were downregulated included those associated with chromosomal structure and aging. BT2 upregulated (FDR<0.05) pathways associated with cell surface receptor signaling, cell adhesion and anatomical structure. Bulk RNA-seq analysis. To gain further insights into the mechanism of action of BT2, RNA was isolated from the AAR of rats treated with vehicle or BT2, 4h or 24 h after M / IR injury, and bulk next-generation RNA sequencing (RNA-seq) was performed. Multidimensional scaling (MDS) showed clear separation between treatment groups. Of 14927 genes in the dataset, at 4 h, there were 2540 significantly DEG (FDR<0.05). The number of upregulated (logFC>0 & FDR<0.05) and downregulated (logFC<0 & FDR<0.05) genes was 1032 and 1508, respectively. At 24 h, there were substantially more (6060) DEG (FDR<0.05); the number of upregulated (logFC>0 & FDR<0.05) and downregulated (logFC<0 & FDR<0.05) genes was 2895 and 3165, respectively. BT2 reduces expression of neutrophil and other immune cell markers in the AAR. Neutrophils provide the primary cellular response to M / IR injury and mediate inflammation and severity associated with M / IR injury. BT2 reduced expression of several neutrophil markers in the AAR within 4 h of M / IR injury such as Cd11b / Itgam (logFC -1.85 fold), Cd18 / Itgb2 (logFC -1.97 fold), Cd33 (logFC -1.68 fold), Cd44 (logFC -1.52 fold) and Cd45 / Ptprc (logFC -1.28 fold). Additionally, compared with vehicle 24 h following M / IR, BT2 reduced expression of CD11b / Itgam (logFC -1.84), Cd18 / Itbg2 (logFC -2.02), Cd33 (logFC - 1.48), Cd15 / Fut4 (logFC -1.00), Cd16 / Fcgr3a (logFC -1.60), Cd32 / Fcgr2b (logFC -1.21), Cd44 (logFC -1.77), Cd45 / Ptprc (logFC -1.10) and Cd62L / Selplg (logFC -1.00). At 24h after M / IR injury, BT2 inhibited monocyte (e.g., Cd14, Cd11b / Itgam) and M1 macrophage (e.g., Cd86, Cd83, Cd80) biomarker expression. In line with reduced neutrophil and monocyte / macrophage biomarker expression in the AAR, BT2 inhibited levels of cell surface adhesion receptors expressed by monocytes and / or neutrphiles (e.g., Cxcr2, CD44) and their cognate endothelial ligands (e.g., Cxcl2, Sele). BT2 reduces expression of transcription factors, cytokines, metalloproteinases, inflammasome, DAMPs, DAMP-sensing receptors and HF biomarkers in AAR. Compared with vehicle 4 h after M / IR injury, BT2 reduced expression in the AAR of pro-inflammatory transcription factors (e.g., Egr1, Jun, Junb, Fosl1, Klf5), matrix metalloproteinases (e.g., Mmp3, Mmp8, Mmp25), pro-inflammatory cytokines and their receptors (e.g., Osm, I1b, Il1r2, Tnfsf18, Tnfrsf12a, Tnfrs1b), DAMP biomarkers (e.g., S100a8, S100a9, Hspa1a), DAMP receptors (e.g., Tlr1, Tlr2, Tlr13, Ripk2, Nlrp3, Trem1, Trem3, Trpm2), CXC chemokines (e.g., Cxcl1, Cxcl2), chemokines (e.g., Ccl2, Ccl3, Ccl6, Ccl17, Ccl22, Ccl24), Nppb (B-type natriuretic peptide, Bnp) and growth factors (e.g., Hbegf) (Table 2). Among genes with the largest fold inhibition of expression at 4 h were Mmp3 (logFC -4.67 fold), metallopeptidases Adam12 (logFC -3.05 fold) and Adam8 (logFC -2.74 fold), and chemokines Ccl7 (logFC -4.88 fold) and Ccl22 (logFC -2.84 fold). Conversely, at 4 h, BT2 increased the expression of Sned1 (logFC 2.49 fold), an anti-inflammatory factor, and Tcam1 (logFC 2.91 fold), which mediates Treg suppressive function. Similarly, compared with vehicle 24 h after M / IR injury, BT2 in the AAR reduced the expression of pro-inflammatory transcription factors (e.g., Atf3, Fos, Fosl1, Fosl2), matrix metalloproteinases (e.g., Mmp10, Mmp12, Mmp16), pro-inflammatory cytokines and their receptors (e.g., Il6, Il6r, Il17ra, Il18, Tnfsf18, Tnfrsf12A), macrophage biomarkers (e.g., cd68), DAMP biomarkers (e.g., Hspb1), DAMP receptors (e.g., Tlr1, Tlr2, Tlr6, Tlr7, Nlrp3, Trem1, Trem3, Trpm2), CXC chemokines (e.g., Cxcl1, Cxcl6), chemokines (e.g., Ccl2, Ccl6, Ccl17, Ccl22, Ccl24), Nppa (atrial natriuretic peptide, Anp) and Nppb (Bnp) (Table 3). Among genes with the largest fold inhibition of expression with BT2 compared to vehicle at 24 h were Mmp10 (logFC -5.79 fold), Mmp12 (logFC -3.60 fold), Il6 (logFC -3.70 fold), Ccl12 (logFC -3.14 fold) and Ccl7 (logFC -3.03 fold). At 24 h, BT2 increased the expression of the Treg biomarker Foxp3 (logFC 2.20 fold), the autophagy regulator Trim50 (logFC 2.89 fold), the negative regulator of inflammation Ctla2a (logFC 1.28 fold), Treg suppressor biomarker Tcam1 (logFC 1.51 fold) and antioxidant enzyme SOD3 (logFC 1.92 fold) (Fig. 3C, Table 3). Early phase infiltrating monocytes serve as mediators of M / IR injury. For example, siRNA (targeting CCR2) knockdown of monocytes reduces IF size in rodent models. BT2 strongly reduced mRNA levels of Ccl2 (whose main receptor is CCR2) at 4 h (logFC -2.21) and 24 h (logFC -2.85). BT2 reduced mRNA levels of monocyte markers in the AAR within 24 h of M / IR injury, such as CD62L / Sell at 4 h (logFC -2.31), Cd14 at 24 h (logFC -1.26), Cd16 at 24 h (logFC -1.60), CD115 / Csf1r at 24 h (logFC -0.94) and Cx3cr1 at 24 h (logFC -1.88). Monocytes are the main source of macrophages, BT2 reduced mRNA level of the macrophage marker Cd68 at 4 h (logFC -0.75) and 24 h (logFC -1.71) in the AAR. BT2 also reduced levels of Nlrp3, which in monocytes, mediates and exacerbates the inflammatory response following reperfusion onset, at 4 h (logFC -2.17) and 24 h (logFC -1.45) (Tables 2 & 3). Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis was performed to determine whether DEG following BT2 treatment were over-represented in specific biochemical pathways (p<0.05). Among the top hits in downregulated genes at 4 h were cytokine-cytokine receptor interactions, lipid and atherosclerosis, TNF signaling pathway, chemokine signaling, MAPK signaling, NF-κB signaling, IL17 signaling, Toll-like receptor signaling and leukocyte transendothelial migration. Among the top hits in downregulated genes at 24 h were cell cycle, chemokine signaling, cytokine-cytokine receptor interaction, lipid and atherosclerosis and NF-κB signaling. BT2 reduces cardiac fibrosis 2 weeks after M / IR injury. Cardiac fibrosis is typified by accumulation of extracellular matrix proteins such as collagen in the interstitium, causing cardiac dysfunction. Cross sections of hearts from rats treated with BT2 or vehicle 2 weeks after M / IR were stained with Sirius Red (stains collagen). Despite no difference in ST elevation between Veh and BT2 groups during coronary artery ligation (Fig.10A), BT2 reduced scarring in the LV by ~50% after 2 weeks (Fig.10B). Inhibition of cardiac fibrosis by BT2 after 2 weeks is in line with its ability to preserve heart function and prevent adverse remodeling at this later time (Fig.10B). This agrees with BT2’s downregulation of genes linked with cardiac remodeling, fibrosis and oxidative stress namely growth factors, extracellular superoxide dismutase (SOD) and lysyl oxidase. This includes Fgf2r2 (logFC -0.92 fold), Tgfb1 (logFC -0.97 fold), Tgfb2 (logFC -2.26 fold), Tgfbr1 (logFC -0.49 fold), Sod3 (logFC -1.92 fold) and Lox (logFC -1.15) (Table 2). BT2 inhibits IF size and preserves heart function after M / IR injury when delivered during myocardial ischaemia. To determine whether BT2 is cardioprotective when it is administered during an ischaemic event, rats were dosed i.p. with BT2 prior to reperfusion and assessed IF size and heart function 24 h after M / IR injury (Fig.11A). BT2 lowered IF / AAR (Fig. 11B), and neutrophil infiltration in the border zone (Fig. 11C). Conversely, BT2 increased both EF and FS (Fig. 12A and 12B) and reduced LVIDs (Fig.12C). These data, taken together, highlight the cardioprotective effects of BT2 in an animal model of M / IR injury and its potential to reprogram gene expression signatures that typify ischaemic disease severity and HF in ACS patients. BT2 inhibits multiple genes associated with disease severity in AMI patients. Interrogation of the STARNET database (Koplev, S., et al. Nat Cardiovasc Res 1, 85-100 (2022)) comparing gene expression in aortic tissue from CAD patients with those without CAD revealed that genes suppressed by BT2 in rodents are associated with disease severity. This includes EGR1 (padj=3.9x10-10), FOS (padj=3.9x10-5), FOSL1 (padj=3.2x10-19), JUN (padj=3.8x10-4) and ATF3 (padj=9.5x10-18). In STARNET, EGR1 and ATF3 are highly significant for DUKE CAD severity score (which predicts all-cause mortality) and JUN correlates with SYNTAX (Synergy between Percutaneous Coronary Intervention with Taxus and Cardiac Surgery) risk score (predictive of MACE). Gene expression in the Holvoet et al database (Holvoet, P., et al. PLoS One 14, e0225621 (2019)) constructed from monocytes sourced from AMI patients compared with stable CAD (GSE129935) was analysed. This comparison showed that mRNA levels of ATF3, FOSL1, FOSL2, KLF5, IL1ß, IL6R, TNFRSF21, HBEGF, OSM, CXCL1, CXCL2, TLR2, RIPK2, NLRP3, TREM1, TSC22D1 and NPPA are significantly increased DEG in the setting of AMI (Fig.13). Levels of all these DEG are reduced by BT2. In contrast, mRNA levels of SNED1 (nidogen), reduced in the setting of AMI, were increased by BT2. Nidogen increases heart function and reduces fibrosis in a mouse M / IR model (Zbinden, A., et al. Adv Sci (Weinh) 8, 2002500 (2021)). Gene expression in the Hall et al. database (Hall, J.L., et al. Physiol Genomics 17, 283-291 (2004)) constructed from LV apex samples harvested from AMI patients at the time of implant of an LV assist device compared with apex material sourced from non-ischaemic patients (GSE974) was analysed. FOSL2, IL1ß, TLR2 and TREM1 mRNA levels were elevated in AMI tissue whereas FOXP3 levels were reduced (Fig. 14). BT2 reduced Fosl2, Il1ß, Tlr2 and Trem1 mRNA levels and increased Foxp3 levels in rat M / IR injury model. Gene expression in the Muse et al. database (Muse, E.D., et al. Sci Rep 7, 12268 (2017)) constructed from circulating endothelial cells sourced from AMI patients and healthy individuals (GSE66360) was analysed. IL1ß, IL6, OSM, CXCL1, CXCL2, TLR2, NLRP3, TREM, FOSL1, FOSL2, ATF3, EGR1 and HB-EGF mRNA levels were elevated in AMI tissue (Fig.15). BT2 reduced mRNA levels of I11ß, Il6, Osm, Cxcl1, Cxcl2, Tlr2, Nlrp3, Trem, Fosl1, Fosl2, Atf3, Egr1 and Hbegf in the heart. Gene expression in the Park et al. database (Park, H.J., et al. Oncotarget 6, 12970-12983 (2015)) constructed from peripheral blood collected from ACS patients comparing STEMI, NSTEMI and UA patients with normal individuals (GSE61145, platform GPL6884) was analysed. OSM, IL6R, IL1R2, CXCL1, TLR2, TREM1 and FOSL2 mRNA levels were elevated in STEMI serum). OSM and TLR2 mRNA levels were also elevated in NSTEMI serum (Fig.16). BT2 reduced mRNA levels of Osm, Il6r, Il1r2, Cxcl1, Tlr2, Trem1 and Fosl2. These findings indicate that BT2 inhibits multiple genes associated with disease severity in ACS patients. BT2 inhibits multiple genes predictive of HF in STEMI patients undergoing PCI. Since the RNA-seq data revealed that BT2 reduced Nppa and Nppb levels in the heart (Table 2), it was explored whether other genes that were suppressed by BT2 may be linked with the progression of HF in STEMI patients. Gene expression was analysed in the Maciejak et al. database (Maciejak, A., et al. Genome Med 7, 26 (2015)) constructed from PBMCs isolated from individuals with stable CAD or 2 groups of STEMI patients that underwent PCI, one group that developed HF within 6 months and the other which did not develop HF (GSE59867). mRNAs predictive of HF progression elevated were NLRP3, TRPM2, FOSL2, IL1ß, CXCL1, CXCR2, CCL24, TLR6, IL1R2, HBEGF, FOS and FOSL1 (Fig. 17). Remarkably, BT2 reduced mRNA levels of Nlrp3, Trpm2, Fosl2, IL1ß, Cxcl1, Cxcl2, Ccl24, Tlr6, Il1r2, Hbegf, Fos and Fosl2. To illustrate the significance of these data in more global terms, all DEG in the HF group one day of AMI (all underwent direct PCI) (vs. stable CAD, padj<0.05) were compared with DEG in the BT2 treated group (vs. vehicle, padj<0.05). There were 4175 upregulated DEG and 3221 downregulated DEG in the HF group at 24 h. In contrast, there were just 77 upregulated and 77 downregulated DEG in the non-HF group (vs. stable CAD) (Fig. 18). Of the upregulated DEG in the HF group, 10.1 % (or 423) were the same DEG that BT2 downregulated in the rat AAR at 4 h (Fig. 18). Conversely, 6.3% (or 202) of downregulated DEG in the HF group were upregulated by BT2 in the AAR at 4 h (Fig. 18). The overlap of upregulated and downregulated DEG in HF patients with DEG downregulated and upregulated by BT2, respectively, effectively doubled when DEG in the HF group were compared with DEG in the BT2 group at 24 h. Remarkably, BT2 downregulated 18.5% (1 in 6) of DEG upregulated in the HF group (Fig. 18). Conversely, BT2 upregulated 12.5% (1 in 8) of DEG downregulated in the HF group (Fig. 18). This demonstrates the ability of BT2 to regulate the expression of genes predictive of heart failure. These data, taken together, highlight the cardioprotective effects of BT2 in an animal model of M / IR injury and its potential to reprogram gene expression signatures that epitomize ischaemic disease severity and HF in ACS patients undergoing PCI.

[0003] Table 2. Curated list of DEG regulated by BT2 (vs Veh) 4 h after M / IR injury. RNA-seq and bioinformatics analysis was performed from total RNA prepared from AAR treated with vehicle or BT24 h after M / IR injury. Genes downregulated by BT2 (vs Veh) at 4 h Gene log2FC Padj Gene type or function Mmp3 -4.6733001 0.01088119 Mmp8 -2.4820688 0.018644 Metalloproteinases Mmp25 -2.2184901 0.02854679 Osm -2.4113487 0.00706115 Il1ß -2.9225489 0.0179278 Tnfsf18 -3.0562752 0.00706115Pro-inflammatory cytokinesTnfrsf1b -1.0957269 0.02010979 Tnfrsf12a -1.6381505 0.01412738 Il1r2 -2.6940215 0.0320841Cytokine receptorsIsg15 -1.3999932 0.01518402 Effector CD8+T cell marker S100a8 -2.0577329 0.03073652 S100a9 -2.1022765 0.03215201DAMP biomarkersHspa1a -2.0265926 0.0185804 Tlr2 -2.2583706 0.01750286 Tlr1 -1.9276837 0.02924155 Tlr13 -1.478631 0.0160618 Ripk2 -1.3946156 0.00970051 DAMP receptors Nlrp3 -2.1778409 0.01518242 Trem1 -3.1087184 0.01318525 Trem3 -2.2896426 0.01799235 Trpm2 -1.7441191 0.02033392 Cxcl2 -2.4583681 0.04811277 Cxcl1 -1.8112501 0.02526025 Ccl6 -2.3634642 0.01493095 Ccl2 -2.2109829 0.04058337 Ccl24 -2.1400611 0.03211466 CXC chemokines and chemokines Ccl17 -1.619901 0.03211466 Ccl21 -1.0196948 0.02500677 Ccl12 -4.8830528 0.01097652 Ccl22 -2.8415787 0.01296949 Ccl3 -2.7227476 0.01973686 Nppb -1.6698998 0.01656057 HF marker Cd11b / Itgam -1.8581842 0.01412738Cd18 / Itgb2 -1.9776255 0.01634728 Neutrophil biomarkerHbegf -2.6837875 0.00970051 Growth factor Jun -0.8379349 0.04446886Junb -0.9219934 0.01559262Transcription factors Fosl1 -2.2360958 0.02741917 Egr1 -1.725542 0.02099399 Klf5 -2.3620146 0.00970051 Tsc22d1 -1.1603509 0.00970051Cnn2 -1.191314 0.00970051 Migration / activation ofmacrophages and neutrophilsXirp1 -1.4338603 0.00979259cardiac hypertrophy and fibrosis,cardiac remodelingPD-L1, upregulated in cardiac Cd274 -1.2095288 0.00970051 fibroblasts during cardiac remodelingCtgf -2.5431662 0.00979259Tgfb2 -1.1833952 0.01097652Growth factors regulating fibrosisGenes upregulated by BT2 (vs Veh) at 4 h Gene log2FC Padj Gene type or function Inhibits production of pro- inflammatory cytokines and Sned1 2.48739346 0.00706758 promotes production of anti- inflammatory cytokines Inhibits TGF-ß signaling and Htra3 1.55451718 0.00979259 ameliorates cardiac dysfunction after pressure overload Tcam1 2.9085497 0.01116205 Treg suppressor function

[0004] Table 3. Curated list of DEG regulated by BT2 (vs Veh) 24 h after M / IR injury. RNA-seq and bioinformatics analysis was performed from total RNA prepared from AAR treated with vehicle or BT224 h after M / IR injury. Genes downregulated by BT2 (vs Veh) at 24 h Gene log2FC Padj Gene type or function Mmp10-5.7985452 0.01920061Mmp12-3.6022046 0.00142161MetalloproteinasesMmp16-1.108544 0.03514491Il6 -3.7050706 0.02065624Il18Pro-inflammatory cytokines-1.5870192 0.0039057Il6r -0.8490598 0.0085099Il17raCytokine receptors-1.2239098 0.00425888Ctgf-2.5324009 0.00093571Tgfb2-2.2688308 0.00040405Growth factorsTgfb1-0.9717146 0.0009252Fgfr2-0.9287469 0.02611114Lox -1.151183 0.00270392Loxl2Growth factor receptors-0.8728095 0.00679259Loxl4-1.5910023 0.00704324Tnfsf18-1.3542744 0.00285686Tnfrsf21-1.1329723 0.00139967TNF superfamily and receptorsTnfrsf12a-2.1489357 0.00171274Cd68-1.708733 0.00043065Macrophage markerCd80-1.2920216 0.02256591Cd83-1.3048662 0.00158053M1 macrophage markerCd86-1.5734843 0.01481355Hspb1-1.7301101 0.00218811DAMP biomarkerTlr1-2.0715793 0.0074108Tlr2-1.6916792 0.01506231Tlr6-1.2198396 0.04115803Tlr7-1.3242122 0.00364128Tlr8-1.0329651 0.01866595DAMP receptors Tlr13-1.198744 0.01037564Nlrp3-1.2579368 0.02806665Trem1-1.7184838 0.03061275Trem3-2.2807912 0.0092105Trpm2-1.4006281 0.01107831Cxcl1-1.9038865 0.01315735Cxcl6-4.4987187 0.02953747Ccl2-2.8501103 0.00838453Ccl7-3.0336667 0.00769621Ccl9-2.5075614 0.01239469Chemokines Ccl12-3.1468874 0.00189169Ccl17-2.1253564 0.02130739Ccl22-1.7340681 0.03233231Ccl27-1.3791552 0.0094059Nppa -2.3792063 0.00139967NppbHF markers-1.1122855 0.02428916Fos-1.2658575 0.00975741Fosl1 -2.861743 0.00762257Fosl2Transcription factors-0.8362928 0.03542744Atf3-1.0363103 0.03017469Genes upregulated by BT2 (vs Veh) at 24 h Gene log2FC Padj Gene type or function Foxp3 2.20462526 0.00386384 Treg biomarker Trim50 2.89433861 0.00043065 Autophagy regulator Negative regulation of Ctla2a 1.28170958 0.00273888 inflammatory response Tcam1 1.51540592 0.00806506 Treg suppressor function Sod3 1.92686304 0.00033785 Antioxidant enzyme DISCUSSION Myocardial injury is caused by restoration, secondary to PCI or thrombolytic therapy, of coronary blood flow following ischaemia. In a rodent model of M / IR injury, in which the LAD is ligated for 30 min followed by 24 or 2 weeks of reperfusion, it was found that BT2 is an inhibitor of IF size, cardiac dysfunction and fibrosis. BT2 inhibited IF size in the AAR after M / IR injury, regardless of whether BT2 was delivered before or during myocardial ischaemia. On the other hand, BT3, a structural analogue, did not affect IF size indicating BT2’s effects on IF size was not due to a mass effect. BT2 also reduced serum levels of the cardiac biomarker troponin I, thereby demonstrating that BT2 reduces damage to the heart following M / IR injury. Remarkably, BT2 preserved heart function (no decrease in EF or FS) and prevented adverse remodeling (no increase in LVIDd or LVIDs) 24 h and 2 weeks after M / R injury. In fact, % EF, % FS, LVIDd and LVIDs in BT2 treated animals at 2 weeks were equivalent to that of the sham group (rats that underwent the surgical procedure without M / IR injury). BT2 also prevented myocardial fibrosis at 2 weeks after M / IR injury. Cardiac fibrosis is predictive of HF (Burchfield, J.S., Xie, M. & Hill, J.A. Circulation 128, 388-400 (2013). This result is important as LVIDd is an independent risk factor for poor long-term cardiovascular outcomes and an independent predictor of all-cause mortality in patients with coronary artery disease (Ito, K., et al. ESC Heart Fail 8, 4997-5009 (2021)). While infiltrating neutrophils provide the primary innate cellular response following M / IR injury, these cells also mediate myocardial damage and affect adverse LV remodeling and HF development in AMI patients. For example, increased neutrophil infiltration within the infarcted zone is associated with larger IF size and impaired LV function (Anderson, J.L. & Morrow, D.A. N Engl J Med 376, 2053-2064 (2017)). BT2 strongly reduced expression of neutrophil markers in the AAR within a few hours of M / IR injury. This suggests that BT2 may help to ameliorate this, most damaging (“The Bad”) phase of inflammation post-AMI (Matter, M.A., et al. Eur Heart J, ehad486 (2023)). Taken together, this work shows that BT2 is a novel small molecule inhibitor of M / IR injury and cardiac fibrosis. Summary ^ BT2 inhibits IF size in the area-at-risk after M / IR injury. ^ BT2 preserves heart function and prevents adverse remodelling after M / IR injury. ^ BT2 reduces inflammation resulting from M / IR injury in multiple cells types. ^ BT2 inhibits multiple genes associated with coronary artery disease (CAD) severity ^ BT2 inhibits multiple genes predictive of HF in ACS and / or STEMI patients. Also described herein are the following items 1 to 18: 1. A method of reducing or preventing inflammation and / or myocardial cell death in heart tissue of a subject following reperfusion of ischaemic tissue in the heart of the subject, comprising administering to the subject an effective amount of a compound of formula (I), or a pharmaceutically acceptable salt thereof: OR2H R1is straight or branched C1-C6alkyl; and R2is straight or branched C1-C6alkyl, or R2is R3 4; and R3is straight or branched C1-C6alkyl. 2. The method of item 1, wherein R1is straight C1-C6alkyl or branched C1-C6alkyl. 3. The method of item 1, wherein R1is -CH2CH3or -CH2CH(CH3)2. 4. The method of item 1, wherein R2is straight C1-C6alkyl or branched C1-C6alkyl. 5. The method of item 1, wherein R2is -CH2CH3or -CH2CH(CH3)2. 6. The method of item 1, wherein the compound of formula (I) is a compound of formula (I-1): O2HR wherein: R2is straight or branched C1-C6alkyl; or R2is: R3wherein q is R3is straight or branched C1-C6alkyl. 7. The method of item 1, wherein the compound of formula (I) is a compound of formula (II): 8. The method of any compound is administered following an ischaemic event. 9. A method of reducing or preventing left ventricular remodelling in a subject following reperfusion of ischaemic tissue in the heart of the subject, comprising administering an affective amount of a compound of formula I, or a pharmaceutically acceptable salt thereof. 10. A method of reducing or preventing cardiac fibrosis following reperfusion of ischaemic tissue in a heart of a subject, comprising administering an affective amount of a compound of formula I, or a pharmaceutically acceptable salt thereof. 11. A method of reducing or preventing loss of heart function following reperfusion of ischaemic tissue in a heart of a subject, comprising administering an affective amount of a compound of formula I, or a pharmaceutically acceptable salt thereof. 12. The method of any one of items 9 to 11, wherein the compound of formula (I) is a compound of formula (II): of items 1 to 12, wherein inflammation is reduced in myocardium of the subject. 14. The method of any one of items 1 to 13, wherein myocardial cell death is reduced in the heart of the subject. 15. The method of any one of items 1 to 14, wherein expression of one or more genes selected from the group consisting of ATF3, FOS, FOSL1, FOSL2, MMP10, MMP16, IL6, IL6R, IL17RA, IL18, TNFSF18, TNFRSF12A, CD68, HSPB1, TLR1, TLR2, TLR6, TLR7, NLRP3, TREM1, TREM3, TRPM2, CXCL1, CXCL6, CCL2, CCL6, CCL17, CCL22, CCL24, NPPA and NPPB is decreased in heart tissue of the subject. 16. The method of any one of items 1 to 15, wherein expression of one or more genes selected from the group consisting of FOXP3, TRIM50, CTLA2a, TCAM1 and SOD3 is increased in heart tissue of the subject. 17. A kit for reducing or preventing inflammation and / or myocardial cell death in the heart of a subject following reperfusion of ischaemic tissue in the heart of the subject, the kit comprising a compound of formula (I), or a pharmaceutically acceptable salt thereof. 18. A kit for reducing or preventing inflammation and / or myocardial cell death in the heart of a subject following reperfusion of ischaemic tissue in the heart of the subject, the kit comprising a compound of formula (II): , (II), or a salt thereof.

Claims

CLAIMS:

1. A method for reducing or preventing myocardial ischaemic perfusion (M / IR) injury in a subject, comprising administering an effective amount of a compound of formula (I), or a pharmaceutically acceptable salt thereof: OR2HR1is straight or branched C1-C6alkyl; and R2is straight or branched C1-C6alkyl, or R2is R32, 3 or 4; and R3is straight or branched C1-C6alkyl.

2. The method of claim 1, wherein the reducing or preventing myocardial ischaemic perfusion injury reduces or prevents inflammation and / or myocardial cell death in heart tissue of a subject following reperfusion of ischaemic tissue in the heart of the subject.

3. The method of any one of claims 1 to 2, wherein the reducing or preventing myocardial ischaemic perfusion injury reduces or prevents cardiac fibrosis in heart tissue of a subject following reperfusion of ischaemic tissue in the heart of the subject.

4. The method of any one of claims 1 to 3, wherein the reducing or preventing myocardial ischaemic perfusion injury reduces or prevents left ventricular remodelling in a subject following reperfusion of ischaemic tissue in the heart of the subject.

5. The method of any one of claims 1 to 4, wherein the reducing or preventing myocardial ischaemic perfusion injury reduces or prevents loss of heart function in a subject following reperfusion of ischaemic tissue in the heart of the subject.

6. The method of any one of claims 1 to 4, wherein the reducing or preventing myocardial ischaemic perfusion injury reduces the risk of heart failure following reperfusion of ischaemic tissue in the heart of the subject.

7. The method of any one of claims 1 to 4, wherein the reducing or preventing myocardial ischaemic perfusion injury reduces or prevents heart failure following reperfusion of ischaemic tissue in the heart of the subject.

8. The method of any one of claims 1 to 7, wherein the reducing or preventing myocardial ischaemic perfusion injury reduces or prevents myocardial infarct formation in heart tissue following reperfusion of ischaemic tissue in the heart of the subject.

9. The method of any one of claims 1 to 8, wherein R1is straight C1-C6alkyl or branched C1-C6alkyl.

10. The method of any one of claims 1 to 9, wherein R1is -CH2CH3or -CH2CH(CH3)2.

11. The method of any one of claims 1 to 10, wherein R2is straight C1-C6alkyl or branched C1-C6alkyl.

12. The method of any one of claims 1 to 11, wherein R2is -CH2CH3or -CH2CH(CH3)2.

13. The method of any one of claims 1 to 12, wherein the compound of formula (I) is a compound of formula (I-1), or a pharmaceutically acceptable salt thereof: O2HRwherein: R2is straight or branched C1-C6alkyl; or R2is: R3wherein q isR3is straight or branched C1-C6alkyl.

14. The method of any one of claims 1 to 13, wherein the compound of formula (I) is a compound of formula (II), or a pharmaceutically acceptable salt thereof:

15. The method of any one of claims 1 to 14, wherein the compound, or a pharmaceutically acceptable salt thereof, is administered prior to, during or after an ischaemic event.

16. The method of any one of claims 1 to 15, wherein the compound, or a pharmaceutically acceptable salt thereof, is administered prior to reperfusion.

17. The method of any one of claims 1 to 16, wherein the compound, or a pharmaceutically acceptable salt thereof, is administered parenterally.

18. The method of claim 17, wherein the parenteral administration is intravenous or intraperitoneal.

19. The method of any one of claims 1 to 18, wherein the subject has undergone myocardial infarction.

20. The method of any one of claims 1 to 19, wherein the myocardial infarction is ST-elevation myocardial infarction.

21. The method of any one of claims 1 to 20, wherein the M / IR injury results from PCI.

22. The method of any one of claims 1 to 20, wherein the M / IR injury results from thrombolysis.

23. The method of any one of claims 1 to 20, wherein the M / IR injury results from coronary bypass.

24. A method of treating a subject suffering from an ischaemic event, comprising: (a) administering a compound of Formula (I), or a pharmaceutically acceptable salt thereof; and (b) restoring blood flow to ischaemic tissue.

25. The method of claim 24, wherein the ischaemic tissue is myocardium.

26. The method of claim 24 or 25, wherein the ischaemic event is an acute coronary syndrome.

27. The method of any one of claims 24 to 26, wherein the ishaemic event is a myocardial infarction.

28. The method of any one of claims 24 to 27, wherein the compound of formula (I) is a compound of formula (II), or a pharmaceutically acceptable salt thereof:

29. The method of any one of claims 1 to 28, wherein expression of one or more genes selected from the group consisting of ATF3, FOS, FOSL1, FOSL2, MMP10, MMP16, IL6, IL6R, IL17RA, IL18, TNFSF18, TNFRSF12A, CD68, HSPB1, TLR1, TLR2, TLR6, TLR7, NLRP3, TREM1, TREM3, TRPM2, CXCL1, CXCL6, CCL2, CCL6, CCL17, CCL22, CCL24, NPPA and NPPB is decreased in myocardium of the subject.

30. The method of any one of claims 1 to 29, wherein expression of one or more genes selected from the group consisting of FOXP3, TRIM50, CTLA2a, TCAM1 and SOD3 is increased in myocardium of the subject.

31. A kit for reducing or preventing myocardial ischaemic-perfusion injury in a subject resulting from reperfusion of ischaemic tissue in the heart of the subject, the kit comprising a compound of formula (I), or a pharmaceutically acceptable salt thereof.

32. A kit for reducing or preventing myocardial ischaemic-perfusion injury in a subject resulting from reperfusion of ischaemic tissue in the heart of the subject, the kit comprising a compound of formula (II): , (II), or asalt thereof.

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