Impact of Myocardial Fibrosis on Cardiovascular Structure, Function and Functional Status in Heart Failure with Preserved Ejection Fraction

The trial design and results of the PIROUETTE trial (Clinicaltrials.gov NCT02932566) have been published previously [12, 14]. In brief, between March 7, 2017, and December 19, 2018, 94 patients with HFpEF and myocardial fibrosis were randomised to receive pirfenidone or placebo treatment for 52 weeks. Eligibility requirements included patients being ≥ 40 years of age, symptoms and signs of heart failure, left ventricular ejection fraction of ≥ 45% and elevated natriuretic peptides (brain natriuretic peptide (BNP) ≥ 100 pg/ml or N-terminal pro-B-type natriuretic peptide (NT-proBNP) ≥ 300 pg/ml or BNP ≥ 300 pg/ml or NT-proBNP ≥ 900 pg/ml if atrial fibrillation (AF) present). Patients deemed eligible underwent CMR, and those with evidence of myocardial fibrosis, defined as having an ECV of 27% or higher, were randomised to treatment with either pirfenidone or matching placebo for 52 weeks stratified by sex. Exclusion criteria included patients having an alternative cause of symptoms such as significant respiratory disease, obesity or anaemia; hypertrophic cardiomyopathy, pericardial constriction or infiltrative cardiomyopathy; and contraindication to CMR imaging. The primary outcome was change in myocardial fibrosis from baseline to 52 weeks, measured using CMR ECV.

The trial was sponsored by Manchester University NHS Foundation Trust, and trial management, independent data management and independent statistical analyses were performed by Liverpool Clinical Trials Centre, a UK Clinical Research Collaboration Clinical Trials Unit. Trial conduct was overseen by a trial steering committee. The study protocol was approved by a research ethics committee. Patients were identified at six UK hospitals and study visits took place at Manchester University NHS Foundation Trust. All patients provided written informed consent.

The trial procedures, analysis methods and outcome measurements have been described previously [12, 14]. In brief, echocardiography, CMR, electrocardiography, 6-min walk test, laboratory tests and the Kansas City Cardiomyopathy Questionnaire (KCCQ) were performed both at baseline and repeated after 52 weeks of treatment. ³¹Phosphorous magnetic resonance spectroscopy (³¹P-MRS) was also performed at baseline and 52 weeks in a subset of patients as part of a predefined sub-study (n = 60).

Myocardial ECV was calculated from basal and mid-left ventricular (LV) short axis pre- and post-contrast T1 maps (Modified Look-Locker Inversion [MOLLI] recovery). Images were acquired before and 15 min after administration of gadolinium contrast (0.15 mmol/kg of gadoterate meglumine), as ECV = (1 – haematocrit) × [ΔR1_myocardium]/[ΔR1_bloodpool], where ΔR1 is the difference in relaxation rates (1/T1) between the pre- and post-contrast [4]. The haematocrit was measured on the same day as the CMR scan. Absolute myocardial extracellular matrix (ECM) volume was calculated as the product of LV myocardial volume (LV mass divided by the known specific gravity of myocardium [1.05 g/ml]) and ECV. Absolute myocardial cellular volume was calculated as the product of LV myocardial volume (LV mass divided by the known specific gravity of myocardium [1.05 g/ml]) and (1 – ECV). Full details can be found in the trial protocol paper [14].

Analysis was conducted on an intention to treat basis, including all randomised patients retained in their randomised treatment groups. Continuous data are presented as mean ± standard deviation (SD) or as median (interquartile range (IQR)), as appropriate. Categorical data are presented as counts and percentages. Correlation analyses were used to assess associations between change in ECV (week 52 value minus baseline value) and change in selected secondary outcome variables that reflected cardiac mechanical and electrical function, circulating biomarkers and functional status. Pearson’s or Spearman’s correlation coefficients were used as appropriate following Shapiro–Wilk testing for normality. Analyses were performed in Stata (version 14.0, StataCorp, College Station, TX) and SAS (version 9.4, SAS Institute, Inc., Cary, NC).

Mediation analyses were conducted in order to determine whether changes in myocardial fibrosis, measured using ECV and absolute ECM volume, and changes in myocardial cellular volume (the mediator variables) following antifibrotic therapy, caused changes in cardiovascular structure and function, circulating biomarkers and functional status (the outcome variables) (Fig. 1).

Mediation analyses were conducted using the Baron and Kenny approach [15], under a structural equation modelling (SEM) framework (Stata package medsem), in order to estimate the average causal mediation effect (ACME) of each mediator, adjusting for confounders (baseline covariates that predicted both the mediator and outcome at the 10% significance level). For each potential outcome, the analysis was only performed if both of the following conditions were satisfied:

The outcome variables for this study were all secondary outcome measurements in the main PIROUETTE trial. The outcome variables reflect cardiovascular structure and function, circulating biomarkers and functional status. They were selected for use in this study because they are variables that, based on the published literature and clinical judgement, are associated with adverse outcome in HFpEF or were hypothesised to be impacted by myocardial fibrosis. The selection of the outcome variables was prospective; i.e. outcome variables were selected before data lock for the trial had occurred, thus before the results of the trial were known and were prespecified in an ‘Additional Statistical Analysis Plan’ that was written before data lock.

Baseline characteristics of the 94 patients that were randomised are presented in Table 1. The mean age of patients was 78 years, and 46% were female. Nearly all patients had New York Heart Association functional class II or III symptoms (95%), the mean LV ejection fraction was 64%, and the median NT-proBNP was 1104 pg/ml. The mean myocardial ECV was 30.1%. At the end of the trial, 12 patients had withdrawn from the study and two had died. No patient was lost to follow-up. Only patients with complete data were included in the mediation analysis (n = 79).

Change in myocardial ECV from baseline to week 52 showed a positive correlation with change in left ventricular end-diastolic volume indexed for body surface area (r = 0.23, p = 0.039), and inverse correlations with 6-min walk test distance (r =  − 0.28, p = 0.021), and KCCQ Clinical Summary Score (r =  − 0.23, p = 0.045) (Fig. 2 and Table S1 in Supplementary Appendix), although all the associations were weak.

Pirfenidone had a significant effect on the mediator variables measuring myocardial fibrosis (myocardial ECV and absolute myocardial ECM volume). The effect of pirfenidone on the mediator variable myocardial cellular volume was not significant at the conventional 5% level but was significant at the 10% level, and thus, as prespecified, this mediator variable was also included in the mediation analysis (Table 2).

The only outcome variable that demonstrated a treatment effect was LV ejection fraction (p = 0.011) (Table 3).

No baseline covariates were found to predict the mediator variables and LV ejection fraction; thus, no baseline covariates were required to be included in the mediation analysis (Table S2 in Supplementary Appendix).

In the mediation analysis, the estimated average causal mediation effects of myocardial ECV, absolute myocardial ECM volume and absolute myocardial cellular volume on LV ejection fraction were 6.1%, 21.5% and 13.7%, respectively, none of which was significant (p = 0.608, p = 0.123 and p = 0.186, respectively) (Table 4 and Fig. 3).

As discussed, the PIROUETTE trial was not powered for secondary outcomes; thus, the findings of this study are considered exploratory. The analyses conducted as part of the current study were not included in the Statistical Analysis Plan for PIROUETTE and thus are considered post hoc. Nevertheless, the analyses conducted in this study were prespecified in an ‘Additional Statistical Analysis Plan’ that was written before trial data lock. As discussed, the selection of outcome variables to include in this study was prospective and performed before data lock. Finally, no adjustment for multiple comparisons was performed; therefore, false positive results cannot be excluded.