A Randomized Controlled Trial of Angiotensin-Converting Enzyme Inhibition for Skeletal Muscle Dysfunction in COPD

BACKGROUND: Skeletal muscle impairment is a recognized complication of COPD, predicting mortality in severe disease. Increasing evidence implicates the renin-angiotensin system in control of muscle phenotype. We hypothesized that angiotensin-converting enzyme (ACE) inhibition would improve quadriceps function and exercise performance in COPD. METHODS: This double-blind, randomized placebo-controlled trial investigated the effect of the ACE inhibitor, fosinopril, on quadriceps function in patients with COPD with quadriceps weakness. Primary outcomes were change in quadriceps endurance and atrophy signaling at 3 months. Quadriceps maximum voluntary contraction (QMVC), mid-thigh CT scan of the cross-sectional area (MTCSA), and incremental shuttle walk distance (ISWD) were secondary outcomes. RESULTS: Eighty patients were enrolled (mean [SD], 65 [8] years, FEV1 43% [21%] predicted, 53% men). Sixty-seven patients (31 fosinopril, 36 placebo) completed the trial. The treatment group demonstrated a significant reduction in systolic BP (Δ−10.5 mm Hg; 95% CI, −19.9 to −1.1; P = .03) and serum ACE activity (Δ−20.4 IU/L; 95% CI, −31.0 to −9.8; P < .001) compared with placebo. No significant between-group differences were observed in the primary end points of quadriceps endurance half-time (Δ0.5 s; 95% CI, −13.3-14.3; P = .94) or atrogin-1 messenger RNA expression (Δ−0.03 arbitrary units; 95% CI, −0.32-0.26; P = .84). QMVC improved in both groups (fosinopril: Δ1.1 kg; 95% CI, 0.03-2.2; P = .045 vs placebo: Δ3.6 kg; 95% CI, 2.1-5.0; P < .0001) with a greater increase in the placebo arm (between-group, P = .009). No change was shown in the MTCSA (P = .09) or ISWD (P = .51). CONCLUSIONS: This randomized controlled trial found that ACE inhibition, using fosinopril for 3 months, did not improve quadriceps function or exercise performance in patients with COPD with quadriceps weakness. TRIAL REGISTRY: Current Controlled Trials; No.: ISRCTN05581879; URL: www.controlled-trials.com

Skeletal muscle impairment is a key extrapulmonary complication of COPD, aff ecting approximately onethird of patients independent of the degree of airfl ow obstruction. 1,2 In particular, quadriceps weakness in COPD has been associated with reduced exercise capacity, 3 impaired quality of life, 4 and mortality in patients with moderate to severe disease. 5 Importantly, pulmonary rehabilitation, which improves exercise performance and reduces health-care utilization, also increases quadriceps strength. 6,7 Th e mechanisms responsible for skeletal muscle dysfunction in COPD remain a matter of controversy and are likely multifactorial, but there is evidence that chronic activation of the IM renin-angiotensin system may be a key pathophysiologic pathway. 8 In animal models, [9][10][11] angiotensin II promotes muscle loss via an inhibitory eff ect on the insulin-like growth factor (IGF)-1 system and stimulation of a catabolic pathway mediated by two ubiquitin ligases, the atrogenes: muscle RING fi nger protein-1 and atrogin-1. Th rough ubiquitinproteasome degradation, these ligases have been postulated to play a key role in the muscle atrophy observed in patients with COPD. 12 In addition, an endogenous reduction in serum and tissue angiotensin-converting enzyme (ACE) levels as a result of polymorphism of the human ACE gene has been associated with an enhanced endurance phenotype 13 with the presence of a deletion allele (D) shown to correlate with greater quadriceps strength in COPD. 14 A polymorphism determining a reduction in bradykinin

Materials and Methods
Participants Th e study was approved by the Joint University College London Committees on the Ethics of Human Research (reference number 08/H0715/90), and each participant gave informed written consent. Study inclusion criteria were patients diagnosed with COPD based on GOLD (Global Initiative for Chronic Obstructive Lung Disease) criteria 23 and the presence of quadriceps weakness defi ned as a quadriceps maximum voluntary contraction (QMVC) in kilograms , 120% of the patient's BMI. 5 Exclusion criteria were patients within 3 months of pul-monary rehabilitation or 1 month of an exacerbation, and those with a comorbidity including cardiac failure, diabetes, renal disease, or rheumatoid arthritis. Patients receiving ACE inhibitors, angiotensin II receptor blockers, or warfarin (because the study entailed a vastus lateralis biopsy) were also excluded. receptor expression has also been associated with a reduced fat-free mass and quadriceps strength in patients with COPD. 15 Further evidence has come from observational studies in hypertensive cohorts where treatment with an ACE inhibitor has been associated with increased locomotor muscle size 16 and strength. 17 Th is is supported by randomized controlled trials where ACE inhibition has FOR EDITORIAL COMMENT SEE PAGE 878 increased 6-min walking distance in elderly subjects 18 and angiotensin II receptor blockade has shown a trend toward an improvement in quadriceps strength in subjects with COPD. 19 Given this evidence base and the increasing focus toward the development of pharmacotherapy targeting skeletal muscle, 20,21 we hypothesized that ACE inhibition would have a benefi cial eff ect on quadriceps function in patients with COPD. To strengthen the design of the trial we used a stratifi ed medicine approach, selecting patients with quadriceps weakness using the cutoff that has been found to be associated with increased mortality in COPD. 5 Quadriceps endurance, measured by repetitive magnetic stimulation, was used as the primary outcome due to the asso ciation of reduced ACE levels with greater endurance in healthy subjects 13 and the advantages of a nonvolitional test in the context of a population with severe airfl ow limitation. 22 Resting BP and renal function were reviewed at 1 week by an independent assessor and if satisfactory, the daily dose was increased to two capsules (fosinopril 20 mg maximum or placebo). Dose was not escalated if the systolic BP was , 110 mm Hg. A pharmacy controlled, 1:1 randomization in blocks of four using consecutive numbers was performed by the Clinical Trials Department, Royal Free Hampstead NHS Trust UK. Primary outcomes were change in quadriceps endurance measured nonvolitionally by repetitive magnetic stimulation 22 and vastus lateralis atrophy signaling using atrogene expression from biopsies taken at baseline and 3 months. Secondary outcomes included change in QMVC, quadriceps twitch force (TwQ), mid-thigh CT scan of the crosssectional area (MTCSA), incremental shuttle walk distance (ISWD), health status, and serum infl ammatory markers. Measurements of fatfree mass, BP, lung function, and anthropometrics were also made and a single assessment of baseline physical activity was conducted using a multisensor armband worn for 1 week as previously described. 2 Further details of the trial protocol and methods are available online ( e-Appendix 1 , e-Tables 1, 2 ).

Analysis and Statistics
Detecting a 25% increase in time to fatigue in the fosinopril vs placebo groups, with an 80% power at the 5% signifi cance level, would require 54 patients randomized on a 1:1 basis. To allow for a 30% dropout rate, 80 patients were targeted for recruitment. Data were normally distributed and a per-protocol analysis was conducted using paired or independent t tests. Response variables were tested by analysis of variance with post hoc correction for more than two groups. Multiple linear regression analysis was performed to identify baseline parameters infl uencing response to treatment. Analysis was performed using StatView 5.0 (Abacus Concepts) with P , .05 considered statistically signifi cant.

Results
One hundred seventeen patients were screened for study participation, and 80 patients underwent randomization. Th ere were eight withdrawals from the treatment group and five from the placebo group. Further details are shown in the CONSORT diagram ( Fig 1 ).

Baseline Anthropometrics and Muscle Measurements
Th e placebo and treatment groups were well matched for age, sex, and lung function parameters and there were no statistically signifi cant baseline diff erences in body composition and quadriceps muscle function ( Table 1 ). Th ere were also no signifi cant baseline diff er-ences in vastus lateralis atrogene expression and serum measurements between the groups ( e- Table 3 ).  ( Fig 2A ). QMVC improved in both groups (fosinopril: ⌬ 1.1 kg; 95% CI, 0.03-2.2; P 5 .045 vs placebo: ⌬ 3.6 kg; 95% CI, 2.1-5.0; P , .0001) with a greater increase in the placebo arm (betweengroup diff erence, 2.5 kg; 95% CI, 0.7-4.3; P 5 .009) ( Fig 2B ). TwQ response did not diff er in the placebo vs treatment  Lung function parameters including FEV 1 % predicted, diff using capacity of the lung for carbon monoxide % predicted, residual volume-to-total lung capacity ratio, and arterial blood gas measurements showed no significant change between groups at 3 months, as shown in Table 2 . Health-related quality-of-life measures, which included St. George's Respiratory Questionnaire (SGRQ) and COPD assessment test (CAT) score, also did not vary signifi cantly between groups ( Table 2 ).

Molecular Outcomes Following ACE Inhibition
Vastus Lateralis messenger RNA Expression: At 3 months, no signifi cant diff erences were observed in vastus lateralis atrogin-1 messenger RNA (mRNA)  2D ). No signifi cant diff erences were found in serum IGF-1, high-sensitivity C-reactive protein, N-terminal pro-B-type natriuretic peptide, fi brinogen, or serum infl ammatory cytokines between the groups as shown in e- Table 4 . A post hoc stratifi cation based on ACE genotype did not infl uence response to treatment as detailed online ( e-Tables 5, 6 ). Multiple linear regression analysis did not identify any baseline demographic, anthropometric, lung function, or physical activity variables infl uencing response to treatment. Data on dropouts from the study are shown in e- Table 7 .

Discussion
ACE inhibition by fosinopril did not have a benefi cial eff ect on quadriceps muscle function over a 3-month period in patients with COPD selected for quadriceps weakness, although measurements of BP and serum ACE activity confi rmed both adherence and biologic activity of the drug. Moreover, we were unable to detect any changes in atrophy signaling pathways in the participants. Th e present data do not support the use of ACE inhibitors alone to augment muscle phenotype in patients with COPD.
Th ese fi ndings were unexpected given the evidence for ACE inhibition on skeletal muscle function, and reinforce the importance of conducting prospective controlled trials. An observational study by Onder et al 17 assessed the relationship between ACE inhibitor use and muscle strength in 641 elderly hypertensive women. Th ey found that at 3 years' follow-up, participants taking an ACE inhibitor continuously had a lower mean decline in both knee extensor muscle strength and walking speed than those using other antihypertensives and those not on any antihypertensive medications. Intermittent use of ACE inhibitors was also associated with a signifi cantly larger decline in walking speed compared with continuous use. Th e study group had poor mobility but no concomitant heart failure at baseline. In addition, crosssectional data from 2,431 hypertensive subjects was used to evaluate whether ACE inhibitor treatment is associated with a larger lower extremity muscle mass compared with the use of other antihypertensive medications, 16 and found that lower extremity muscle mass was larger in the ACE inhibitor group in a manner proportional to the length of use.
Interventional studies of ACE inhibition have also suggested a treatment eff ect. A randomized controlled trial in 95 elderly subjects with self-reported diffi culties in mobility, showed that 5 months of perindopril treatment signifi cantly improved 6-min walk distance (31.4 m; 95% CI, 10.8 m-51.9 m; P 5 .003) compared with placebo. 18 Interestingly, this cohort included current and ex-smokers and the improvement was observed in the absence of heart failure in the participants. In addition, a small randomized controlled trial of 21 subjects with COPD evaluating the eff ects of 4 weeks' treatment with enalapril on exercise performance found a signifi cant improvement in O 2 pulse and peak work rate in the treatment group. 24 Th ere are a number of possible factors that may explain why ACE inhibition was not eff ective in the present study given previous data. Th ese are discussed here and include the patient population studied, the infl uence of physical inactivity, and the possibility that ACE inhibition created a more "benign" IM environment, eff ectively removing a training stimulus.

Patient Selection
We adopted a stratifi ed medicine approach, selecting a quadriceps weakness patient phenotype, to focus on those patients with COPD with a level of skeletal muscle dysfunction known to be associated with worse survival. 5 It may, however, be that at this stage the weakest patients have a limited ability to respond to treatment and the low physical activity level found at baseline may refl ect this. Th e level of inactivity could explain the discrepancy between the present data and the eff ect of ACE inhibitors in relatively healthy populations being treated for hypertension. 17 Th ere is also evidence that an exercise stimulus may be needed for ACE inhibition to promote adaptive changes, such as an increase in capillary density, in skeletal muscle. 25 Th erefore, the current data do not preclude the possibility that the use of ACE inhibition in the context of pulmonary rehabilitation may yield benefi t.
Skeletal muscle impairment in COPD involves both fi ber atrophy and fi ber shift away from an oxidative, fatigue-resistant phenotype. 26 In any individual, these processes occur to a varying extent with diff erent eff ects on muscle function. The D allele of the ACE (I/D) polymorphism, which is associated with higher ACE activity, was associated with greater quadriceps strength in patients with COPD. 14 Th e eff ects of ACE inhibition may therefore counteract strength adaptations seen in COPD, in favor of achieving a more aerobic phenotype. Th is trial was not prospectively powered for stratifi cation by ACE genotype, so although post hoc analysis did not identify a genotype-specifi c infl uence on quadriceps function, this method of stratifi cation would be an important consideration for future studies.

Eff ect of ACE Inhibition on IM Environment
Th e improvements observed in quadriceps strength in the current trial warrant further discussion. As subjects were included based on the presence of quadriceps weakness, it is not completely unexpected that a potential placebo eff ect with regression to the mean would be observed in relation to volitional quadriceps strength, although more detailed monitoring of physical activity throughout the trial period would be needed to support this. Th e existence of a placebo eff ect has been well doc-umented in the context of functional exercise capacity in clinical trials of ACE inhibition in heart failure 27 and in other studies of physical performance. 28 However, the fi nding that this eff ect was greater in the placebo arm of the trial was unexpected. While this could be a chance eff ect, this seems unlikely because of the high level of statistical signifi cance shown ( P 5 .009). A possible reduction in skeletal muscle blood fl ow secondary to a reduced systemic BP may explain the attenuated response observed with treatment. Of note, captopril has been shown to increase maximal blood lactate during exercise and reduce exercise capacity in normotensive, sedentary rats. 29 Despite evidence from animal models of the ability to prevent angiotensin II-induced cachexia via IGF-1 overexpression and atrogene downregulation, 10 30 However, local tissue renin-angiotensin system can generate angiotensin II, independent of ACE activity, through serine proteases such as chymase and cathepsin G 31 and this may explain the lack of influence of ACE inhibition on skeletal muscle. The timing of biopsies may have also influenced the ability to detect changes following ACE inhibition and, therefore, early biopsies, where feasible, may guide further mechanistic work in this area.

Critique of the Method
A strength of this study was that the quadriceps assessment was comprehensive including both volitional and nonvolitional physiologic outcomes as well as molecular outcomes. Although no signifi cant changes were shown in the activity domain of the SGRQ, the infl uence of treatment on physical activity level was not objectively assessed in this trial. Th e baseline inclusion of physical activity monitoring did, however, objectively confi rm that both groups were well matched for their level of physical activity when commencing the trial. Importantly, this activity level was in keeping with published data on patients from other northern European countries, 32 highlighting the overall generalizability of the study population findings. However, the inclusion of more patients with varying degrees of muscle impairment may have enabled a wider assessment of potential responders.
Finally, a 3-month study duration was chosen as this was comparable to the time period used in other relevant studies 19,24 and would minimize potential confounding by intercurrent exacerbations. However, the time needed for skeletal muscle adaptation following pharmacotherapy in these patients is unclear, with ACE inhibitor treatment duration ranging from 10 weeks to 12 months for studies of improved 6-min walk distance in heart failure. 27 The potential role of longer-term therapy remains to be evaluated in further trials.

Conclusions
In summary, despite a strong theoretical basis for the study, we found that the ACE inhibitor fosinopril did not improve quadriceps function over a 3-month period, in a COPD population selected for quadriceps weakness. Th is study does not support a role for ACE inhibition alone in the treatment of skeletal muscle dysfunction in patients with COPD. Future work should focus on the use of pharmacotherapy during pulmonary rehabilitation to investigate augmenting exercise eff ects in these patients.