Efficacy and Safety of PCSK9 Inhibition With Evolocumab in Reducing Cardiovascular Events in Patients With Metabolic Syndrome Receiving Statin Therapy: Secondary Analysis From the FOURIER Randomized Clinical Trial.

IMPORTANCE: The PCSK9 inhibitor evolocumab reduced low-density lipoprotein cholesterol and cardiovascular events in the FOURIER randomized clinical trial. Patients with metabolic syndrome (MetS) are at increased cardiovascular risk. OBJECTIVE: To investigate outcomes with evolocumab in patients with and without MetS. DESIGN, SETTING, AND PARTICIPANTS: The FOURIER trial randomized patients worldwide with stable atherosclerotic cardiovascular disease receiving statin to evolocumab vs placebo with follow-up for a median of 2.2 years. Data were collected February 2013 to November 2016. For this prespecified analysis, patients with the requisite data were stratified based on the National Cholesterol Education Program Adult Treatment Panel III MetS criteria; in secondary analyses, patients were further substratified by diabetes at baseline. Analysis was intention to treat. Analysis began March 2018 and ended April 2020. INTERVENTIONS: Patients were randomized to evolocumab or placebo. MAIN OUTCOMES AND MEASURES: The primary end point was cardiovascular death, myocardial infarction, stroke, hospitalization for unstable angina, or coronary revascularization. The key secondary end point was cardiovascular death, myocardial infarction, or stroke. RESULTS: Of 27 342 patients (mean [SD] age, 63 [9] years; 20 623 men [75.4%]) included in this analysis, 16 361 (59.8%) with baseline MetS were, when compared with patients without MetS, at higher risk of cardiovascular events (adjusted hazard ratio [95% CI], 1.31 [1.18-1.46]; P < .001 for the primary and 1.38 [1.20-1.57]; P < .001 for the key secondary end point). Evolocumab reduced low-density lipoprotein cholesterol similarly in patients with MetS (median [interquartile range], 92 [79-109] mg/dL vs 30 [19-48] mg/dL; P < .001) and without MetS (median [interquartile range], 92 [81-108] mg/dL vs 29 [18-44] mg/dl; P < .001). For the primary end point, the hazard ratios (95% CI) with evolocumab vs placebo were 0.83 (0.76-0.91) and 0.89 (0.79-1.01) in patients with and without MetS (P for interaction = .39). For the key secondary end point, the corresponding hazard ratios (95% CIs) were 0.76 (0.68-0.86) and 0.86 (0.74-1.01) (P for interaction = .23), respectively. Evolocumab did not increase the risk of new-onset diabetes or other major safety outcomes including worsening glycemic control, compared with placebo in patients with MetS. CONCLUSIONS AND RELEVANCE: Patients with atherosclerotic cardiovascular disease and MetS have substantial residual risk of cardiovascular events despite statin therapy. Evolocumab significantly reduced low-density lipoprotein cholesterol and cardiovascular risk in patients with MetS without increasing new-onset diabetes, worsening glycemic control, or other major safety events. These data suggest the addition of evolocumab to statin therapy in patients with atherosclerotic cardiovascular disease and MetS is safe and efficacious to reduce residual cardiovascular risk. TRIAL REGISTRATION: ClinicalTrials.gov Identifier: NCT01764633

Clinical Efficacy And Safety Of Evolocumab In High-Risk Patients Receiving A Statin Secondary Analysis Of Patients With Low Ldl Cholesterol Levels And In Those Already Receiving A Maximal-Potency Statin In A Randomized Clinical Trial

IMPORTANCE Current guidelines for atherosclerotic cardiovascular disease focus on high-intensity statins and targeting or using a threshold low-density lipoprotein cholesterol (LDL-C) level of less than 70 mg/dL for the highest-risk patients. Whether further reduction of LDL-C beyond these boundaries would be beneficial is unknown. OBJECTIVE To compare outcomes of evolocumab vs placebo in patients with stable atherosclerotic cardiovascular disease and a baseline LDL-C of less than 70 mg/dL and in those receiving background treatment with a maximal-potency statin. DESIGN, SETTING, AND PARTICIPANTS This secondary ad hoc analysis of the Further Cardiovascular Outcomes Research With PCSK9 Inhibition in Subjects With Elevated Risk (FOURIER) trial compared randomized treatments in 2 subgroups of patients with stable atherosclerotic cardiovascular disease currently receiving statin. Patients were classified by a baseline LDL-C of less than 70 or at least 70 mg/dL and by statin intensity (maximal: atorvastatin calcium, 80 mg/d, or rosuvastatin, 40 mg/d; submaximal: all other dosages). Patients with baseline LDL of less than 70 mg/dL either had a final screening LDL-C of at least 70 mg/dL or a final screening non-high-density lipoprotein cholesterol level of at least 100 mg/dL. Datawere retrieved from 2013 to 2016 and analyzed in 2017 based on intention to treat. MAIN OUTCOMES AND MEASURES The primary efficacy end point was the composite of cardiovascular death, myocardial infarction, stroke, hospitalization for unstable angina, or coronary revascularization. The secondary efficacy endpoint was the composite of cardiovascular death, myocardial infarction, or stroke. Safety outcomes included adverse events and events of interest identified in the FOURIER trial. Interaction testing was used to assess the consistency of results in patients who did vs did not satisfy the above criteria. RESULTS A total of 27 564 patients (75.4% men and 24.6% women; mean [SD] age, 62.5 [9.0] years) were included in the analysis. Of 2034 patients (7.4%) who had a baseline LDL-C of less than 70 mg/dL, evolocumab reduced the risk for the primary endpoint (hazard ratio [HR], 0.80; 95% CI, 0.60-1.07) to a similar degree as in the 25 529 patients who had baseline LDL-C of at least 70mg/dL (HR 0.86; 95% CI, 0.79-0.92; P = .65 for interaction; 1 patient was missing baseline LDL-C data). Of 7533 patients (27.3%) receiving maximal-potency statins, evolocumab significantly reduced the primary endpoint (HR, 0.86; 95% CI, 0.75-0.98) to a similar degree as in the 20 031 patients not receiving a maximal-potency statin (HR, 0.85; 95% CI, 0.78-0.93; P = .88 for interaction). The key secondary endpoint was reduced to a similar degree in both analyses. No major safety concerns were identified. CONCLUSIONS AND RELEVANCE Evolocumab was equally effective in reducing cardiovascular events in patients with stable atherosclerotic cardiovascular disease regardless of whether the baseline LDL-C was less than 70 or at least 70 mg/dL and whether the background statin was of maximal or submaximal potency.Wo

Air-Adapted <i>Methanosarcina acetivorans</i> Shows High Methane Production and Develops Resistance against Oxygen Stress

<div><p><i>Methanosarcina acetivorans</i>, considered a strict anaerobic archaeon, was cultured in the presence of 0.4–1% O₂ (atmospheric) for at least 6 months to generate air-adapted cells; further, the biochemical mechanisms developed to deal with O₂ were characterized. Methane production and protein content, as indicators of cell growth, did not change in air-adapted cells respect to cells cultured under anoxia (control cells). In contrast, growth and methane production significantly decreased in control cells exposed for the first time to O₂. Production of reactive oxygen species was 50 times lower in air-adapted cells <i>versus</i> control cells, suggesting enhanced anti-oxidant mechanisms that attenuated the O₂ toxicity. In this regard, (i) the transcripts and activities of superoxide dismutase, catalase and peroxidase significantly increased; and (ii) the thiol-molecules (cysteine + coenzyme M-SH + sulfide) and polyphosphate contents were respectively 2 and 5 times higher in air-adapted cells <i>versus</i> anaerobic-control cells. Long-term cultures (18 days) of air-adapted cells exposed to 2% O₂ exhibited the ability to form biofilms. These data indicate that <i>M. acetivorans</i> develops multiple mechanisms to contend with O₂ and the associated oxidative stress, as also suggested by genome analyses for some methanogens.</p></div

Methane synthesis and oxygen consumption in <i>M</i>. <i>acetivorans</i>.

<p>Cells were cultured in methanol (A, C) or acetate (B, D) and the contents of methane (A, B) and O₂ (C, D) in the head space at the indicated times were determined. Control anaerobic cells (■), control anaerobic cells <i>plus</i> air pulses (●) and air-adapted cells (▲). The O₂ concentration in the head space of culture bottles without cells was also determined (□). The increase in the content of O₂ was due to each new air injection which was indicated by arrows. Values are the mean ± SD of at least 4 different independent cell batches. ^a P<0.05; ^b P <0.01 <i>vs</i> air-adapted cells.</p

Antioxidant enzyme activities.

<p>Cytosolic-enriched fractions from anaerobic control (black bars), anaerobic <i>plus</i> 2% O₂ for 2h (grey bars) and air-adapted cells (white bars) grown on methanol (A) or acetate (B) were used to determine activities of SOD, APX and CAT as described under Methods. Rate units for APX are mU (nmol of substrate consumed (min x mg protein)^-1), whereas CAT and SOD activity units are U (mg protein)^-1 as described under Methods. Values shown are the mean ± SD of at least 4 independent preparations. *P<0.01; ** P<0.05 <i>vs</i> anaerobic control cells. ^a P<0.01 <i>vs</i> methanol-grown cells.</p

Formation of a biofilm extracellular matrix induced by O₂ stress in <i>M</i>. <i>acetivorans</i>.

<p>Samples were prepared for and analyzed by scanning electron microscope as described in the Methods section. Extracellular matrix was absent in cells grown in the absence of oxygen (A) but well-defined in air-adapted cells (B). The carbon source was methanol, which was initially 100 mM and further replenished every 3 days. Micrographs are shown at 5000X, bar = 2 μm. (C) Biofilm formation determined by crystal violet staining in control-grown cells and air adapted cells cultured in methanol. *<i>P</i> < 0.01 <i>versus</i> control cells. (D) Extracellular DNA determination; 1 mg protein of control (filled squares) and air adapted cells (open circles) was added to a quartz cuvette with 1.8 mL of TME buffer. After 60 seconds of baseline acquisition, DNAse I was added and the light pass was determined at 540 nm.</p

Effect of high O₂ concentrations on methane production in <i>M</i>. <i>acetivorans</i>.

<p>Cells were cultured in 100 mL bottles with a syringe inserted in the rubber stopper to allow for gas exchange; the bottles were then incubated At 37°C in an 8 L anaerobic jar under steady 9% O₂ (▲), as described in methods. The methane produced by anaerobic control (■) and air-adapted (○) cells throughout the growth curve was determined. Values are the mean ± SD of 4 bottles contained into the anaerobic jar.</p

Genes annotated coding for proteins involved in oxidative stress protection in methanogens.

<p>Genes annotated coding for proteins involved in oxidative stress protection in methanogens.</p

Cellular responses to O₂ in stationary growth phase in anaerobic control and air-adapted cells.

<p>Cellular responses to O₂ in stationary growth phase in anaerobic control and air-adapted cells.</p

Transcript levels of anti-oxidant genes in <i>M</i>. <i>acetivorans</i>.

<p>Anaerobic control cells grown on acetate and harvested in the stationary phase were incubated at 37°C under orbital shaking in the absence (lane 1) or presence (lane 2) of 2% O₂ for 2 h. Air-adapted cells grown and harvested in the same conditions were also exposed to O₂ (lane 3). mRNA isolation and RT-PCR analysis was carried out by the primer dropping method as indicated in the Methods section and the PCR products separated by gel electrophoresis (A). Densitometric analysis (B) was carried out by double normalization <i>versus</i> the internal control MA3998 transcript and the target genes from anaerobic control cultures without O₂ exposure (lane 1). Values are the mean of 4 independent experiments ± SD.</p