Evaluation of Ubiquinol on Mitochondrial Oxidative Capacity in Statin Patients Using 31PMRS

Synopsis: There are important gaps in our understanding of the pathogenesis and management
of statin-associated myalgia. Data suggest that statins block the production of CoQ10, an
essential component in electron transport during oxidative phosphorylation leading to
mitochondrial dysfunction. Preliminary data suggest that CoQ10 dietary supplements may
improve statin-associated muscle symptoms; however, correlation with plasma CoQ10 and
objective tests to assess statin-associated myalgia and response to treatment are lacking.

Phosphorus-31 magnetic resonance spectroscopy (31P-MRS) is unique in its ability to study,
continuously and non-invasively, the biochemical pathways for the supply and utilization of
energy in muscle. The post-exercise recovery rate of skeletal muscle phosphocreatine (PCr)
is a validated index of mitochondrial oxidative capacity in vivo. In a preliminary study, we
measured PCr recovery kinetics following exercise, using 31P-MRS before and after initiating
statin therapy and found that the return to metabolic equilibrium was longer after 4 weeks
of statin therapy, suggesting mitochondrial impairment. i

It is our primary hypothesis that statin drugs impair skeletal muscle mitochondrial function
and that ubiquinol supplementation will block impairment of PCr recovery kinetics in
patients using statins. We propose a pilot study to extend our research to examine PCr
recovery kinetics in 20 statin users randomized in a parallel arm study to either ubiquinol
or placebo over 4 weeks.

This study will include 10 individuals with statin myalgia on confirmed statin rechallenge
(5 assigned to each arm), to be recruited from an ongoing clinical trial in which statin
users with suspected statin myalgia (based on clinical history) undergo an observed statin
re-challenge to confirm statin myalgia. Confirmation by re-challenge is important, as only
about one-third to one-half of those who report a clinical history of statin myalgia
demonstrate statin myalgia, confirmed by improvement of symptoms when statin is discontinued
and return of symptoms when undergoing an observed statin re-challenge. ii

Our team is unique in 1) having demonstrated the feasibility to perform 31P-MRS studies in
patients with hypercholesterolemia and 2) having access to individuals that have proven
statin myalgia on standardized observed statin-rechallenge. We believe that the data
obtained from this study will provide the critical data needed to support application to the
NIH for a full-scale clinical trial exploring the relationship between statin use, PCr
recovery rate, and ubiquinol therapy.

Aims:

1. o Aim: To compare the effects of ubiqunol on post exercise PCr recovery kinetics.

We hypothesize that statin users randomized to 300 mg of ubiquinol twice daily will
demonstrate shorter PCr recovery kinetics following exercise compared to statin users
randomized to placebo.

2. o Aims: To assess the association between plasma CoQ10 and 1) post exercise PCr
recovery kinetics and 2) inflammatory and vascular biomarkers We hypothesize that lower
plasma CoQ10 concentrations will be associated with longer PCr recovery kinetics and
lower inflammatory/vascular biomarkers.

Exploratory Aims: In each arm we aim to include 5 individuals with previously confirmed
statin myalgia on statin rechallenge and to explore differences among statin users with and
without statin myalgia. We will evaluate for trends to assess whether statin users with
previously confirmed statin myalgia show longer PCr recovery kinetics compared to statin
users without statin myalgia.

Study Design: A pilot randomized placebo-controlled double-blinded study (pre-post statin
use) comparing the effects of ubiquinol on post exercise PCr recovery kinetics over 4 weeks.

Background and Rationale Statins, HMG-CoA reductase inhibitors are generally safe; however,
myalgia (e.g., pain, cramping, muscle soreness and/or weakness) occurs in about 5% of users
iii,iv and can seriously impact quality of life and adherence to statin medications.
Furthermore, there are no objective, non-invasive method for assessing and tracking
mild-moderate muscle symptoms, which are most common and often occur without significant
muscle enzyme "CK" elevation. Muscle biopsy is used to assess severe myopathy and has been
used to evaluate statin myalgia without CK elevation, but is invasive, painful, and provides
only a single time point measurement.

In contrast to invasive muscle biopsy, 31P-MRS is a non-invasive and validated technique
that has been applied to study the metabolic processes in skeletal muscle in many muscle
applications and diseases.v,vi The rate of PCr recovery following exercise is an accepted
gold standard for measuring mitochondrial function.

We propose to apply 31P-MRS to the important problem of statin-associated myalgia to measure
the rate of PCr recovery following exercise in statin users before and after statin therapy
(with concurrent randomization to ubiquinol or placebo). Ultimately, we believe this work
could have an important impact on the care and management of those with statin myalgia by 1)
elucidating the relationship between muscle symptoms and mitochondrial function through a
quantifiable, validated and non-invasive biochemical index and 2) by determining the effects
of supplemental ubiquinol on mitochondrial function and muscle side effects in statin users.

Methods We will measure the rate of PCr resynthesis (the time constant of PCr recovery)
following a controlled exercise protocol that will recruit muscles in the lower leg. We
will evaluate muscle symptoms using a standardized questionnaire and measure muscle strength
using an ergometer that was designed to apply a controlled variable pressure to a foot pedal
through a pneumatic cylinder. We will perform baseline measurements after participants have
been taken off statin medications and CoQ10 supplements (if applicable) for at least 2
weeks. Additionally, those with statin myalgia will have improvement of symptoms before
baseline measures are taken. Following baseline testing, statin therapy+ubiquinol or statin
therapy+placebo will be given to participants for 4 weeks. Repeat measurements will be
performed at 4 weeks to compare the effects of statins+placebo versus statin+ubiquinol on
PCr resynthesis. The primary outcome of this study will be the PCr recovery time constant.
The post-exercise PCr recovery measurements will be performed in our MRI research facility
using our dedicated 3T research MR scanner. Assessment of safety outcomes (CK, AST, ALT),
and physiological parameters (BP, HR, height and weight measured), and blood measures of
lipids and biomarkers (CRP, IL-6, TNF- α, E-selectin, VCAM-1, ICAM-1) will be collected on
site and analyzed at the Harvard Catalyst Clinical Research Center (CRC) Core laboratory.
Analysis of reduced and oxidized plasma CoQ10 will be performed at a qualified outside lab
with a strong track record of using validated methods.

Data Analysis Statistical Analysis: Each subject will have PCr concentration collected at
the following 2 time points: pre-statin (t1), and after 4-weeks of statin therapy (with
ubiquinol or placebo) (t2). At each of these time points, a series of measurements of PCr
concentration are collected. For each subject, we will estimate this parameter using maximum
likelihood estimation method. Thus we will obtain the point estimate and standard error for
the time constant parameter for each subject at each of the time points. We will compute the
relative percent change from baseline to post-statin (ubiquinol vs. placebo) time points and
a linear mixed effects model to obtain the estimate of the relative change from baseline and
the relative change between the post-statin time point. The linear mixed effects model, with
the compound symmetry structure for the variance-covariance matrix of the within-subject PCr
relative changes from baseline takes into account the within-subject correlation. The
estimation of the linear mixed-effects model will be done via restricted maximum likelihood
method. The PCr recovery time constant will be calculated using a monoexponential fit of PCr
versus time, beginning at exercise completion and will be compared between the groups.

Additional analyses will be performed to assess associations between plasma CoQ10 (adjusted
for LDL) and PCr data, inflammatory and vascular biomarkers, measured muscle strength (in
all), and reported symptoms (among those with statin myalgia).

Inclusion Criteria:

1. Persons age 18 years and older who have been advised to use a statin medication by
their physician.

2. Up to ten individuals will have a history of statin myalgia on observed statin
rechallenge. All others will have a history (>6 months) of tolerating a statin at a
standard starting dose or higher.

3. All persons will be able to cooperate for a period of 1-2 hours, able to provided
informed consent and have no known adverse effects or contraindications to an MRI
study.

Exclusion Criteria:

1. Having diabetes or known lower extremity peripheral vascular disease, as these
conditions may interfere with interpretation of the MRI spectroscopy of the lower
extremity (the only site we are examining in this study).

2. Current users of a statin with a recent heart attack, stroke, or revascularization
procedure (within the last 1 year) who could be at higher risk of stopping a statin.

3. Creatine kinase (CK) > 5 x ULN. If CK levels are greater than 3 x ULN, the
participant will be given the option of returning for a repeat blood draw at their
earliest convenience. Any elevation in CK will be treated per the safety guidelines
outlined in section B5.

4. Alanine transferase (ALT) or aspartate transferase (AST) > 3 x ULN. If ALT and/or
AST levels are greater than 3 x ULN, the participant will return for a repeat blood
draw at their earliest convenience. If the participant's ALT and/or AST is found
elevated at this repeat analysis, they will be withdrawn from the study. Any
elevation in ALT and/or AST will be treated per the safety guidelines outlined in
section B5.

5. Severe uncontrolled medical problems or medications that may influence measurements
or impair ability to participate in the study exams (e.g. poorly controlled
hypertension; (>170/>100); known creatinine > 2.5 md/dl or GFR < 30; anemia with Hgb
< 10, etc.).

6. Pregnancy or breastfeeding (a contraindication for statin use)