Clinical trial NCT04594733
Conditions: Fibromyalgia
Sponsor: Medical University of South Carolina
Start date: 2021-12-06; Completion date: 2022-08-31
Enrollment: 14 participants; Allocation: RANDOMIZED; Intervention model: CROSSOVER

Brief summary:
Evaluation of the effect of combinatorial treatment of fibromyalgia patients (standard of care + minocycline + N-acetylcysteine (NAC) relative to standard of care on subjective pain measurement Revised Fibromyalgia Impact Questionnaire (FIQR).

Detailed description:

Arms: arm 1: Participants will be randomized to 8 weeks of placebo, minimum 2 week washout period, and then 8 weeks of a combination minocycline (200 mg daily) and NAC (1200 mg daily) followed by a final 2 week washout arm 2: Participants will be randomized to 8 weeks of a combination minocycline (200 mg daily) and NAC (1200 mg daily), minimum 2 week washout period, and then 8 weeks of placebo followed by a final 2 week washout
Interventions: intervention 1: minocycline (200 mg daily) and NAC (1200 mg daily) followed by placebo intervention 2: Placebo followed by minocycline (200 mg daily) and NAC (1200 mg daily)
Intervention descriptions: intervention 1: minocycline (200 mg daily) and NAC (1200 mg daily) followed by placebo intervention 2: Placebo followed by minocycline (200 mg daily) and NAC (1200 mg daily)

Protocol documents:
Prot_SAP_000.pdf: 
 
 
 
The effect of multi-modal minocycline and N-acetylcysteine for the treatment of 
fibromyalgia: a double-blind, randomized, crossover pilot study. 
 
Study Protocol 
Statistical Analysis Plan 
 
NCT # NCT04594733 
 
12-2-2021 


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PROTOCOL TITLE: 
The effect of multi-modal minocycline and N-acetylcysteine for the treatment of 
fibromyalgia: a double-blind, randomized, crossover pilot study. 
 
PRINCIPAL INVESTIGATOR: 
M. Gabriel Hillegass, III, M.D. 
 
1.0 
Objectives / Specific Aims 
Our objective is to design and test treatments for chronic pain that target multiple non-
neuronal cell types in the clinical setting. 
 
The forefront of the science of pain inhibition, and the most fruitful area of research for drug 
development, is the complete understanding of the molecular mechanisms that govern pain 
sensation. Towards that end, gaining a better understanding of the physiological 
mechanisms linked to pain signaling, like inflammation,1,2 are of the utmost importance. In 
the fields of cellular and molecular neuroscience, it has been demonstrated that the action 
and signaling of non-neuronal cells, astroglia and microglia, serve as potent regulators of 
neuronal communication.3-5 Over the last decade, preclinical scientists have begun to 
incorporate bidirectional communication between neurons and neighboring non-neuronal 
cells into their working models of synaptic plasticity.6,7 Recently, activation of microglial 
cells, and the release of pro-inflammatory cytokines like tumor necrosis factor alpha (TNF) 
and Interleukin 1 (IL-1) have been directly linked to the induction of pain sensation and 
the expression of pain-related phenotypes including hyperalgesia and allodynia.8 Further, 
microglia have been linked to the mechanisms that govern chronic pain.9-12 In addition, 
injury-related decreases in astrocyte-mediated glutamate clearance have also been shown 
to contribute to elevated pain responses following nerve injury.13 
 
Our central hypothesis is that a multimodal approach, designed to inhibit microglial 
activation and promote healthy homeostatic regulation of glutamate by astrocytes, will 
provide a therapeutic advantage in treating pain and other symptoms related to 
fibromyalgia. Enticingly, two FDA approved compounds exist that can be used to 
implement this precise multi-modal approach, minocycline, a tetracycline antibiotic and 
inhibitor of microglia14,15 and N-acetylcysteine (NAC), a powerful antioxidant shown to 
promote homeostatic glutamate regulation by astrocytes.16,17 This proposal aims to 
address the current lack of evidence of both classes of non-neuronal cells to alleviate pain. 
We posit that our multimodal approach will lessen the severity of chronic pain and enhance 
efficacy and implementation of traditional methods for inhibiting pain in patients. This pilot 
study will assess the feasibility of this approach and provide important data for refining this 
research. 
 
2.0 
Background 
Microglia, pain signaling and pain sensitivity  
Microglia are a subset of neuroglia or non-neuronal cells, located throughout the central 
and peripheral nervous system. Microglia act as the resident macrophages of nervous 
tissue, recognizing and responding to external pathogens by engaging the 
neuroinflammatory response. When microglia transition from a quiescent or resting state, 
to a state associated with the release of pro-inflammatory neuromodulators, they are said 
to be ‘active’, or ‘activated’. Remarkably, microglia are incredibly structurally plastic and 


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their activation states are associated with specific alterations in cellular architecture and 
the dimensions of particular cellular compartments.18-20 
 
Recent discoveries have illuminated that microglia and microglia activation are central in 
engaging the cellular signaling responsible for the sensation of pain.8,21 Specifically, spinal 
microglia are strongly activated after peripheral nerve injury,22,23 resulting in elevated pro-
inflammatory cytokine release in the central and peripheral nervous system.24 Microglia are 
activated in response to pain,22,25 and are directly implicated in pain, as microglia respond 
to injury by becoming more sensitive to future insults by upregulating cell surface receptors 
and secreted pro-inflammatory molecules,26 including TNF.27 In rodent models, inhibition 
of microglia activation following nerve injury with the antibiotic minocycline28 has been 
shown to reduce the severity of nociception-related phenotypes.29-32 Mechanistically, 
minocycline inhibits or attenuates microglial activation by altering the program of cytokine 
release,33-35 countering pro-inflammatory signaling.36  Outside of microglia, minocycline 
also reduces expression of receptors for TNF in nearby neurons, limiting the ability of 
microglia to engage TNF-mediated neuroinflammatory signaling and synaptic scaling.37 
Importantly, minocycline treatment also reverses the reorganization of microglia 
ultrastructure occurring concomitantly with activation.15,38-40 These data support the 
efficacy of minocycline in attenuating microglia activation and expression of pain-related 
phenotypes, while underscoring the well-defined structure-function relationship for this cell 
type.41-43 
 
Coordinated non-neuronal cell regulation of the cellular signaling underlying 
chronic pain  
Microglia and astroglia serve as potent regulators of neuronal communication.3-5 However, 
the concomitant bidirectional chemical communication between these three cell types has 
only recently begun to be been incorporated into working models of chronic pain.6,7,44  
The non-neuronal and neuronal signaling mechanisms in chronic pain are illustrated in 
Figure 1.  
Figure 1: Illustration of microglia and astroglial components of chronic pain, and minocycline and N-Aceytlcysteine treatment. A) Disease state at the 
dorsal horn spinal cord synapse. 1) chronic pain causes activation of microglia. 2) Activation of microglia engages TNF (red spheres) and IL1 (orange 
sphere) release. 3) TNF and IL1 engages postsynaptic glutamate receptor insertion in neurons. 4) TNF and IL1 engage GLT-1 downregulation in 
astroctyes. 5) Sensory stimuli then evoke, 6) a potentiated glutamate release (grey spheres) due to the presynaptic action of TNF. 7) Glutamatergic 
communication, and pain sensation is evoked and enhanced due to strengthening of this synapse. B) The dorsal horn spinal cord synapse following 
treatment. 1) Treatment. Minocycline inhibits persistent microglia activation. 2) this limits TNF and IL1 release. 3) Reduced TNF and IL1 release 
limits postsynaptic strengthening, and the negative regulation of GLT-1 expression. 4) The NAC component of the treatment restores GLT-1 function 
and expression. Allowing for removal of glutamate from the cleft and termination of signaling. 5) Following treatment, sensory stimuli should no longer 
provoke an exaggerated glutamatergic response due to the reduced levels of TNF. 7) Finally, following treatment, allodynia, hyperalgesia and other 
pain related phenotypes will be reduced / eliminated. 


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During nerve injury or chronic pain, microglial activation engages a TNF-dependent 
enhancement of glutamatergic synaptic communication with a multimodal molecular 
mechanism.45,46 In neurons, TNF engages TNF receptors, resulting in enhanced 
glutamate release and insertion of glutamate receptors, respectively.47,48 Within the 
astrocyte itself, TNF receptor activation acts to down-regulate the astroglial glutamate 
transporter, required for the termination of sensory synaptic signaling.48 Our overarching 
hypothesis is that our combined non-neuronal cell targeted treatment will reverse this 
maladaptive plasticity by both suppressing microglial activation, and restoring astroglial 
glutamate homeostasis, thus providing analgesia and potentially other symptoms of 
fibromyalgia. 
 
3.0 
Intervention to be studied  
Recent research demonstrates microglia are crucial mediators of 
neuroinflammation and central sensitization, which are both mechanisms that 
underlie the altered synaptic plasticity responsible for chronic widespread pain.49 
Another microglial inhibitor, naltrexone, has been found to reduce fibromyalgia 
associated symptoms.50 While N-acetylcysteine has not been studied clinically in 
patients suffering from chronic pain, it has been successfully used to treat types of 
addiction and compulsivity.51  
• 
Minocycline is FDA approved. Typical dosing is 100 mg every 12 hours. It is 
prescribed chronically for certain latent bacterial infections as well as for 
adjunctive therapy in severe acne vulgaris. Tetracyclines may render oral 
contraceptives less effective. Like other tetracycline antibiotics, minocycline 
crosses the placenta and can adversely affect skeletal development and may 
have other embryotoxic effects. It is also excreted in breast milk and can 
permanently discolor developing teeth. Photosensitivity may result in 
exaggerated sunburn reactions, although minocycline is less likely to produce 
this reaction than other tetracyclines. Sunscreen is not protective from this 
reaction. CNS side effects may include light-headedness, dizziness, or 
vertigo, which may dissipate during therapy. These symptoms could affect 
driving or the operation of heavy machinery. Pseudotumor cerebri (benign 
intracranial hypertension) has been associated with tetracyclines. Alteration 
of the normal gut flora may cause diarrhea or even predispose for the 
development of Clostridium difficile associate diarrhea and related 
complications, which could be mild or as severe as causing fatal colitis if not 
treated. This potential complication has been reported with nearly all 
antibiotics. It may also result in the overgrowth of fungi. If minocycline 
becomes stuck in the esophagus it could cause esophageal irritation and 
ulceration. Hypersensitivity reactions that included, but were not limited to 
anaphylaxis, angioedema, urticaria, rash, swelling of the face, and pruritus 
have been reported. Postmarketing cases of anaphylaxis and serious rash 
such as exfoliative dermatitis, Stevens-Johnson syndrome, and erythema 
multiforme have been reported with minocycline. Postmarketing reports have 
included cases of severe hypersensitivity reactions and autoimmune 
syndromes including a lupus-like syndrome. Drug Rash with Eosinophilia and 
Systemic Symptoms (DRESS), including fatal cases, have been reported with 
minocycline use. Hepatotoxicity has been reported with minocycline; 
therefore, minocycline should be used with caution in patients with hepatic 


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disease and in conjunction with other hepatotoxic drugs. Hepatitis, including 
autoimmune hepatitis, and liver failure have been reported. The antianabolic 
action of the tetracyclines may cause an increase in BUN. While this is not a 
problem in those with normal renal function, in patients with significantly 
impaired function, higher serum concentrations of minocycline or other 
tetracyclines may lead to azotemia, hyperphosphatemia, and acidosis. 
• 
N-acetylcysteine (NAC) is FDA approved in some formulations and 
considered a nutritional supplement in others. We will be utilizing pure 
powder of NAC that will have a Certificate of Analysis. The CoA confirms the 
chemical powder has the potency labeled, the pH, whether or not it contains 
heavy metals, the particle size and density, that is it free from bacteria, mold 
or fungus, and the physical description and appearance of the product. An 
Investigation New Drug (IND) application will be submitted to the FDA and 
approval will be required prior to initiation of this study. NAC was introduced 
in the 1960s as a mucolytic drug for chronic respiratory diseases (e.g., 
COPD, tuberculosis, bronchiectasis, etc.). It is also used for nephrotoxicity 
prophylaxis against radiographic-contrast-induced reactions in patients with 
preexisting renal insufficiency. It has a well-established safety profile and is 
still commonly used orally at doses of 600 mg/day as a mucolytic. However, 
at higher doses (≥1200mg), acetylcysteine also acts as an antioxidant 
through complex mechanisms which can combat conditions of oxidative 
stress. Acetylcysteine is a derivative of the natural amino acid cysteine, which 
serves as a substrate for the synthesis of glutathione (GSH) in the body 
which an antioxidant effect. This reduces the formation of proinflammatory 
cytokines, such as IL-9 and TNF-α and also has vasodilator properties by 
increasing cyclic GMP levels and by contributing to the regeneration of 
endothelial-derived relaxing factor. 
• 
The control/placebo will contain only microcrystalline cellulose, which is 
refined wood pulp. Chemically, it is an inert substance, is not degraded 
during digestion and has no appreciable absorption. This has been used by 
our collaborating pharmacist in previous NAC studies, with no issues or 
problems reported, however large quantities provide dietary bulk and may 
lead to a laxative effect. This adverse effect may be seen with a daily dose of 
30 grams of M-cellulose. The placebo for the NAC capsule will contain about 
300 mg of M-cellulose per capsule, and the placebo for the minocycline 
capsule will contain about 100 mg of M-cellulose per capsule. The empty 
placebo capsules will be exposed to a “NAC bomb” prior to being filled so 
that they smell similar to the active drug. A “NAC bomb” is essentially 
powdered NAC inside of a panty hose that prevents the medication from 
escaping but allows the empty placebo capsules to take up the rotten egg 
smell. 
• 
There will be two different sized capsules used: 600 mg NAC (or placebo) will 
be in a #1 size capsule and 100 mg of minocycline (or placebo) will fit into a 
smaller #3 size capsule. Blister packs will be utilized to improve compliance. 
These will fit both a #1 and #3 capsule in a single blister pack and be labeled 
“Take both capsules every 12 hours”. 
 
4.0 
Study Endpoints  


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• Primary Outcome: Change in the Revised Fibromyalgia Impact Questionnaire (FIQR) 
scores. 21 questions; 3 domains = function, overall impact, symptoms; 11-point 
numeric rating scale; questions framed with the context of the past 7 days. Measured 
weekly (48-hour response time with reminders every 12 hours until lockout occurs at 
48 hours past due). 
Secondary Outcomes: (1) Patient Global Impression of Change (PGIC)56: Likert scale of 
patient's rating of overall improvement. The PGIC and the modified 2010 American 
College of Rheumatology Preliminary Diagnostic Criteria for Fibromyalgia 
(modified ACR 2010)57 questionnaire will be utilized at baseline and conclusion of 
each treatment period. 
Other Data Collected: (2) “Pain on average”, “interference with Enjoyment of life”, 
“interference with General activity” (PEG)53; (3) Patient Health Questionnaire-2 
(PHQ-2)54: 2 question depression severity measure; (4) Generalized Anxiety 
Disorder scale (GAD-2)55: 2 question anxiety severity measure; The PEG will be 
measured at baseline and weekly. The PHQ-2 and GAD-2 will be measured at 
baseline & biweekly during treatment phases & washout periods.  
5.0 
Inclusion and Exclusion Criteria/ Study Population 
The PI and coinvestigators will screen patients based on their pre-existing 
diagnosis of fibromyalgia. A chart review will also be performed to ensure the 
subject meets the inclusion and exclusion criteria.  
Inclusion Criteria  
• Female 
• Age 18 years or older 
• Formal diagnosis of fibromyalgia and at least 6 months of widespread 
pain (confirmed by the modified 2010 ACR Diagnostic Criteria for 
Fibromyalgia Widespread Pain Inventory and Symptom Severity 
Scale) 
• Currently on birth control or unable to become pregnant 
• Willingness to avoid taking opioid and opiate medications during the 
duration of the study (5-6 months) 
Exclusion Criteria 
• Known hypersensitivity to minocycline or tetracycline antibiotics or to 
N-acetylcysteine 
• Unwillingness to pause opioid therapy thirty days prior to starting the 
study and during the duration of the study  
• Active pregnancy, lactation or plans to become pregnant in the next 
6 months 
• Significant hepatic disease as indicated by an AST or ALT greater 
than twice the upper limits of normal or bilirubin greater than twice 
the upper limits of normal at baseline 
• Significant renal disease as indicated by an estimated glomerular 
filtration rate less than 60 mL/min/1.73m2 at baseline or during the 
first washout period 
• History of autoimmune syndromes (systemic lupus erythematosus, 
myasthenia gravis, rheumatoid arthritis) 
• History of intracranial hypertension or pseudotumor cerebri 
• History of IBD (Crohns disease, ulcerative colitis), Clostridium difficile 
infection 


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• History of esophagitis, esophageal obstruction, achalasia or 
esophageal dysmotility 
• History of GI hemorrhage or known risk factors for GI hemorrhage, 
such as esophageal varices, peptic ulcer disease, etc. 
• Unwillingness to limit intake of divalent and trivalent cations such as 
oral iron supplements, certain dietary supplements (multivitamins) 
that contain manganese or zinc, or antacids that contain aluminum, 
calcium, or magnesium, for at least three hours before and after 
taking study drugs, which would decrease minocycline absorption 
• Subjects taking anticoagulant medication since minocycline can 
decrease plasma prothrombin activity 
• Subjects taking isotretinoin 
• Subject taking ergot alkaloids for migraines 
• Subjects taking penicillin antibiotics 
• Subjects who work outdoors or otherwise have prolonged exposure 
to UV light and sunlight 
• Lack of access to reliable technology to be able to complete emailed 
REDCap questionnaires 
• Cognitive deficits that may make it difficult to adhere to the 
medication regimen or provide consistent and timely completion of 
questionnaires 
• Inability or unwillingness to give informed consent 
• Unwillingness to use two forms of birth control for the entire duration 
of participation in the study (if capable of becoming pregnant) 
• Unwillingness to complete home pregnancy tests throughout the 
study (if capable of becoming pregnant) 
• Inability to swallow large pills 
 
6.0 
Number of Subjects 
14 subjects recruited with the goal of 12 patients completing the study 
 
7.0 
Setting 
The protocol will mostly be conducted remotely (telephone, video conference, text 
or email), however three in-person lab visits will be required to obtain blood draws 
for a comprehensive metabolic panel and a urine sample for pregnancy testing (if 
applicable) at baseline and during each washout period. The baseline study visit 
appointment will require an in person visit with study personnel for consenting, 
which will be done electronically using the REDCap eConsent platform, and 
enrollment, distribution of study meds and administration of study instruments. 
Subjects will receive all 8 weeks of study medication at the start of each treatment 
period. Subjects will complete all measures via REDCap surveys that will be 
delivered electronically to them via email with a secure link to REDCap or the 
REDCap application.  
 
8.0 
Recruitment Methods 
Subjects will be recruited through the MUSC Pain Clinic by the PI and coinvestigators, 
but some patients may be referred to the study from MUSC colleagues who routinely 
collaborate with the Pain Clinic staff.  
 


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The flyer will also be mailed, emailed and sent via mychart message to potential 
participants diagnosed with fibromyalgia utilizing MUSC’s patient outreach recruitment 
strategies.  The communications will include the required direct contact information letter. 
Doing this will allow us to be inclusive of all potential subjects diagnosed with 
fibromyalgia since they are seen in various clinics that we may not have access to. 
 
Placement in flyers in community areas such as pharmacies, MUSC clinical areas 
(waiting rooms and bulletin boards). 
 
Those who express interest in participation will be provided a phone number and email to 
contact our research support staff who will then provide additional information and a pre-
screening survey. A chart review of each potential participant who completes the 
screening survey will be conducted to ensure that no exclusionary criteria are indeed 
present. 
 
9.0 
Consent Process 
Informed consent will occur in person during the initial screening visit. At this visit, 
participants will give consent electronically by using the REDCap eConsent 
platform. Potential participants will be given time to read the consent. Adequate 
time will be taken to review the consent and HIPAA documents with the subject 
and answer all questions. 
 
10.0 Study Design / Methods 
• A double-blind, randomized, crossover design will be used. Fourteen female 
fibromyalgia patients will be recruited with a goal of 12 patients completing the study. 
This pilot study will only include women, which reflects the affected population with 
most diagnosed cases being in women. Population based studies have shown lifetime 
prevalence to be 2:1 for women compared to men. This study seeks to assess the 
multimodal effects of the intervention in the most at-risk population. 
• Prior to enrollment, a urine sample for a pregnancy test (if appropriate) and a blood 
draw for a comprehensive metabolic panel (CMP) will be assessed. This lab work will 
be completed at a MUSC lab and will not be billed to the patient. If the potential 
participant is pregnant or has evidence of significant kidney or liver dysfunction they 
will not be enrolled. 
• A simple randomization method will be used to assign the initial treatment and then 
the opposite treatment will ensue for the second treatment period. Patients will be 
randomized to (i) 8 weeks of placebo minocycline and placebo NAC, minimum 2 week 
washout period, and then 8 weeks of a combination minocycline (200 mg daily) and 
NAC (1200 mg daily) followed by a final 2 week washout or (ii) 8 weeks of a 
combination minocycline (200 mg daily) and NAC (1200 mg daily), minimum 2 week 
washout period, and then 8 weeks of placebo minocycline and placebo NAC followed 
by a final 2 week washout. The washout period is intended to capture any post-
treatment changes in the primary and secondary outcomes following the conclusion of 
each active treatment phase. Treatment will be in addition to standard of care. Our 
primary outcome is change in the Revised Fibromyalgia Impact Questionnaire (FIQR) 
score52. Secondary clinical outcomes include subjective measurements of pain and 
interference (PEG)53, depression (PHQ-2)54, anxiety (GAD-2)55, patient global 
impression of change (PGIC)56. The modified 2010 American College of 


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Rheumatology Preliminary Diagnostic Criteria for Fibromyalgia (modified ACR 2010)57 
questionnaire will be utilized at baseline and conclusion of each treatment period.  
• Subjective pain measurements: Validated psychometric assessments will be utilized 
and delivered directly to the patient’s email for response via the REDcap survey 
instrument. 
o Primary Outcome: Change in the Revised Fibromyalgia Impact Questionnaire 
(FIQR). 21 questions; 3 domains = function, overall impact, symptoms; 11-point 
numeric rating scale; questions framed with the context of the past 7 days. 
Measured weekly (48-hour response time with reminders every 12 hours until 
lockout occurs at 48 hours past due). 
o Secondary Outcomes: Impression of Change (PGIC)56: Likert scale of patient's 
rating of overall improvement.    
o Other Collected Date: (1) “Pain on average”, “interference with Enjoyment of life”, 
“interference with General activity” (PEG)53; (2) Patient Health Questionnaire-2 
(PHQ-2)54: 2 question depression severity measure; (3) Generalized Anxiety 
Disorder scale (GAD-2)55: 2 question anxiety severity measure; (4) Patient Global 
The PEG will be measured at baseline and weekly. The PHQ-2 and GAD-2 will be 
measured at baseline & biweekly during treatment phases & washout periods. The 
PGIC and modified ACR 2010 will be measured at the end of each treatment phase. 
 
 
Clinical Procedure 
Virtual/Phone 
Screening 
Visit 
Pre-
enrollment 
Labs 
In-person 
Baseline Visit 
& Enrollment 
Treatment 
Period 1  
(8 weeks) 
1st Washout 
(2 weeks)  
In-person 
Visit 
Treatment 
Period 2 
(8 weeks) 
 2nd 
Washout 
In-person 
Visit 
 
Screening 
Inclusion/Exclusion  
X 
 
 
 
 
 
 
Medical 
History/Chart 
Review 
 
 X 
 
 
 
 
 
 
Informed Consent 
 
 
 
X 
 
 
 
 
Fibromyalgia 
Diagnosis (ACR) 
X emailed to 
potential 
participant 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
Randomization 
 
 
X 
 
 
 
 
Study drug 
administration 
 
 
Provided by 
study staff 
according to 
randomization 
 
Provided by 
study staff 
according to 
randomization 
 
 
CMP 
 
X 
 
 
X 
 
X 
Urine HCG 
 
X 
 
 
X 
 
X 
FIQR & PEG 
 
 
X 
Weekly 
Weekly 
Weekly 
Weekly 
PHQ-2 & GAD-2 
 
 
X 
Biweekly 
Biweekly 
Biweekly 
Biweekly 
PGIC & ACR 
 
 
 
End of 
treatment 
 
End of 
treatment 
 
Telephone call to 
assess medication 
side effects 
 
 
 
End of 1st 
week 
 
End of 1st 
week 
 
Adverse event 
assessment & 
recording 
 
 
 
Weekly 
Weekly 
Weekly 
Weekly 
 
 


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Analysis Plan: The primary goal is to estimate the effect of multimodal treatment of 
fibromyalgia patients (standard of care + minocycline + NAC) relative to standard of care on 
subjective pain measurement (FIQR). All outcomes will be evaluated using a linear mixed model 
(LMM) approach.  Models will include fixed effects for treatment regimen, time, and a treatment 
by time interaction as well as a random subject effect to account for repeated measures on 
subjects.  Differences within and between treatment regimens will be estimated using a series of 
linear contrasts from the LMM.  We will enroll 14 subjects to allow for up to 14% loss to follow-
up.  A sample size of 12 subjects provides a two-sided 95% confidence interval with a distance 
from the mean paired difference of ± 0.64 standard deviations.  The analysis will be conducted 
as an intent-to-treat analysis. 
 
 
11.0 Data Management  
• Study-related records will be kept in the research office of the Department of 
Anesthesia & Perioperative Medicine, which is a physically secure location 
(locked office door and locked file cabinets). The REDCap survey instruments 
and associated data will be stored within the secure MUSC network. All study 
personnel will conduct themselves with integrity and work to maintain 
confidentiality at all times. All personnel will have current CITI certifications.  
• Quality control will be ensured as all data will be collected utilizing the same 
REDCap survey instruments. 
 
 
12.0 Provisions to Monitor the Data to Ensure the Safety of Subjects 
• The study will be reviewed annually by the Department of Anesthesia’s Data Safety 
Monitoring Board (DSMB). Minutes and outcomes from these meetings will be reported 
to the IRB as required. 
• Adverse events will be recorded, reviewed and reported to the Department of 
Anesthesia’s DSMB and the IRB per policy. 
• PHI will be managed in a manner that complies with institutional rules and regulations. 
There will be an enrollment log that links the study ID number to the patient. This log 
will be kept on a MUSC password protected server that can only be accessed by IRB 
approved study personnel.  
• Any side effects from the study will be evaluated by the principal investigator or co-
investigators. The research nurse will unblind investigators if necessary, to further 
assess side effects or other study-related adverse events. 
• In the event that a woman becomes pregnant during her study participation she will 
be followed until resolution of the pregnancy.  Any fetal anomalies will need to be 
submitted to the IRB (and FDA) as SAEs. 
• The weekly electronic surveys are very detailed with respect to assessing for adverse 
drug reactions and worsening symptoms. Should the patient report “NEW or 
WORSENING symptoms and/or medication side effects over the past week” in the 
“mild” range, a member of the research team will contact the patient to get more 
information. A virtual office visit with the PI or one of the co-I’s will be offered if desired 
by the participant. If the severity is rated as “moderate” or “severe”, a member of the 
research team will contact the patient to get more information and to coordinate a 
virtual or in-person office visit with the PI or one of the Co-I’s. 
• Regarding depressed patients, we anticipate that comorbid depression will be present 
in some participants as this is commonly associated with FMS. If a participant’s 


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depression score (PHQ-2) is worsening a member of the research team will contact 
the patient to get more information and to coordinate a virtual office visit with the PI or 
one of the co-I’s. The investigators will assess the patient and determine the 
appropriate treatment plan and whether or not the participant should remain in the 
study. 
 
13.0 Withdrawal of Subjects 
• The following circumstances could result in subject withdrawal: 
o Development of any significant adverse drug effects 
o Positive test for COVID-19 
• Subjects may be terminated by the investigator for the following: 
o Failure to submit two consecutive weekly questionnaires or more than three 
weekly questionnaires over an 8-week treatment phase 
o Poor adherence to taking the study medication as instructed 
• Subjects who voluntarily withdraw will no longer receive their weekly REDCap 
assessments and be instructed to stop taking the study medication and discard it 
immediately. Their data will remain for analysis as part of our intention-to-treat. 
 
14.0 Risks to Subjects 
• Adverse medication effects could cause subjects to stop the study prematurely. 
These are listed above. All subjects will be monitored for these events as they 
will be asked weekly about any new or worsening symptoms and side effects. 
Common adverse drug effects are generally mild/low risk. 
• Relying on email-generated survey instruments for data collection could be a 
pitfall if subjects experience local internet or power outages or other barriers 
that prevent timely completion of the assessments. This could also be 
psychologically stressful for subjects if there are technical problems with 
completing the questionnaires. 
• The study drug minocycline may have risks to an embryo or fetus should the 
subject be or become pregnant. We will require that subjects who have the 
capability of becoming pregnant take a pregnancy test at baseline, at their 
appointment during the first washout period and at the end of the study  
• Loss of confidentiality is a risk with any clinical research. 
• The study may have risks to the subjects that are currently unforeseeable.  
• The study is highly unlikely to put others at risk who are not study subjects. 
 
15.0 Potential Benefits to Subjects or Others 
• Subjects may experience decreased pain and improvement in symptoms 
related to their fibromyalgia (fatigue, mood, cognition, etc.) 
• There may be no direct benefit during the placebo phase or treatment phase of 
the study. 
 
16.0 Sharing of Results with Subjects 
We do not intend to share the results of the study with subjects outside of allowing 
them to read the abstract and manuscript if desired. Should a subject request their 
FIQR scores or any secondary outcome results, we’ll supply that information on a 
case-by-case basis. In order to maintain blinding we would not provide this 
information until the conclusion of the study or completion of subject disenrollment. 
 


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17.0 Drugs  
Dispensing of all study-related medication (both active drugs and placebo) will 
occur by our study personnel during the baseline visit for the first treatment phase 
and initial washout visit for the second treatment phase. Pitt Street Pharmacy will 
appropriately compound our subjects’ medication for each treatment period 
according the randomization allocation provided by our statistician. The PI, study 
personnel distributing the medication nor the subjects will know if they are 
receiving active drug or placebo in order to maintain blinding. Social distancing, 
face coverings, and other standard precautions in accordance to MUSC Policy and 
CDC guidance for COVID-19 will be employed. Proper storage instructions for the 
medication will be included with receipt of the medication. 
 
 
 


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Statistical Analysis Plan: The primary goal is to estimate the effect of combinatorial treatment 
of fibromyalgia patients (standard of care + minocycline + NAC) relative to standard of care on 
subjective pain measurement (FIQR). All outcomes will be evaluated using a linear mixed model 
(LMM) approach.  Models will include fixed effects for treatment regimen, time, and a treatment 
by time interaction as well as a random subject effect to account for repeated measures on 
subjects.  Differences within and between treatment regimens will be estimated using a series of 
linear contrasts from the LMM.  We will enroll 16 subjects to allow for up to 25% loss to follow-
up.  A sample size of 12 subjects provides a two-sided 95% confidence interval with a distance 
from the mean paired difference of ± 0.64 standard deviations.  The analysis will be conducted 
as an intent to treat analysis.