Targeting Inflammation With Salsalate in Type 1 Diabetes Neuropathy-TINSAL -T1DN
Phase: PHASE2
Conditions: Type 1 Diabetes, Peripheral Neuropathy
Sponsor: University of Michigan

Study Title:  Targeting Inflammation With Salsalate in Type 1 Diabetes Neuropathy 
(TINSAL-T1DN)
PI: Rodica Pop-Busui, MD, PhD
NCT:  NCT02936843
Protocol Version IRB Approval Date: 10/10/2022

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CONFIDENTIAL 
CLINICAL PROTOCOL 
TARGETING INFLAMMATION WITH SALSALATE IN TYPE 1 DIABETES NEUROPATHY-
(TINSAL -T1DN STUDY) 
A phase II-III, multi-site randomized, double-blind, placebo-controlled clinical trial
US IND Number: 113650 
Protocol Number:  02 
Version and Date:  Version 2.3, September 19, 2022 
Protocol Committee:  
University of Michigan: Rodica Pop-Busui, MD, PhD; Study co-Chair, Eva Feldman, MD, PhD 
Study Co-Chair; Lead Coordinator, Catherine Martin, MS, RN, BC-ADM, CDE 
Joslin Diabetes Center: Allison Goldfine, MD: consultant 
Data Coordinating Center University of Michigan: Dr. Morton Brown, Department of 
Biostatistics, University of Michigan, 1415 Washington Heights M4507, Ann Arbor, MI 48109-
2029 
NOTE: To effectively manage study medication supply chain disruptions, changes to protocol-
items are to be made to minimize disruptions and ensure participate safety. These changes are 
listed in Appendix A of the protocol (Contingency Operation Plan). 
This document contains confidential information.  Except as otherwise agreed to 
in writing, by accepting or reviewing this document, you agree to hold this 
information in confidence and not to disclose it to others (except where required 
by applicable law) or to use it for unauthorized purposes.   

 
 
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SYNOPSIS 
Indication:   Prevention of diabetic neuropathy (DN) progression or improvement in patients 
with Type I diabetes (T1DM) and mild DN  
Rationale:  DN is the most common chronic complication of diabetes, ultimately affecting 
half of patients with T1DM and leading to severe morbidity, high mortality, major physical 
disability and poor quality of life.  Attainable intensive glycemic control, although necessary, 
is insufficient to prevent adverse nervous system effects, justifying a therapeutic need to 
identify new drug targets to treat DN early in its course. Evidence for an important role of 
low-grade inflammation and of nuclear factor kappa B (NF-κB) activation in the 
pathogenesis of DN is emerging from both experimental and clinical studies. Salsalate, a 
nonacetylated prodrug of salicylate, is an FDA approved treatment for osteoarthritis and 
other rheumatologic conditions. It is a highly effective drug in blocking the inhibitor of κB 
kinase (IKKβ)/NF-κB pathway, with a large margin of safety, and low cost. 
Objectives:  
Primary:  Determine whether a 12-month course of oral salsalate (up to 3 grams daily) will 
prevent the progression of DN, as measured by intraepidermal nerve fiber density (IENFD) 
at the proximal thigh and distal leg in subjects with T1DM and mild DN. 
Secondary:  Determine whether a 12 month course of up to 3 grams of salsalate daily will 
have an effect on:  
1. The change from baseline to months in IENFD at the distal thigh;  
2. The proportion of subjects with confirmed clinical neuropathy at baseline, and 12 months 
defined as the combination of:  
          a) Definite clinical neuropathy requiring at least two positive responses among 
neuropathic symptoms (pain, numbness, paresthesias), sensory signs, and abnormal ankle 
reflexes (diminished or absent) consistent with a distal symmetrical polyneuropathy and 
without causal explanation aside from diabetes, as assessed clinically using a standardized 
neurological examination; and 
        b) NC abnormalities involving ≥ 2 nerves among the sural, peroneal, and ulnar nerves.  
3. The proportion of subjects with definite small fiber neuropathy, defined as neuropathic 
symptoms and signs (decreased pinprick or temperature sensation) consistent with small 
fiber predominant dysfunction, and an abnormality in IENFD (one standard deviation below 
normal for age/gender/weight) or abnormal cardiovascular autonomic testing.  
4. Measures of cardiovascular autonomic neuropathy derived from cardiovascular reflex 
testing (R-R interval ratio during deep breathing, postural change, and Valsalva) at baseline 
and 12 months; resting HR will be obtained at baseline and 3 month intervals through 12 
months of follow-up.  
5. Measures of quality of life as assessed by the NeuroQOL domain and overall scores at 

 
 
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baseline, 6, and 12 months.  
6. Autonomic symptoms will be assessed using the Survey of Autonomic Symptoms (SAS). 
7. Measures of pain as assessed by the Neuropathy Pain Scale  
8. Functional Assessments: BERG Balance Scale; Falls Efficacy Scale, and 8 Foot Get-Up- 
and-Go assessments.  
Exploratory: Assessment of expression of pain-related molecules in skin biopsies, including 
sodium channels, TrkA receptors, neuropeptides, such as substance P and CGRP and 
TNFα. Serum inflammatory markers TNFα, IL1β, IL6 and CCL2 will also be measured.  
 
Trial Design:  This is a phase II-III, multi-site randomized, double-blind, placebo-controlled 
clinical trial to evaluate the effect of salsalate on measures of small fiber neuropathy in 70 
subjects with T1DM and mild DN. Subjects will be randomized in a 3:2 fashion to (up to) 3 
grams/day salsalate or matched placebo and receive study medication for 12 months. 
Randomization will be stratified by baseline HbA1c ((≤8.0 or >8.0%)). Subjects will be 
evaluated every 3 months for a total of 12 months.  
 
Endpoints: 
Primary outcome:  Change from baseline to month 12 in the intraepidermal nerve fiber 
density (IENFD) at the distal thigh. 
Secondary outcomes:  
IENFD at the ankle at baseline and 12 months  
Electrophysiological measures 
o Nerve Conduction Studies of the sural, peroneal, and ulnar nerves will be obtained at 
screening and 12 months 
o Quantitative Sensory Testing: Cold and vibration thresholds are assessed at 
screening and 12 months. 
o Measures of CAN will comprise cardiovascular reflex testing using the DCCT/EDIC 
protocol (R-R interval ratio during deep breathing, postural change, and Valsalva). 
Clinical measures of DN will be assessed at baseline and 12 months and will include:  
         a) DN symptoms as evaluated by the Neuropathy Total Symptom Score-6 (NTSS-6)  
         b) Survey of Autonomic Symptoms (SAS) 
         c) The DCCT/EDIC Structured Neurological Examination 
 
Measures of DN- specific pain will include the pain domain part of the NeuroQOL and the 
Neuropathic Pain Scale that will be administered at baseline, 6, and 12 months.  

 
 
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Functional scales will be assessed at baseline and 12 months and include:  
    a) The Berg Balance Scale which measures balance in 14 separate activities of daily 
living such as going from sitting-to-stand and standing on one leg; the unipedal stance 
portion that had been shown to be particularly reflective of neuropathy-related mobility loss;  
    b) The 8 Foot Up and Go Test, a test of functional mobility that assesses the time needed 
for a subject to arise from sitting position, walk 8 ft and turn 180 degrees around a cone and 
return sitting;  
     c) The Modified Falls Efficacy Scale assessing patient’s self-reported ability to perform 
activities of daily living (e.g. get dressed/ undressed, walking, shopping). 
 
Quality of life measures will be administered at baseline, 6, and 12 months and include the 
NeuroQOL, a validated measure of quality of life specific to peripheral neuropathy.  
 
Exploratory outcomes: will be assessed at baseline and 12 months and include:  
     a) Assessment of expression of pain-related molecules in distal leg skin biopsies, 
including the ion channel TRPV1, TrkA receptors, and neuropeptides, e.g. substance P and 
CGRP, using our published protocol;  
   b) skin nerve fiber inflammatory markers including TNFα, IL1β, CD68c, using our 
published protocol; and  
    c) a complex panel of serum inflammatory markers using the Luminex 200 platform 
(Luminex Corporation, Austin, TX) and Millipore multiplex immunoassay panels (Millipore 
Corporation, Chicago, IL, which offers customization of 40 different cytokines including 
TNFα, sTNF receptor, IL1β, IL6, MCP-1, adiponectin, IL8, IL17, m-CSF, sVCAM-1, s-ICAM-
1, IL10; and  
    d) urine inflammatory markers using the Luminex 200 platform which offers customization 
for urinary cytokines as outlined above 
Trial Treatments: 
Oral administration of salsalate up to 3 grams/daily for 12 months, or 
Oral administration of matched placebo daily for 12 months.  
Statistical Methods: 
We reported that the average IEFND in the distal leg in T1DM subjects (mean disease 
duration of 27 years) was 2.2 fibers/mm. We assume that the targeted diabetic subjects 
have the average IENFD for ages 20-29 shown by published normatives at the start of their 
disease, and the decline due to diabetes is 10 fibers/mm or an average of 0.25 fibers/mm 
/year. If the decline is more rapid as neuropathy progresses, then the average decline in 
IEFND may be as high as 0.5 fibers/mm/year. Based on our assumption about the 
mechanism by which salsalate will influence IEFND (reduction of inflammation), and 
published effect of other interventions, we believe that an increase of 1.65 fibers/mm (0.25 + 
1.4 = 1.65) in fiber density is possible with treatment with salsalate. The standard deviation 

 
 
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of the change is approximately 2 fibers/mm. With 25 subjects per group there will be 81% 
power to reject the null hypothesis of no difference between groups using a two-tailed t-test 
at a 5% level of significance.  No formal interim analysis for stopping for efficacy benefit or 
futility is planned, given the projected duration of follow-up required for the assessment of 
the primary endpoint.  Rather, safety will be assessed periodically and reviewed by a Data 
and Safety Monitoring Board. 
A general linear model will be fitted to the change in IENFD score with factors for treatment 
group, baseline HbA1c (≤8.0 or >8.0%), and baseline IEFND as fixed covariates. If the 
distributional assumptions are not met, transformations of IENFD or the use of 
nonparametric methods will be employed. Sensitivity analyses will be performed to assess 
how subjects who withdrew may affect conclusions of the analysis. An intent-to-treat 
analysis set (including subjects who are randomized, treated and have a baseline IENFD) 
will be used to assess treatment effects with multiple imputation methods employed to 
impute outcomes for subjects with missing post-baseline data. 
 
 
SUMMARY OF CHANGES: 
Protocol Version 1.2 
July 13, 2016:   
 
Section 4.1 Exclusion Criteria:  Updated to allow continued low-dose (81 mg daily) 
aspirin therapy. 
 
Section 2, 6.1, 6.2, Table 1: Updated to move quantitative sensory testing to the 
screening visit, rather than baseline visit. 
 
Section 2:  Corrected to indicate that nerve conduction studies are done at the screening 
visit. 
 
Section 2, 4.1, 6.1, 7.1.2:  Updated to indicate that the ulnar nerve will be tested, rather 
than median nerve. 
 
 
Protocol Version 1.3 
October 19th, 2016: 
 
Table 1:  Proposed visit schedule corrected for spelling and timing of events. 
 
Section 1 & 10.1:  Corrected to indicate that this is a single center (not multi-center) 
study. 
 
Section 4.1:  Added history of keloid scarring to list of exclusions. 
 
Section 6.2:  Removed language related to subject medication log 
 

 
 
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Section 10.6:  Added reference that MICHR will provide study monitoring. 
 
Throughout protocol provided clarification that research pharmacy used is Belmar 
Pharmacy (not UMHS Research Pharmacy). 
 
Minor typographical, spelling and grammatical errors fixed throughout. 
 
 
Protocol Version 1.4 
December 6th 2016 
 
 
 
Section 3:  Added that randomization will be by gender as well as by HbA1c. 
 
 
 
Section 4.1: 
Clarified inclusion and exclusion criteria as follows; 
 
-Severe hypoglycemia is defined by hypoglycemia resulting in coma or 
seizure.  
 
-Active Hepatitis C is an exclusion. 
 
-Active foot ulcers are an exclusion. 
 
-Exclusion related to DKA clarified to a history of recurrent DKA or any 
DKA in the prior 3 months. 
 
-Lithium use added to list of exclusions 
 
-Upper age for eligibility increased from 65 to 70 years. 
 
Protocol Version 1.5 
February 28, 2017 
 
Section 4.1:  Inclusion criteria amended follows;  
 
-Inclusion criteria # 4 amended to include provision to use peroneal motor 
nerve conduction velocity of ≥ 35 m/second as inclusion criteria if sural 
nerve amplitude is 0 μV (unrecordable). 
 
Section 4.1:  Clarified exclusion criteria as follows;  
 
-Exclusion # 9 clarified with addition of chronic immunosuppressive 
therapy to long-term glucocorticoid therapy, exception specified for those 
using steroid inhalers for asthma management. 
 
-Exclusion # 15 corrected to reflect treatment period of 12 months rather 
than 24 months. 
 
-Exclusion # 16 added (exclusions related to hypersensitivity to salsalate, 
aspirin and other NSAIDS). Language related to hypersensitivity is 

 
 
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included in section 9.3 of protocol, but has now been added to the 
exclusion list for clarity. 
 
-added language indicating that people with known hypersensitivity to 
lidocaine or epinephrine may not be able to participate as these agents 
are used for local anesthesia during skin biopsies. 
 
Protocol Version 1.6 
May 01, 2017 
 
Section 4.1:  Clarified exclusion criteria as follows;  
 
- 
Exclusion # 6 clarified to include “lung” transplant to match ICD 
 
Section 5.2.3 and 6.2:  Revised study drug titration schedule. 
 
Section 9.3:  Added language re: increased risk for fluid retention and edema to be 
consistent with risks for NSAIDs that are listed in the Salsalate package insert.   
 
Protocol Version 1.7 
August 10, 2017 
 
Section 3: Trial Design.  Amended to reflect increased enrollment to 70 subjects and to 
modify randomization distribution from 1:1 (Salsalate:Placebo) to 3:2 
(Salsalate:Placebo).   
 
Section 5.1:  Removed reference to 1:1 randomization ratio. 
 
Throughout:  
-Reference to enrollment of 60 subjects changed to 70 subjects 
 
-Reference to salsalate dose clarified from “3 grams per day” to “up to 3 grams 
per day” to better reflect intent to use a lower dose for participants who 
experience dose-dependent, treatment related side effects.  
 
-Clarified the randomization stratification by A1c from <, 8% to >8% to ≤ 8.0% to 
>8.0%. 
 
Protocol Version 1.8 
December 4th, 2017 
 
 
 
Table 1: Amended to increase the allowable visit window between screening (visit 1) 
 
and baseline (visit 2) 
 
 
Section 6.1: Amended last paragraph to reflect increase in allowable duration 
 
between screening (visit 1) and baseline (visit 2). 
 
 
Throughout: Minor formatting changes and spelling/grammar corrections. 
 
Protocol Version 1.9 

 
 
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July 11th, 2019 
 
Section 2.0: Sudoscan removed from study procedures due to the bankruptcy of the 
manufacturing company and the subsequent inability to obtain supplies 
 
 
Section 4.1: Clarified exclusion criteria as follows;  
 
 - Amputation Exclusion - edited exclusion criteria to allow patients who have had 
a lower limb amputation from a cause other than neuropathy  
 - Cancer Exclusion - edited exclusion criteria to only exclude patients with active 
cancer (other than basal cell or squamous cell skin cancer) rather than also 
excluding those who have had it in the last five years.  
 
- Macroabluminuria Exclusion - clarify that a history of persistent 
macroalbuminuria is checked by random urine microalbumin creatinine ratio 
rather than a 24-hour test. 
  
Section 5.3: Pill Disposal - edited the method of pill disposal in the protocol such that 
the pills will be disposed of in a U of M pharmaceutical black bin upon patient return 
and pill counting, rather than returning them to Belmar.  
Section 3.0 and 10.1: Changed single center to multi-site to reflect the addition of 
Henry Ford Health System as a performance site. In section 10.1, it is also noted that 
the previous version of this protocol contained an error that stated that the purpose of 
this single center study was the justify the conduction of single center study. This 
should have stated that the purpose of this single center study was the justify the 
conduction of multi-center study. Thus, this has now been edited to acknowledge that 
this is now a multi-site study that would justify a larger multi-site study. 
Protocol Version 2.0 
December 16th, 2019 
 
 Table 1:  Proposed Visit Schedule for TINSAL-T1DN Study: Removed Sudomotor 
testing row that was missed in the changes made for protocol version 1.9. 
 
Section 6.2: Removed Sudomotor testing from visit 2 tests as that was missed in the 
changes made for protocol version 1.9. 
 
Section 6.3: Removed Sudomotor testing from visit 7 tests as that was missed in the 
changes made for protocol version 1.9. 
 
 
Protocol Version 2.1 
December 18th 2020 
 
 
 
Hold over reference to 1:1 randomization changed to 3:2 on page 3.  
 

 
 
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 Section 8.8.1 updated to reflect SAE reporting in accordance with current IRB 
reporting guidelines. 
 
 Table one updated to reflect that visits 3-7 windows are based on medication start 
following visit 2 due to the shipping of the medication from Belmar pharmacy directly 
to the participant.  
 
Protocol Version 2.2 
January 11, 2022 
 
Section 6 was updated to include option for conducting visit number 4 (3 months) and 
6 (9 months) remotely. 
 
Table one updated to include a footnote indicating that visit number 4 and number 6 
may be conducted remotely.    
 
Protocol Version 2.3 
September 19, 2022 
 
 
Appendix A was added.  Appendix A provides a contingency plan in case of disruption 
to supplies of medication (salsalate or placebo) 
 
 
 

 
 
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Table 1:  Proposed Visit Schedule for TINSAL-T1DN Study 
 
Visit Number 
Visit 1 
Visit 2 
Visit 3 
Visit 4** 
Visit 5 
Visit 6** 
Visit 7 
Visit Description 
Screening 
Baseline 
Monitoring 
Monitoring 
Monitoring/ 
Outcomes 
Monitoring 
Monitoring/ 
Outcomes 
Time point 
Day -180 to 
0 
Day 0 
Month 1 
Month 3 
Month 6 
Month 9 
Month 12 
Visit window 
 
NA 
± 7 days 
± 14 days 
± 14 days 
± 14 days 
± 14 days 
Informed Consent 
X 
 
 
 
 
 
 
Medical History /Physical 
Examination 
X 
X 
 
X 
X 
X 
X 
Laboratory Measures* 
X 
 
 
X 
X 
X 
X 
Skin Biopsies (proximal/distal leg) 
 
X 
 
 
 
 
X 
Neurological History /Physical 
Examination 
X 
 
 
 
 
 
X 
Nerve Conduction Studies 
X 
 
 
 
 
 
X 
Neuropathy Total Symptom Score  
X 
X 
 
 
X 
 
X 
Cardiovascular Autonomic Studies 
 
X 
 
 
 
 
X 
Resting Heart Rate 
 
X 
X 
X 
X 
X 
X 
Quantitative Sensory Testing 
X 
 
 
 
 
 
X 
NeuroQOL 
 
X 
 
 
X 
 
X 
Survey of Autonomic Symptoms 
 
X 
 
 
X 
 
X 
Neuropathy Pain Scale 
 
X 
 
 
X 
 
X 
Functional Assessments 
   Berg Balance Scale 
   8-foot Get Up and Go Test 
   Modified Falls Efficacy Scale 
 
X 
 
 
 
 
X 
Medication Dispensing§ 
 
X 
 
X 
X 
X 
 
Medication Compliance 
 
 
X 
X 
X 
X 
X 
Medication Return 
 
 
 
X 
X 
X 
X 
Adverse Events Monitoring 
 
X 
X 
X 
X 
X 
X 
 Baseline history and examination will include diabetes duration, diabetes complications, diabetes treatment, concurrent medical problems, relevant 
past surgical history, height, weight, blood pressure and pulse, and foot examination.  Follow up visits will include changes in medical or surgical 
history, weight, blood pressure, pulse and foot examination * HbA1c, Metabolic Comprehensive Panel, Complete Blood Count with Platelets, TSH, 
Lipid Profile, Cystatin–C, Uric Acid and Urine Albumin performed at screening visit (unless results for these tests are documented in the medical record 
as being done in the prior 3 months for all except HbA1c (1 month) and Urine Albumin (6 months).  Urine pregnancy test at screening for all women of 
childbearing potential regardless of contraceptive practiced.  HbA1c, Metabolic Comprehensive Panel, CBCP, Cystatin-C, Uric Acid and Urine Albumin 
will be repeated at 3,6,9 and 12 months of follow-up. Lipids will be repeated at 3, 6 and 12 months of follow-up. Luminex inflammatory marker assays 
on blood and urine performed at screening or baseline, and again at study end. 
 
*Visit windows for visit 3-7 are based off when the patient starts the study medication following visit 2 due to the shipping of the medication from 
Belmar Pharmacy directly to the participant. 
** Visits 4 and 6 may be conducted remotely with laboratory samples collected at a Michigan Medicine affiliated lab. 
§Medication dispensing,and return may be modified if study drug supplies (salsalate, placebo) are disrupted (Appendix A). 

 
 
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TABLE OF CONTENTS 
1. INTRODUCTION ............................................................................................................................. 13 
1.1. Background ............................................................................................................................................ 13 
1.2. Rationale .................................................................................................................................................. 13 
2. TRIAL OUTCOMES........................................................................................................................ 14 
3. TRIAL DESIGN ................................................................................................................................ 15 
4. STUDY POPULATION .................................................................................................................. 16 
4.1. Inclusion Criteria .................................................................................................................................. 16 
5. Trial TREATMENTS....................................................................................................................... 18 
5.1. Allocation to Treatment ..................................................................................................................... 18 
5.2. Study Drug .............................................................................................................................................. 18 
5.2.1. Description of drug product ...................................................................................................................... 18 
5.2.2. Preparation and Dispensing..................................................................................................................... 19 
5.2.3. Administration.................................................................................................................................................. 19 
5.2.4. Compliance ....................................................................................................................................................... 19 
5.3. Drug Storage and Drug Accountability ...................................................................................... 20 
5.4. Concomitant Medication(s) .............................................................................................................. 20 
6. Study VISITS .................................................................................................................................... 20 
6.1. Recruitment/Screening ...................................................................................................................... 20 
6.2. Baseline .................................................................................................................................................... 21 
6.3. Follow-up Visits .................................................................................................................................... 22 
6.4. Scheduled Interim Contact .............................................................................................................. 23 
6.5. Subject Withdrawal.............................................................................................................................. 23 
7. ASSESSMENTS .............................................................................................................................. 23 
7.1. Efficacy ..................................................................................................................................................... 23 
7.1.1. Interaepidermal Nerve Fiber Density (IENFD)................................................................................ 23 
7.1.2. Additional Measures of Neuropathy ..................................................................................................... 24 
7.2. Safety ......................................................................................................................................................... 25 
7.3. Exploratory.............................................................................................................................................. 25 
Assessment of expression of pain-related molecules in skin biopsies, including sodium 
channels, TrkA receptors, neuropeptides, such as substance P and CGRP and TNFα. Serum 
and urine inflammatory markers TNFα, IL1β, IL6 and CCL2 will also be measured ................. 25 
8. Adverse Event Reporting ........................................................................................................... 25 
8.1. Adverse Events ..................................................................................................................................... 25 
8.2. Definition of an Adverse Event ...................................................................................................... 25 
8.3. Abnormal Laboratory Findings ...................................................................................................... 26 
8.4. Serious Adverse Events .................................................................................................................... 26 
8.5. Hospitalization ....................................................................................................................................... 27 
8.6. Severity Assessment .......................................................................................................................... 27 
8.7. Discontinuations (See also Subject Withdrawal, Section 6.5) .......................................... 28 
8.8. Eliciting Adverse Event Information ............................................................................................ 28 
8.8.1. Reporting Requirements (Serious and Nonserious Adverse Events) ................................. 28 
8.8.2. Reporting Serious Adverse Events ...................................................................................................... 28 
8.8.3. Reporting Non-Serious Adverse Events ............................................................................................ 29 

 
 
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9. Expected Risks Associated with the Protocol .................................................................. 29 
9.1. Risks Associated with Study Participation ............................................................................... 29 
9.2. Risks Associated with Blood Collection .................................................................................... 29 
9.3. Risks Associated with Study Medication .................................................................................. 30 
9.4. Risks Associated with Nerve Conduction Studies ................................................................ 31 
9.5. Risks Associated with Skin Biopsies .......................................................................................... 31 
9.6. Risk Associated with Cardiovascular Autonomic Testing ................................................. 31 
10. DATA ANALYSIS/STATISTICAL METHODS .................................................................... 31 
10.1. Sample Size Determination ........................................................................................................... 31 
10.2. Efficacy Analysis ............................................................................................................................... 32 
The two treatment groups will be compared with respect to demographic and baseline 
variables (e.g., age, BMI, smoking, etc). If a significant difference is found in any variable, the 
models described below will be fitted both with and without the variables that differ at 
baseline. ............................................................................................................................................................. 32 
10.3. Analysis of Exploratory Endpoints ............................................................................................ 33 
10.4. Safety Analysis ................................................................................................................................... 33 
10.5. Interim Analysis ................................................................................................................................. 34 
10.6. Data and Safety Monitoring Board ............................................................................................. 34 
11. DATA HANDLING AND RECORD KEEPING .................................................................... 34 
11.1. Case Report Forms / Electronic Data Record ....................................................................... 34 
12. ETHICS ............................................................................................................................................ 35 
12.1. Institutional Review Board (IRB) ................................................................................................. 35 
12.2. Ethical Conduct of the Trial .......................................................................................................... 35 
12.3. Subject Information and Consent............................................................................................... 35 
12.4. Subject Confidentiality .................................................................................................................... 35 
12.5. Sample and Data Storage ............................................................................................................... 35 
References ............................................................................................................................................ 36 
Appendix A………………………………………………………………………………..………...….38 
 
 

 
 
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1.  INTRODUCTION 
For patients with type 1 diabetes (T1DM) and diabetic neuropathy (DN), no disease modifying 
treatment other than glycemic control is available. The IKKβ/NF-κB pathway is unique in that it 
is implicated in both the pathogenesis of nerve fiber loss, pain and other symptoms in DN 1-4. 
This will be the first study to confirm beneficial effects of the anti-inflammatory agent, salsalate, 
on preventing DN progression and on DN pain. The proposed study design employs a 
quantifiable early measure of DN, intraepidermal nerve fiber density (IENFD) of the thigh, 
allowing for accurate assessment of actual nerve fiber density over time5, while also 
incorporating traditional large fiber DN outcomes, the Diabetes Control and Complications 
Trial/Epidemiology of Diabetes Interventions and Complications (DCCT/EDIC) primary DN 
endpoint 6,7, measures of pain and quality of life.   
1.1. Background 
Diabetic neuropathy is the most common chronic complication of diabetes, ultimately affecting 
half of patients with T1DM and leading to severe morbidity, high mortality, major physical 
disability, poor quality of life 8; and estimated total annual costs of $22 billion 
(www.diabetes.org). Due to the complex structure and anatomy of the peripheral nervous 
system, DN presents with a very broad spectrum of clinical symptoms and deficits, including 
severe pain, sensory deficits, foot ulcers and amputations.  Despite the high morbidity 
associated with DN, most randomized clinical trials evaluating therapies for established DN 
have been disappointing. To date there is no pathogenetic treatment for this condition 8-10. The 
Diabetes Control and Complications Trial (DCCT) demonstrated that intensive control designed 
to achieve near-normal glycemia is essential in reducing the risk of DN development in type 1 
diabetes 6,7.  However, attainable intensive glycemic control, although necessary, is insufficient 
to prevent adverse nervous system effects 11, justifying a therapeutic need to identify new drug 
targets to treat DN early in its course. 
1.2. Rationale 
Evidence for an important role of low-grade inflammation and of nuclear factor kappa B (NF-kB) 
activation in the pathogenesis of DN and in the pain syndrome associated with DN is emerging 
from both experimental 1,2,12 and clinical studies 13.  This suggests that agents with known anti-
inflammatory properties, such as salicylates, may prevent the development of DN and the pain 
associated with DN. Salsalate, a pro-drug form of salicylate, is indicated for relief of the signs 
and symptoms of rheumatoid arthritis, osteoarthritis and related rheumatic disorder. It is a highly 
effective drug in blocking the IKKβ/NF-κB pathway, and can be obtained at a low cost.  
Salsalate has recently been shown to have glucose lowering effects. 
The recently published NIDDK-funded “Targeting Inflammation Using Salsalate in Type 2 
Diabetes (TINSAL-T2D)” trial confirmed salutary effects of 3.5 gram/day salsalate on markers of 
inflammation, glucose control and overall safety after 48 weeks in patients with type 2 diabetes 
(T2DM).  
Our initial NIDDK funded R03 (DK 094499) grant confirmed the safety and feasibility of targeting 
inflammation with salsalate treatment in T1DM subjects with DN (26). Our current study builds 
upon and expands our initial promising results and will either confirm or refute the therapeutic 
efficacy of salsalate in a larger T1DM cohort.  

 
 
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Traditionally the diagnosis of DN is made by the presence of abnormal nerve conductions (NC), 
and a combination of symptoms and signs of distal sensorimotor polyneuropathy. However, NC 
studies assess only large myelinated nerve fiber function and in early DN, these conventional 
measures can be normal and if abnormal, change slowly. Recent data have also described 
minimal worsening 14 or improvement in NC 15 in placebo and epidemiological cohorts with little 
relation to other measures of small fiber nerve function in diabetic patients. This was further 
confirmed by the relative modest changes in all NC studies in the DCCT/EDIC cohort during 13-
14 years of follow-up while adhering to current standard of care 16.  
 
In T1DM, DN is a diffuse, symmetrical injury to the entire peripheral nervous system (5, 30-33). 
The smallest Aδ thinly-myelinated fibers and the unmyelinated C-fibers are likely the earliest to 
undergo damage in the natural history of DN. These fibers mediate pain, temperature 
discrimination, touch and autonomic responses, and constitute 80% to 90% of all peripheral 
nerve fibers (34). Current evidence on DN natural history point to a progressive course leading 
to later stage phenotypes that include abnormality in the morphology and function of larger 
nerve fibers (as documented by changes in NC studies).  
Therefore, when studying a population with earlier stages of disease which would be most 
amenable to a therapeutic intervention, it is important to utilize outcome measures that best 
identify these patients (in this case a measure that sensitively capture small fiber deficits).  
 
IENFD is endorsed by several societies and authorities in the field including the Toronto 
Consensus Panel on Diabetic Neuropathy as a reliable, efficient and valid outcome measure to 
assess early neuropathy in clinical trials 5,8,17,18. IENFD, is a continuous measure of small fiber 
neuropathy, with high positive and negative predictive values along with a high diagnostic 
efficiency in differentiating between subjects with and without neuropathy 5,19,20.The 
morphometric quantification of IENFD, most commonly expressed as the number of IENFs per 
length of section (IENF/mm e.g. density) is possible with skin biopsy, a minimally invasive 
procedure, with less than 1% of mild adverse events such as bleeding, swelling or erythema. 
Inter-observer variability for the assessment of IENFD demonstrates good agreement, 
especially with assessment at the thigh, and a large normative data base was recently created 
by an international consortium 5,19,20. Patient refusal rate for this procedure is minimal (less than 
2 % in several past and ongoing trials of the two Principal Investigators). 
 
2. TRIAL OUTCOMES  
Primary outcome:  Change from baseline to month 12 in the intraepidermal nerve fiber density 
(IENFD) at the distal thigh. 
Secondary outcomes:  
IENFD at the ankle at baseline and 12 months  
Electrophysiological measures 
o Nerve Conduction Studies of the sural, peroneal, and ulnar nerves will be obtained at 
screening and 12 months 
o Quantitative Sensory Testing: Cold and Vibration thresholds are assessed at screening 
and 12 months. 

 
 
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o Measures of CAN will comprise cardiovascular reflex testing using the DCCT/EDIC 
protocol (R-R interval ratio during deep breathing, postural change, and Valsalva). 
Clinical measures of DN will be assessed at baseline and 12 months and will include:  
         a) DN symptoms as evaluated by the Neuropathy Total Symptom Score-6 (NTSS-6)  
         b) Survey of Autonomic Symptoms (SAS) 
         c) The DCCT/EDIC Structured Neurological Examination 
 
Measures of DN- specific pain will include the pain domain part of the NeuroQOL and the 
Neuropathic Pain Scale that will be administered at baseline, 6, and 12 months.  
 
Functional scales will be assessed at baseline and 12 months and include:  
    a) The Berg Balance Scale which measures balance in 14 separate activities of daily living 
such as going from sitting-to-stand and standing on one leg; the unipedal stance portion that 
had been shown to be particularly reflective of neuropathy-related mobility loss;  
    b) The 8 Foot Up and Go Test, a test of functional mobility that assesses the time needed for 
a subject to arise from sitting position, walk 8 ft. and turn 180 degrees around a cone and return 
sitting;  
     c) The Modified Falls Efficacy Scale assessing patient’s self-reported ability to perform 
activities of daily living (e.g. get dressed/ undressed, walking, shopping). 
 
Quality of life measures will be administered at baseline, 6, and 12 months and include the 
NeuroQOL, a validated measure of quality of life specific to peripheral neuropathy.  
 
Exploratory outcomes: will be assessed at baseline and 12 months and include:  
     a) Assessment of expression of pain-related molecules in distal leg skin biopsies, including 
the ion channel TRPV1, TrkA receptors, and neuropeptides, e.g. substance P and CGRP, using 
our published protocol;  
   b) skin nerve fiber inflammatory markers including TNFα, IL1β, CD68c, using our published 
protocol; and  
    c) a complex panel of serum inflammatory markers using the Luminex 200 platform (Luminex 
Corporation, Austin, TX) and Millipore multiplex immunoassay panels (Millipore Corporation, 
Chicago, IL, which offers customization of 40 different cytokines including TNFα, sTNF receptor, 
IL1β, IL6, MCP-1, adiponectin, IL8, IL17, m-CSF, sVCAM-1, s-ICAM-1, IL10; and  
    d) urine inflammatory markers using the Luminex 200 platform which offers customization for 
urinary cytokines as outlined above 
3. TRIAL DESIGN 
This is a phase II-III, multi-site randomized, double-blind, placebo-controlled clinical trial to 
evaluate the effect of salsalate on measures of small fiber neuropathy in 70 subjects with T1DM 
and mild DN. In the original study design we proposed to randomize up to 60 subjects in a 1:1 
fashion to up to 3 grams of salsalate or matched placebo.  In August 2017 the protocol was 
modified by recommendation of the DSMB to address differential study medication 
discontinuation rates and to increase the number of subjects exposed to study drug.  
Specifically, the protocol will enroll up to 70 subjects and randomize them in a 3:2 fashion to 

 
 
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salsalate or placebo.  All subjects enrolled subsequent to the approval of the revised protocol 
will be randomized in 3:2 fashion, up to 3 grams per day of salsalate or placebo respectively.  
Treatment duration will be 12 months.  Randomization will be stratified by baseline HbA1c (≤8.0 
or >8.0%) and by gender within the HbA1c groups. Subjects will be evaluated every 3 months 
for a total of 12 months.  
4. STUDY POPULATION 
70 subjects with type 1 diabetes mellitus (T1DM) will be enrolled in this study.  
This clinical trial can fulfill its objectives only if appropriate subjects are enrolled.  The following 
eligibility criteria are designed to select subjects for whom protocol treatment is considered 
appropriate.  All relevant medical and non-medical conditions should be taken into consideration 
when deciding whether this protocol is suitable for a particular subject. 
4.1. Inclusion Criteria 
1. T1DM;  
 
2. age 18-70;  
3. mild DN as defined by symptoms and/or signs, confirmed by at least one abnormality in 
electrophysiology studies (abnormality of at least one attribute among conduction 
velocity, latency, amplitude or F-Wave in at least one nerve among sural sensory, ulnar 
sensory, or peroneal motor);  
4. sural nerve amplitude > 0 μV. If sural nerve amplitude is 0 μV (unrecordable) peroneal 
motor nerve conduction velocity must be ≥ 35 m/second*; 
5. on a stable insulin regimen for the 3 months prior to enrollment;    
6. be willing and capable of signing the IRB approved consent form and willing and able to 
cooperate with the medical procedures for the study duration;    
7. be willing to accept random treatment assignment to salsalate or placebo; and    
8. women of childbearing age agree to use an appropriate contraceptive method 
(hormonal, IUD, or diaphragm) for the duration of the study and must have a negative 
urine pregnancy test at screening.  
Exclusion Criteria  
1. history of severe DN, active lower limb ulceration or lower limb amputation directly 
caused by diabetic neuropathy, or risk factors for any other causes of neuropathy (e.g. 
active hepatitis C, end stage renal disease, systemic lupus erythematosus or a known 
hereditary neuropathy) as determined through medical history, family history, history of 
medications, occupational history, history of exposure to toxins, physical and 
neurological examinations); 

 
 
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2. history of recent severe hypoglycemia (within prior 6 months) as defined by 
hypoglycemia resulting in coma or seizure or a history of recurrent diabetic ketoacidosis 
(DKA) or any diabetic ketoacidosis within the last three months.  
3. history of persistent macroalbuminuria [random urine microalbumin creatinine ratio 
(ACR) >300 mg/gm]. ACR up to 300 mg/gm is acceptable if serum creatinine is <1.4 for 
women, <1.5 for men AND estimated GFR (eGFR) is > 60; 
4. serum creatinine >1.4 for women and >1.5 for men or eGFR <60 [possible chronic 
kidney disease stage 3 or greater calculated using the Modification of Diet in Renal 
Disease (MDRD) equation]; 
5. pregnancy or lactation, or intention to become pregnant in next 12 months; 
6. history of previous lung, kidney, pancreas, liver, cardiac or bone marrow transplant;    
7. history of drug or alcohol abuse within the previous 5 years, or current weekly alcohol 
consumption >10 units/week; 
8. use of warfarin (Coumadin), clopidogrel (Plavix), dipyridamole (Persantine), heparin or 
other anticoagulants, probenecid (Benemid, Probalan), sulfinpyrazone (Anturane) or 
other uricosuric agents; Subjects must agree to not use high-dose aspirin during the 
course of the study.  Daily low-dose aspirin treatment (not more than 81 mg per day) 
may be continued if currently prescribed. 
9. requiring long-term glucocorticoid therapy or chronic immunosuppressive therapy; 
Inhaled steroid use for management of asthma is not an absolute exclusion. 
10. use of lithium   
11. participation in an experimental medication trial within 3 months of starting the study;    
12. current therapy for malignant disease other than basal- cell or squamous-cell skin 
cancer;    
13. history of gastrointestinal bleeding or active gastric ulcer; screening laboratory 
abnormalities including AST (SGOT) and or ALT (SGPT) > 2.5 x the upper limit of 
normal (ULN), total bilirubin > 1.5 x ULN, platelets < 100,000; 
14. developed keloid scarring in the past.  Keloids are large, thick masses of scar tissue. 
These are more common among dark-skinned people.      
15. presence of any condition that, in the opinion of the investigator would make it unlikely 
for the subject to complete 12 months of study participation, e.g., history of non- 
adherence to therapeutic regimens, presence of conditions likely to limit life expectancy, 
living situation that would interfere with study visit schedules such as a job requiring 
frequent or extended travel  
16. known hypersensitivity to salsalate.  Patients who have experienced asthma, hives, or 
other allergic-type reactions to aspirin or other NSAIDs are excluded from participation.  

 
 
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Patients with known or suspected aspirin or NSAID-sensitive asthma are excluded. 
In addition, subjects with concurrent chicken pox, influenza, flu-like symptoms or other 
symptomatic viral illnesses should not be enrolled in the study until the illness or condition has 
resolved.  
Subjects with known or suspected hypersensitivity to lidocaine or epinephrine may not be able 
to participate as these agents are used for local anesthesia during skin biopsies.  The study 
investigators should consider the nature and severity of past reported reactions to these agents, 
and may consider alternative anesthesia options for local anesthesia on a case by case basis. 
*Because obtaining the sural nerve response can be technically challenging, the absence of 
response does not necessarily indicate advanced neuropathy.  Criteria for peroneal motor nerve 
conduction velocity is added as a secondary measure of neuropathy severity so that otherwise 
eligible candidates are not excluded based solely on inability to obtain the sural response. 
5. TRIAL TREATMENTS 
5.1. Allocation to Treatment 
Randomization to Salsalate or Placebo: After signing informed consent, confirming eligibility and 
completion of baseline assessments (visit 2), subjects will be randomized to salsalate, up to 3 
grams/daily or matched placebo for 12 months. Randomization will be stratified by baseline 
HbA1c (≤8.0% or >8.0%).  
Procedure for Subject Treatment Assignment: Subjects will be randomized to treatment with 
either Salsalate or Placebo.  The procedures for randomization will be determined by the study 
biostatistician. 
Breaking the Blind  
In the event of an emergency and a need to know study medications, the investigator will 
contact Belmar Research Pharmacist (the research pharmacy contracted for this study) and 
request identification of subject’s test article assignment. The Belmar Pharmacist will indicate 
that the blind was broken on the Pharmacy Randomization Log and will document the date of 
the request, the name of the research personnel making the request, and the pharmacist’s 
signature. The research coordinator will report the emergent event on the Adverse Event CRF 
and the unmasking event on the Unmasking Report CRF and then signed by the investigator.   
5.2. Study Drug 
This study will be conducted under IND 113650 (Dr. Pop-Busui, Sponsor-Investigator). 
Neither subjects nor their insurance will incur any additional cost from their participation in this 
study.  Salsalate and matching placebo will be provided by Belmar Research Pharmacy. 
5.2.1. Description of drug product 
Salsalate tablets 500mg 

 
 
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Salsalate is a nonsteroidal anti-inflammatory agent for oral administration.  Chemically, salsalate 
(salicylsalicylic acid or 2 hydroxy-benzoic acid, 2-carboxyphenyl ester) is a dimer of salicylic 
acid; its structural formula is shown below: 
 
5.2.2. Preparation and Dispensing 
The Belmar Research Pharmacy will act as the central pharmacy for this study, and will provide 
salsalate and matching placebo to each participant.   
5.2.3. Administration 
Each subject will be given sufficient study medication to last until the next scheduled visit.   
Subjects will be instructed to take one tablet with breakfast for the first four days beginning with 
the first breakfast after receipt of the study medication.  The study medication dose will be 
increased slowly to the maximum tolerated dose (2 tablets, 3 times daily) using the following 
schedule:   Beginning on day 5, take 1 tablet with breakfast and 1 tablet with lunch.  Beginning 
on day 9, take 1 tablet at breakfast, 1 tablet at lunch, 1 tablet at dinner.  Beginning on day 23, 
take 2 tablets at breakfast, 1 tablet at lunch, 1 tablet at dinner.  Beginning on day 27 take 2 
tablets at breakfast, 2 tablets at lunch, 1 tablet at dinner.  The maximum dose of study 
medication, 2 tablets at breakfast, 2 tablets at lunch, and 2 tablets at dinner, may be started no 
sooner than the 31st day from receiving the first supply of study medication.   Should side effects 
(e.g., stomach upset, tinnitus) develop, the subject will be instructed to contact the study staff for 
guidance re: reducing the dose of study medication. 
5.2.4. Compliance 
The number of tablets dispensed at each visit will be recorded in the case report form.  Subjects 
are reminded to return all unused study medication at each visit in the original container.  The 
study staff will collect all unused medications and record the number of tablets remaining.  
Discrepancies between expected versus actual study medication returned will be evaluated by 
the local study staff to address issues related to non-adherence to the study protocol.   Subjects 
will be encouraged to continue on the maximally tolerated dose of study medication for the 
duration of the treatment period.  Consistent with the intention to treat model, all subjects will be 
followed for the duration of the study treatment period according to the treatment schedule, 
unless consent is withdrawn, or, in the opinion of the investigator, it is unsafe for the subject to 
continue participation. 

 
 
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5.3. Drug Storage and Drug Accountability 
The Belmar Research Pharmacy shall be responsible for storage and dispensing of study drug 
for the cohort of subjects followed at the University of Michigan and Henry Ford Health System.  
The clinical coordinator will be responsible for drug accountability. Upon the completion of drug 
accountability, the coordinator will discard the remaining drug in a University of Michigan 
pharmaceutical bin.  
5.4. Concomitant Medication(s) 
The research staff will record in the source documents all concomitant drugs used by the 
subject.  This is to include prescription, non-prescription, homeopathic medications and 
dietary/vitamin supplements.  At the screening visit, medication use is to be carefully reviewed 
to ensure that no contraindicated medications as described in the protocol are being used (e.g., 
blood thinning agents, immune-modulating medications, NSAIDS).  At all subsequent visits, 
subjects will be asked to report any changes to their reported medication use and the source 
document updated accordingly.  Queries regarding medication exclusions and contraindications 
are directed to the study co-chairs. 
6. STUDY VISITS 
The schedule of study assessments is shown in Table 1.  After obtaining informed consent, 
verifying eligibility at screening and completing all required baseline assessments, subjects are 
randomized, receive study medication and are followed every 3 months for one year. The staff 
should query the subject regarding any changes to their health, changes to their medication 
regimen, including changes in their insulin treatment, and frequency and severity of 
hypoglycemia.  
Visits 4 and 6 may be conducted remotely if the study investigator agrees that a remote visit is 
appropriate for the participant, and the participant has access to a University of 
Michigan/Michigan Medicine affiliated laboratory.  The remote visit option for Visits 4 and 6 will 
reduce the burden of study participation for those participants who live at a distance from the 
study site, or have limited ability to take time off from work or have other limiting obligations, or 
who are concerned about travel during the COVID-19 pandemic. Study staff will conduct the 
visit via telephone or site-approved and HIPPA compliant electronic conferencing platform.  
Study staff will place an order for required laboratory assessments and instruct the participant to 
have the laboratory measures obtained at the nearest Michigan Medicine affiliated laboratory. 
6.1. Recruitment/Screening 
Visit 1: Screening/Eligibility.  
An eligibility visit will be scheduled for subjects who, based on preliminary screening (telephone, 
email, medical chart review) meet the general entry criteria (age, T1DM, diabetes duration, 
symptomatic peripheral neuropathy, no contraindications for salsalate, etc.). At visit 1 the 
investigator or her designee will obtain written informed consent, after which all required 
eligibility examinations will be performed. The eligibility examination will include the following 
laboratory assessments: HbA1c, metabolic comprehensive panel (METCOMP), complete blood 
count with platelets (CBCP), thyroid stimulating hormone (TSH), lipid profile, cystatin-C, uric 
acid, urine albumin assessment, and in all women of child bearing age and potential, a urine 

 
 
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pregnancy test, even if an accepted method of contraception is being used. If any of the 
required laboratory measures are documented in the subject’s medical record as being 
completed in the prior 3 months (1 month for HbA1c, and 6 months for urine albumin), they will 
not need to be repeated at the screening visit. Additional blood and urine will be obtained for 
Luminex assays of inflammatory markers. The Luminex assays though may be postponed until 
the baseline examination. 
Weight, height and seated blood pressure and pulse will be obtained.  
A structured neurologic history and physical examination and nerve conduction studies will be 
performed at the screening visit to verify and document the presence of signs or symptoms of 
peripheral neuropathy, the presence of a sural nerve response, and abnormality of at least one 
abnormal nerve attribute (amplitude, latency, conduction velocity or f-wave) in the sural, 
peroneal or ulnar nerve. In the absence of a sural nerve response (e.g., sural amplitude is 0 µV, 
response is unobtainable, then the presence of a peroneal motor nerve conduction velocity of ≥ 
35 meters per second will be required for inclusion.  Quantitative sensory testing will also be 
conducted at the screening visit. 
All visit 1 measures are ideally to be completed within a 4-week time frame but may take up to 6 
months (180 days) if repeated measures are needed, and may be done at one, or over several 
visits if that is more convenient for the subject.  
6.2. Baseline 
Visit 2: Baseline (Day 0).  
The key activities of the baseline visit are to obtain the initial skin biopsies and to initiate 
treatment with either Salsalate or placebo. Cardiovasular autonomic neuropathy (CAN) 
assessment, as well as quality of life and symptom questionnaires, and a short battery of 
functional assessments, will also be performed at the baseline visit. Two skin biopsies are 
obtained; one at the proximal thigh, and one at the distal leg. These will be done on the 
subjects’ dominant side unless contraindicated (e.g., presence of a cast).  
Upon satisfactory completion of all eligibility and baseline assessments, including the skin 
biopsy, a 3 month + 14-day supply of Salsalate or placebo will be provided, and written 
instructions for use of the medication reviewed with and provided to the subject. Subjects will be 
instructed to take one tablet with breakfast for the first four days beginning with the first 
breakfast after receipt of the study medication.  The study medication dose will be increased 
slowly to the maximum tolerated dose (2 tablets, 3 times daily) using the following schedule:   
Beginning on day 5, take 1 tablet with breakfast and 1 tablet with lunch.  Beginning on day 9, 
take 1 tablet at breakfast, 1 tablet at lunch, 1 tablet at dinner.  Beginning on day 23, take 2 
tablets at breakfast, 1 tablet at lunch, 1 tablet at dinner.  Beginning on day 27 take 2 tablets at 
breakfast, 2 tablets at lunch, 1 tablet at dinner.  The maximum dose of study medication, 2 
tablets at breakfast, 2 tablets at lunch, and 2 tablets at dinner, may be started no sooner than 
the 31st day from receiving the first supply of study medication.    
  Subjects are to be reminded to return any unused study medication at their next visit.  
Study staff will contact the subject within one week of the baseline visit to assess medication 
compliance and tolerance and to inquire as to the status of the biopsy sites.  

 
 
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6.3. Follow-up Visits 
Visit 3: Safety Monitoring (Month 1).  
At this visit, the study staff will measure the weight, blood pressure, and resting heart rate, will 
observe and note the condition of the baseline biopsy sites, and will ask the subject about any 
changes in health status or physical symptoms and record the findings in the source  
documents. The staff will assess medication compliance and tolerance. Adverse events will be 
similarly recorded and reported as required.  
 
Visit 4: Safety Monitoring and Drug Dispensing (Month 3).  
At this visit, the study staff will measure the weight, blood pressure, and resting heart rate, will 
observe and note the condition of the baseline biopsy sites, and will ask the subject about any 
changes in health status or physical symptoms and record the findings in the source 
documents. Adverse events will be similarly recorded and reported as required. Laboratory 
assessments (HbA1c, METCOMP, CBCP, cystatin-C, lipids, uric acid and urine albumin) will be 
obtained.  Urine pregnancy testing may be performed at the discretion of the investigator for all 
women of childbearing potential.  The study staff will collect any unused study medication, and 
will dispense 3 months + 14 day of medication. The number of unused salsalate tablets will be 
recorded in the source document and unused medication secured until returned for final drug 
disposition.  
Visit 5: Safety Monitoring and Drug Dispensing (Month 6).  
At this visit, the study staff will measure the weight, blood pressure and resting heart rate, and 
will ask the subject about any changes in health status or physical symptoms. Laboratory 
assessments (HbA1c, METCOMP, CBCP, cystatin-C, lipids, uric acid and urine albumin) will be 
obtained.  Urine pregnancy testing may be performed at the discretion of the investigator for all 
women of childbearing potential.  The study staff will collect any unused study medication, and 
will dispense a 3 month + 14-day supply of study medication. The number of unused salsalate 
tablets will be recorded in the source document and unused medication secured until returned 
for final drug disposition. The subjects will be asked to complete quality of life and neuropathy 
symptom surveys.  
Visit 6: Safety Monitoring and Drug Dispensing (Month 9).  
Weight, blood pressure and resting heart rate are recorded and inquiries made re: changes in 
health status or physical symptoms. Laboratory assessments (HbA1c, METCOMP, CBCP, 
cystatin-C, uric acid and urine albumin) will be obtained.  Urine pregnancy testing may be 
performed at the discretion of the investigator for all women of childbearing potential.  The study 
staff will collect any unused study medication, and will dispense a 3 month + 14-day supply of 
medication. The number of unused salsalate tablets will be recorded in the source document 
and unused medication secured until returned for final drug disposition.  
Visit 7: Final Visit (Month 12).  
Weight, blood pressure and resting heart rate are recorded and inquiries made re: changes in 
health status or physical symptoms. Laboratory assessments (HbA1c, METCOMP, CBCP, 
cystatin-C, lipids, uric acid and urine albumin) will be obtained as will blood and urine samples 

 
 
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for Luminex assays of inflammatory markers.  Urine pregnancy testing may be performed at the 
discretion of the investigator for all women of childbearing potential.  The study staff will collect 
any unused study medication. The number of unused salsalate tablets will be recorded in the 
source document and unused medication secured until returned for final drug disposition.  
The structured neurological history and examination and nerve conduction studies performed at 
screening will be repeated.  
Cardiovascular autonomic neuropathy (CAN) assessment, quantitative sensory tests, as well as 
quality of life and symptom questionnaires, and battery of functional assessments, will be 
performed at the final visit. Two skin biopsies are obtained; one at the proximal thigh, and one at 
the distal leg. These will be done on the subjects’ dominant side unless contraindicated (e.g., 
presence of a cast). The biopsies will be obtained from skin which is adjacent (within 1 cm) to 
the original biopsy sites.  
Within 1 week of the final visit, the study staff will contact the subject by phone or email to 
inquire about any changes in health status, and to as inquire as to the status of the skin biopsy 
sites.  
6.4. Scheduled Interim Contact 
Within 1 week of Visit 2 (Baseline, initial biopsy) the study staff will contact (telephone, email, in-
person) the subject to inquire about any problems at the biopsy sites and to inquire about 
medication compliance and any side effects encountered.  This contact will be recorded in the 
source document.   
6.5. Subject Withdrawal 
A subject may withdraw from the trial at any time at their own request, or they may be withdrawn 
at any time at the discretion of the investigator or sponsor for safety, behavioral, or 
administrative reasons.  If a subject does not return for a scheduled visit, every effort should be 
made to contact the subject.  In any circumstance, every effort should be made to document 
subject outcome, if possible.   
If the subject withdraws consent, no further evaluations should be performed and no attempts 
should be made to collect additional data; however, they may be questioned regarding their 
reason for withdrawal and asked to return all unused investigational products. 
7. ASSESSMENTS 
7.1. Efficacy     
7.1.1. Interaepidermal Nerve Fiber Density (IENFD) 
Traditionally the diagnosis of DN is made by the presence of abnormal nerve conductions (NC) 
studies, and a combination of symptoms and signs of distal sensorimotor polyneuropathy. 
However, NC studies assess only large myelinated nerve fiber function and in early DN, these 
conventional measures can be normal and if abnormal, change slowly. Recent data have also 
described minimal worsening 14 or improvement in NC 15 in placebo and epidemiological cohorts 
with little relation to other measures of small fiber nerve function in diabetic patients. This was 

 
 
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further confirmed by the relative modest changes in all NC studies in the DCCT/EDIC cohort 
during 13-14 years of follow-up while adhering to current standard of care 16.  
Therefore, when studying a population with earlier stages of disease which would be most 
amenable to a therapeutic intervention, it is important to utilize outcome measures that best 
identify these patients (in this case, a measure that sensitively captures small fiber deficits).  
 
In patients with confirmed or more advanced DN, the prevalence of abnormal NC, quantitative 
sensory testing and IENFD was comparable 5,18. Data from several trials show also that IENFD 
was significantly reduced in patients with normal NC studies and that there is an inverse 
correlation between IENFD and the severity of DN, defined by the Neurological Disability Score  
and the Neuropathy Impairment Score 18, suggesting early damage to small nerve fibers. These 
data suggest that IENF loss is an early feature of DN, progresses with increasing neuropathic 
severity and may improve with appropriate intervention. Additionally, IENFD was endorsed by 
several societies and authorities in the field including the Toronto Consensus Panel on Diabetic 
Neuropathy as a reliable, efficient and valid outcome measure to assess small fiber neuropathy 
in clinical trials 5,8,17,18.  
 
IENFD is a continuous measure of small fiber neuropathy, with high positive and negative 
predictive values along with high diagnostic efficiency in differentiating between subjects with 
and without neuropathy 5,19,20.The morphometric quantification of IENFD, most commonly 
expressed as the number of IENF per length of section (IENF/mm e.g. density) is possible with 
skin biopsy, a minimally invasive procedure, with less than 1 % of mild adverse events such as 
bleeding, swelling or erythema. Intra- and inter-observer variability for the assessment of IENFD 
demonstrates good agreement and a large normative data base was recently created by an 
international consortium 5,19,20. Patient refusal rate for this procedure is minimal. 
 
The change in IENFD at the distal thigh and ankle will be assessed by skin biopsies obtained at 
baseline and 12 months (as described in detail in the Manual of Operations. 
7.1.2. Additional Measures of Neuropathy 
Considering the complexity of nerve fiber involvement in patients with diabetes, we have 
included in our trial design several additional outcomes including the change in the composite 
measure of large fiber neuropathy (the endpoint used in the DCCT/EDIC study) as a secondary 
outcome and definite small fiber neuropathy. 
1. The change from baseline to months in IENFD at the distal thigh;  
2. The proportion of subjects with confirmed clinical neuropathy at baseline, and 12 months 
defined as the combination of:  
a) Definite clinical neuropathy requiring at least two positive responses among neuropathic 
symptoms (pain, numbness, paresthesias), sensory signs, and abnormal ankle reflexes 
(diminished or absent) consistent with a distal symmetrical polyneuropathy and without causal 
explanation aside from diabetes, as assessed clinically using a standardized neurological 
examination and, 
b) NC abnormalities involving ≥ 2 nerves among the sural, peroneal, and ulnar nerves.  
3. The proportion of subjects with definite small fiber neuropathy, defined as neuropathic 
symptoms and signs (decreased pinprick or temperature sensation) consistent with small fiber 

 
 
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predominant dysfunction, and an abnormality in IENFD (one standard deviation below normal 
for age/gender/weight) or abnormal cardiovascular autonomic testing.  
4. Measures of cardiovascular autonomic neuropathy derived from cardiovascular reflex testing 
(R-R interval ratio during deep breathing, postural change, and Valsalva) at baseline and 12 
months; resting HR will be obtained at baseline and 3 month intervals through 12 months of 
follow-up.  
5. Measures of quality of life as assessed by the NeuroQOL domain and overall scores at 
baseline, 6, and 12 months.  
6. Autonomic symptoms will be assessed using the Survey of Autonomic Symptoms (SAS). 7. 
Measures of pain as assessed by the Neuropathy Pain Scale  
8. Functional Assessments: BERG Balance Scale; Falls Efficacy Scale, and 8 Foot Get-Up- 
and-Go assessments.  
7.2. Safety 
Safety will be assessed through the collection of adverse events and laboratory parameters. 
7.3. Exploratory 
Assessment of expression of pain-related molecules in skin biopsies, including sodium 
channels, TrkA receptors, neuropeptides, such as substance P and CGRP and TNFα. Serum 
and urine inflammatory markers TNFα, IL1β, IL6 and CCL2 will also be measured  
8. ADVERSE EVENT REPORTING 
8.1. Adverse Events 
All observed or volunteered adverse events regardless of treatment group or suspected causal 
relationship to the investigational product(s) will be recorded on the adverse event page(s) of 
the case report form (CRF).  
For all adverse events, the investigator must pursue and obtain information adequate both to 
determine the outcome of the adverse event and to assess whether it meets the criteria for 
classification as a serious adverse event requiring immediate notification to the IRBMED. For all 
adverse events, sufficient information should be obtained by the investigator to determine the 
causality of the adverse event. The investigator is required to assess causality and indicate that 
assessment on the CRF. Follow-up of the adverse event, after the date of therapy 
discontinuation, is required if the adverse event or its sequelae persist. Follow-up is required 
until the event or its sequelae resolve or stabilize at a level acceptable to the investigator and 
the IRBMED.  
8.2. Definition of an Adverse Event 
An adverse event is any untoward medical occurrence in a clinical investigation subject 
administered a product or medical device; the event need not necessarily have a causal 

 
 
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relationship with the treatment or usage.  Examples of adverse events include but are not 
limited to:   
• 
Abnormal test findings; 
• 
Clinically significant symptoms and signs; 
• 
Changes in physical examination findings; 
• 
Hypersensitivity; 
• 
Progression/worsening of underlying disease; 
8.3. Abnormal Laboratory Findings 
The criteria for determining whether an abnormal objective test finding should be reported as an 
adverse event are as follows:  
• 
Test result is associated with accompanying symptoms, and/or 
• 
Test result requires additional diagnostic testing or medical/surgical intervention, and/or 
• 
Test result leads to a change in trial dosing outside of protocol-stipulated dose adjustments, 
discontinuation from the study, significant additional concomitant drug treatment, or other 
therapy, and/or 
• 
Test result is considered to be an adverse event by the investigator or sponsor. 
Merely repeating an abnormal test, in the absence of any of the above conditions, does not 
constitute an adverse event.  Any abnormal test result that is determined to be an error does not 
require reporting as an adverse event. 
8.4. Serious Adverse Events 
A serious adverse event or serious adverse drug reaction is any untoward medical occurrence 
at any dose that: 
• 
Results in death; 
• 
Is life-threatening; 
• 
Requires inpatient hospitalization or prolongation of existing hospitalization; 
• 
Results in persistent or significant disability/incapacity; 
• 
Results in congenital anomaly/birth defect. 
Medical and scientific judgment should be exercised in determining whether an event is an 
important medical event.  An important medical event may not be immediately life-threatening 
and/or result in death or hospitalization.  However, if it is determined that the event may 
jeopardize the subject and may require intervention to prevent one of the other outcomes listed 
in the definition above, the important medical event should be reported as serious. 

 
 
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8.5. Hospitalization 
Adverse events reported from clinical trials associated with hospitalization or prolongation of 
hospitalization are considered serious.  Any initial admission (even if less than 24 hours) to a 
healthcare facility meets these criteria.   
Hospitalization does not include the following: 
• 
Rehabilitation facilities; 
• 
Hospice facilities; 
• 
Respite care (e.g., caregiver relief); 
• 
Skilled nursing facilities; 
• 
Nursing homes; 
• 
Routine emergency room admissions; 
• 
Same day surgeries (as outpatient/same day/ambulatory procedures). 
Hospitalization or prolongation of hospitalization in the absence of a precipitating, clinical 
adverse event is not in itself a Serious Adverse Event.  Examples include:  
• 
Admission for treatment of a preexisting condition not associated with the development of a 
new adverse event or with a worsening of the preexisting condition (e.g., for work-up of 
persistent pre-treatment lab abnormality); 
• 
Social admission (e.g., subject has no place to sleep); 
• 
Administrative admission (e.g., for yearly physical exam); 
• 
Protocol-specified admission during a clinical trial (e.g., for a procedure required by the trial 
protocol); 
• 
Optional admission not associated with a precipitating clinical adverse event (e.g., for 
elective cosmetic surgery); 
• 
Pre-planned treatments or surgical procedures should be noted in the baseline 
documentation for the entire protocol and/or for the individual subject. 
Diagnostic and therapeutic non-invasive and invasive procedures, such as surgery, should not 
be reported as adverse events.  However, the medical condition for which the procedure was 
performed should be reported if it meets the definition of an adverse event.  For example, an 
acute appendicitis that begins during the adverse event reporting period should be reported as 
the adverse event, and the resulting appendectomy should be recorded as treatment of the 
adverse event. 
8.6. Severity Assessment 
If required on the adverse event case report forms, the investigator will use the 
adjectives MILD, MODERATE, or SEVERE to describe the maximum intensity of the 
adverse event.  For purposes of consistency, these intensity grades are defined as 
follows: 

 
 
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MILD 
Does not interfere with subject's usual function. 
MODERATE 
Interferes to some extent with subject's usual function. 
SEVERE 
Interferes significantly with subject's usual function. 
 
Note the distinction between the severity and the seriousness of an adverse event.  A severe 
event is not necessarily a serious event.  For example, a headache may be severe (interferes 
significantly with subject's usual function) but would not be classified as serious unless it met 
one of the criteria for serious adverse events, listed above. 
8.7. Discontinuations (See also Subject Withdrawal, Section 6.5) 
The reason for a subject discontinuing from the trial will be recorded in the CRF.  A 
discontinuation occurs when an enrolled subject ceases participation in the study, regardless of 
the circumstances, prior to completion of the protocol.  The investigator must determine the 
primary reason for discontinuation.  Withdrawal due to adverse event should be distinguished 
from withdrawal due to insufficient response, according to the definition of adverse event noted 
earlier, and recorded on the appropriate adverse event CRF page.   
When a discontinuation is due to a serious adverse event, the serious adverse event must be 
reported in accordance with the reporting requirements defined below. 
8.8. Eliciting Adverse Event Information 
The investigator is to report all directly observed adverse events and all adverse events 
spontaneously reported by the trial subject.   
8.8.1. Reporting Requirements (Serious and Nonserious Adverse Events) 
Each adverse event is to be classified by the investigator as serious or nonserious.  This 
classification determines the reporting procedures to be followed.  If a serious adverse event 
occurs, expedited reporting will follow local and international regulations, as appropriate. 
SAEs are reportable from the time that the subject provides informed consent, which is obtained 
prior to the subject’s participation in the clinical trial, i.e., prior to undergoing any trial-related 
procedure and/or receiving investigational product.   
8.8.2. Reporting Serious Adverse Events 
Serious adverse events will be reported to the UMIRBMED in accordance with IRB Reporting 
guidelines.  
In the event that the investigator does not become aware of the occurrence of a serious adverse 
event immediately (e.g., if an outpatient trial subject initially seeks treatment elsewhere), the 
investigator is to report the event within 24 hours after learning of it and document the time of 
his/her first awareness of the adverse event. 

 
 
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The investigator’s assessment of causality must be provided.  An investigator’s causality 
assessment is the determination of whether there exists a reasonable possibility that the 
investigational product caused or contributed to an adverse event.  If the investigator’s final 
determination of causality is unknown and the investigator does not know whether or not study 
drug caused the event, then the event will be handled as “related to study drug” for reporting 
purposes.  If the investigator's causality assessment is "unknown but not related to study drug", 
this should be clearly documented on study records.  In addition, if the investigator determines 
the adverse event is associated with trial procedures, the investigator must record this causal 
relationship in the source documents and CRF, as appropriate, and report such an assessment 
in accordance with the serious adverse event reporting requirements, if applicable.  
8.8.3. Reporting Non-Serious Adverse Events 
Adverse events reported by subjects that are unrelated or unlikely related to study participation, 
regardless of expectedness, will not routinely be reported, unless the severity and duration of 
the event reaches the threshold for defining a serious adverse event. Local reporting guidelines 
should be followed. 
 
Adverse events reported by subjects that are related (definitely related, probably related or 
possibly related) to study participation and are expected based on known risks of participation, 
will not be routinely reported, unless the severity and duration of the event reaches the 
threshold for defining a serious adverse event.  For example, an episode of mild, self-treated 
hypoglycemia would be expected because of the underlying diagnosis of type 1 diabetes and 
expected as a known side-effect of salsalate.  This would be classed as a mild expected 
adverse event possibly related to study treatment and would not be reported 
 
9. EXPECTED RISKS ASSOCIATED WITH THE PROTOCOL 
9.1. Risks Associated with Study Participation 
Risks associated with study participation in general are related to the time commitment for the 
study, and risk for loss of privacy.  There will be 7 “in-person” visits required for the study, as 
well as 2-3 “remote” visits conducted by phone or email.    The study staff will try to minimize the 
burden of the visits to the extent practical by scheduling on days and at times that are most 
convenient for the patient.  Privacy risks will be reduced by maintaining a separate research 
record that uses a coded identifier, rather than personal identities.  Links to the coded identifier 
and personal identity shall be maintained in a secure electronic file by the study coordinator and 
the links will ultimately be destroyed.  Destruction of the links will occur no sooner than 2 years 
after the study has been acknowledged as terminated by the University of Michigan IRB (UM 
IRBMED). 
9.2. Risks Associated with Blood Collection 
Risks associated with blood collection are pain, discomfort, bleeding, bruising or infection.  
There is also the risk of dizziness, light-headedness, nausea or even fainting.   Blood sampling 
will be performed by staff with skill and experience in venipuncture. 

 
 
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9.3. Risks Associated with Study Medication 
The study is a randomized, double blind study and so neither the subject nor the study staff will 
know whether the medication is salsalate or a placebo.  Not knowing what the actual study 
medication is may cause anxiety.  Subjects who are not willing to accept random treatment 
assignment will not be accepted into the study.   
The Belmar Research Pharmacy will maintain a master list of treatment assignments and in the 
event of a medical emergency, may be contacted to “unmask” the subject treatment. 
Once the study is completed in all subjects, subjects may request their treatment assignment.   
The common side effects of salsalate are nausea, loss of appetite, heartburn and feeling of 
ringing in the ears and some degree of deafness.  These effects usually subside with ongoing 
treatment but if they persist, the dose of salsalate may be reduced.  Subjects will be instructed 
to take the medication at mealtimes to reduce the GI side effects of the medication.  While the 
GI and otologic side effects of salsalate tend to subside, if they persist or are troubling, the dose 
of salsalate may be reduced or salsalate may be stopped.   
Salsalate may cause allergic reactions.  People with known allergy to salsalate, aspirin, or to 
other NSAIDS are excluded from participating in the study.  Subjects will be reported to stop 
salsalate treatment should they develop a rash, itching, hives, or difficulty breathing and to 
contact the study team or their usual health care provider right away.  Emergency care may be 
required if the subject has breathing difficulties. 
Salsalate and other salicylate medications in high doses can cause gastrointestinal bleeding, 
especially in patients with previous history of gastric ulcers.  Patients with a history of GI 
bleeding or gastric ulcers are excluded from participation.  Subjects will be advised of the risk 
for GI bleeding and provided information regarding signs and symptoms of GI bleeding and are 
asked to report these symptoms to the study team or to their regular health care provider. 
Salsalate and other drugs in the class of salicylates can, especially in high doses, cause the 
blood sugar to drop, which may increase the risk of hypoglycemia in this population of people 
with type 1 diabetes.  Subjects will be made aware of the risk for low blood sugar and advised to 
check blood glucose levels frequently, and to carry treatment for low blood sugar.  Frequent 
monitoring and carrying treatment for low blood sugar is consistent with routine advice given to 
people with type 1 diabetes and people with type 2 diabetes on insulin or on other hypoglycemic 
agents.  The subjects’ insulin dose may be adjusted to prevent hypoglycemia. 
Salsalates, like other NSAIDs can cause cardiovascular side effects including heart attack or 
stroke.  Subjects will be informed of the symptoms of stroke and heart attack and instructed to 
seek emergent evaluation and treatment should symptoms occur. 
Fluid retention and edema has been associated with NSAIDS, and so these should be used with 
caution in people with fluid retention or heart failure.   
Salsalate, like other NSAIDs can cause serious skin reactions such as exfolliative dermatitis and 
Stevens-Johnson Syndrome.  Subjects will be instructed to report any rash, blistering, hives, 
fever or itching that occurs while on salsalate, and to stop using salsalate and contact the study 
doctor or their own health care provider should a rash or other skin changes appear. 
Salsalate may cause hepatotoxicity; subjects will be asked to report nausea, fatigue, itchiness, 
jaundice, abdominal pain and discomfort and or flu-like symptoms to the study staff or to their 
usual doctor and, to discontinue the medication until an evaluation of the symptoms as been 
done. 

 
 
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Salsalate is not approved for use in pregnant or nursing women. Pregnant and nursing women 
(and women who are planning pregnancy) are excluded from participation. Subjects are asked 
about current contraceptive practices during screening and those with child-bearing potential 
must agree to use an acceptable form of contraception during the study [oral contraceptive pills, 
injectable contraception (e.g. depo-provera) IUDs, barrier contraceptives (diaphragm/condoms), 
or surgical contraception (tubal ligation)].  A urine pregnancy test will be performed on all 
women of childbearing potential at baseline, regardless of contraceptive practices and will be 
repeated during the study if indicated.  Women who suspect pregnancy are asked to contact the 
study staff and may be asked to have a pregnancy test.  Women who suspect pregnancy or 
who are known to be pregnant are instructed to stop study medication immediately and to 
contact the study staff. 
9.4. Risks Associated with Nerve Conduction Studies   
Risks associated with nerve conduction studies are the discomfort associated with these 
studies.  The subjects will be alerted that they will feel a sensation similar to a “carpet shock” or 
snap from a rubber band during the test and will be reassured that the sensation will be 
temporary.  A skilled electromyographer will perform the studies and care will be taken to 
minimize the discomfort associated with the studies.  
9.5. Risks Associated with Skin Biopsies   
The risks associated with skin biopsies are pain, bleeding, bruising, or infection.  Subjects may 
be left with a small scar or discolored area at each biopsy site.  Four sites are biopsied in the 
course of the study, each about 3 mm in diameter.  The skin biopsies will be done by members 
of the staff who have training and experience in doing them. The areas biopsied will be cleaned 
with antiseptics prior to the biopsy and a local anesthetic used to reduce pain. Subjects will be 
given written information about wound care and how and when to report any signs or symptoms 
of infection.  The study staff will call the subject within one week each biopsy to check on how it 
is healing. 
9.6. Risk Associated with Cardiovascular Autonomic Testing 
Participants will be asked to fast in preparation for testing and are therefore at heightened risk 
for hypoglycemia.  The study team will advise the subject regarding adjustments to insulin 
dosing to reduce the risk for hypoglycemia.  Testing will be scheduled in the morning to shorten 
the duration of fasting.  The test itself has few risks associated with it, although some patients 
may become dizzy or lightheaded when moving from supine to standing, or after performing 
valsalva maneuvers.  Subjects will be reminded to report any dizziness or lightheadedness 
immediately to the examiner and will be instructed to return to a seated or lying position as 
needed to relieve symptoms.   
10. DATA ANALYSIS/STATISTICAL METHODS 
Detailed methodology for summary and statistical analyses of the data collected in this trial will 
be documented in a Statistical Analysis Plan, which will be prepared during the R34 and 
finalized prior to breaking of the blind. 
10.1. Sample Size Determination 
The purpose of this clinical trial is to provide the preliminary data necessary to design a larger 

 
 
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multi-site clinical trial. There are two assumptions that are necessary to justify the larger trial. 
The first is that salsalate will cause an increase in nerve fiber density (re-innervation) and the 
second is that the natural progression of DN, even with treatment for diabetes, leads to loss of 
nerve fibers. The primary outcome is an assessment of IENFD at the distal thigh after 12 
months of placebo or salsalate treatment.   
In the largest study of IENFD, Lauria et al57 provided a table of median IEFND values in the 
distal leg as a function of age and gender. In the 20-29 age range the average IEFND was 12.2 
fibers/mm with an average decline of 0.1 fibers/mm per year. We reported that the average 
IEFND in the distal leg in T1DM subjects (mean disease duration of 27 years) was 2.2 
fibers/mm. If we assume that our diabetic subjects have the average IENFD for ages 20-29 
shown by Lauria et al at the start of their disease, then the decline due to diabetes was 10 
fibers/mm or an average of 0.25 fibers/mm /year. If the decline is more rapid as neuropathy 
progresses, then the average decline in IEFND may be as high as 0.5 fibers/mm/year.  
In a study of the effect of physical exercise Smith et al found an increase in the IENFD at the 
thigh of 1.4 fibers/mm after one year of intervention. Based on our assumption about the 
mechanism by which salsalate will influence IEFND (reduction of inflammation), we believe that 
a similar increase in fiber density is possible with treatment with salsalate. In multiple studies 
that reported a change in IEFND over time (including one of our own studies), the standard 
deviation of the change was approximately 2 fibers/mm.  With the above assumptions, there will 
be an expected difference of 1.65 fibers/mm (0.25 + 1.4 = 1.65) in the change between the two 
groups at the thigh with a standard deviation of 2.0. With 25 subjects per group there will be 
81% power to reject the null hypothesis of no difference between groups using a two-tailed t-test 
at a 5% level of significance. Prior data are insufficient to perform a power calculation for the 
distal leg.  
The trial will be considered a success if the primary endpoint demonstrates a significant 
difference between the two groups and one other secondary endpoint trends in a similar 
direction. This trial will also be considered a success if there is sufficient evidence to show that 
IEFND increases with salsalate but is likely to decrease with usual treatment. With 25 subjects 
per group, the standard error of the estimate will be approximately 0.4 [=2/square root (25)] and 
therefore the confidence interval for the mean change in each group will be the estimated mean 
change ± 0.8 fibers/mm; i.e. an increase of greater than 0.8 fibers/mm with salsalate and an 
observed decrease with placebo (since the expected decrease with placebo is not expected to 
be large). With these results, this trial would provide the preliminary data needed for a larger 
trial to validate the effect of salsalate. Assuming 15-20% attrition rates, 70 subjects will be 
recruited to attain the necessary 50. 
10.2. Efficacy Analysis 
The two treatment groups will be compared with respect to demographic and baseline variables 
(e.g., age, BMI, smoking, etc.). If a significant difference is found in any variable, the models 
described below will be fitted both with and without the variables that differ at baseline. 
 
Analysis of Primary Outcome:  
 
The primary outcome is change from baseline to 12 months in IENFD at the distal thigh. A 
general linear model will be fitted to the change in IENFD score with factors for treatment group, 
baseline HbA1c (≤8.0% or >8.0%), and baseline IEFND as fixed covariates. If the distributional 

 
 
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assumptions are not met, transformations of IENFD or the use of nonparametric methods will be 
employed. Sensitivity analyses will be performed to assess how subjects who withdrew may 
affect conclusions of the analysis. An intent-to-treat analysis set (including subjects who are 
randomized, treated and have a baseline IENFD) will be used to assess treatment effects with 
multiple imputation methods101 employed to impute outcomes for subjects with missing post-
baseline data. Depending on the extent and pattern of missingness, other simpler sensitivity 
analyses may be used: for example, change from baseline to month 12 for subjects completing 
the study (completers) may be analyzed using an analysis of variance model. The model will 
include the same covariates that are included in the primary analysis.   
 
Analyses of Secondary Outcomes:  
Due to multiple tests, formal statistical testing won’t be performed; p-values will be reported to 
allow the reader to assess the result. For analysis of repeated measures of continuous data 
(change from screening/baseline to 12 months in NC studies, quantitative sensory testing, CAN 
measures, Neuropathy Symptoms, Pain, and NeuroQOL domain scores), a longitudinal 
approach will be employed. Specifically, each outcome will be analyzed using a mixed effect 
model for repeated measures with the treatment group and baseline HbA1c (≤8.0 or >8.0%)) as 
fixed covariates. Under the model, time of assessment will be a continuous variable and the 
outcome will be modeled as a linear function of time. Appropriate non-linear parametric or non-
parametric tests may be applied if assumptions of the model are not satisfied. 
 
Logistic regression will be used to compare the proportion of subjects who have confirmed 
clinical neuropathy with NC and those who had normal NC and therefore required an 
assessment of IENFD for trial entry, between the salsalate and placebo groups, with covariates 
for treatment group, and baseline HbA1C (≤8.0 or >8.0%). The components of this composite 
endpoint will also be assessed. Rank-transformation analysis of covariance will be used on 
each of the NC values, and an overall treatment group differences in the multiple NC values will 
be assessed using the nonparametric Wei-Lachin rank test of stochastic order102. The impact of 
additional covariates on the relationship between IENFD and confirmed clinical neuropathy and 
treatment will be explored by adding age, sex, height, weight, BMI, cigarette smoking, 
cardiovascular risk factors and concomitant medications into the models described above. A 
forward stepwise selection procedure will be used, with p<0.10 used to retain covariates. 
Interaction terms between treatment groups and select covariates will be investigated. In 
addition, we will adjust for the effect of changes in glucose on outcomes during the course of 
treatment by including HbA1c as a time-varying covariate in models. 
10.3. Analysis of Exploratory Endpoints  
Several exploratory analyses will be employed to investigate phenotypic characteristics 
associated with response to intervention, and the association of biomarkers with outcomes, to 
maximize the scientific knowledge gained from this study.  For example, exploiting the 
longitudinal measures of the phenotypic characteristics, we will jointly model longitudinal and 
event time (e.g., response to treatment) as a function of treatment group and other potential 
prognostic factors 25,26.  We will also use more sophisticated statistical models, such as linear 
and/or non-linear mixed effects models or nonparametric regression, to explore the relationship 
of phenotype and response to treatment, and biomarkers with clinical outcomes 25,27,28. 
10.4. Safety Analysis 
Adverse events and laboratory parameters will be summarized by treatment group.  Chi-square 
or Fisher’s exact tests will compare salsalate to placebo for discrete outcomes, and two-sample 

 
 
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t tests (on appropriately transformed continuous outcomes, if necessary) or nonparametric 
wilcoxon tests will assess treatment differences for continuous outcomes.  Serious adverse 
events (SAEs), unexpected adverse events and all AEs will also be summarized descriptively by 
body system and treatment group, regardless of relatedness to study treatment. 
10.5. Interim Analysis  
Given the expected enrollment period of 2 years and the 1-year follow-up period, an interim 
analysis to stop study enrollment for benefit or futility is not feasible.  Rather, periodic 
assessment of the safety of study subjects will be undertaken by an independent Data and 
Safety Monitoring Board (see below).  Although salsalate is an approved medication (off-label 
for this indication), if there were safety concerns in this study, the DSMB could request a formal 
analysis of safety and efficacy data to delineate the risk-benefit profile of salsalate in this study, 
with the potential to stop the study.   
10.6. Data and Safety Monitoring Board  
Study monitoring will be done by the Michigan Institute for Clinical and Health Research 
(MICHR).  A Data and Safety Monitoring Board (DSMB) will oversee this study.  Members of the 
DSMB are independent of the study investigators and represent Endocrinology, Neurology, 
Biostatistics, as well as possibly a lay member.  The DSMB will meet every three months or 
more frequently if requested by the Chairs either in person or by teleconference.  This protocol 
will be approved by the DSMB prior to initiation of recruitment. 
11. DATA HANDLING AND RECORD KEEPING 
11.1. Case Report Forms / Electronic Data Record 
As used in this protocol, the term case report form (CRF) should be understood to refer to either 
a paper form or an electronic data record or both, depending on the data collection method used 
in this trial. 
A CRF is required and should be completed for each included subject.  It is the investigator's 
responsibility to ensure completion and to review and approve all CRFs.  CRFs must be signed 
by the investigator or by an authorized staff member.  These signatures serve to attest that the 
information contained on the CRFs is true.  At all times, the investigator has final personal 
responsibility for the accuracy and authenticity of all clinical and laboratory data entered on the 
CRFs.  Subject source documents are the physician's subject records maintained at the trial 
site.  In most cases, the source documents will be the hospital's or the physician's chart.  In 
cases where the source documents are the hospital or the physician's chart, the information 
collected on the CRFs must match those charts.  
In some cases, the CRF may also serve as the source document.  In these cases, the DCC and 
the investigator must prospectively document which items will be recorded in the source 
documents and for which items the CRF will stand as the source document. 

 
 
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12. ETHICS 
12.1. Institutional Review Board (IRB)  
It is the responsibility of the investigator to obtain prospective approval of the trial protocol, 
protocol amendments, informed consent forms, and other relevant documents, e.g., 
advertisements, if applicable, from the IRB.  All correspondence with the IRB should be retained 
in the Investigator File. 
12.2. Ethical Conduct of the Trial 
The trial will be performed in accordance with the protocol, International Conference on 
Harmonization Good Clinical Practice guidelines, and applicable local regulatory requirements 
and laws. 
12.3. Subject Information and Consent 
The informed consent form must be agreed to by the Protocol Committee and the IRB and must 
be in compliance with ICH GCP, local regulatory requirements, and legal requirements. 
The investigator must ensure that each trial subject, or her legally acceptable representative, is 
fully informed about the nature and objectives of the trial and possible risks associated with 
participation.  The investigator will obtain written informed consent from each subject or the 
subject's legally acceptable representative before any study-specific activity is performed.  The 
informed consent form used in this trial, and any changes made during the course of the trial, 
must be prospectively approved by the IRB before use.  The investigator will retain a copy of 
each subject's signed consent form. 
12.4. Subject Confidentiality 
Study records with the study participant’s information for internal use at the clinical sites will be 
secured at the study site during the study. At the end of the study, all records will continue to be 
kept in a secure location. There are no plans to destroy the records.  
Study participant data, which is for reporting purposes, will be stored at the Coordinating Center. 
Data will identify participants by the unique study identification number. The study data entry 
and study management systems used by clinical sites and by Coordinating Center research 
staff will be a secured, password protected computer. 
12.5.  Sample and Data Storage 
Data collected for this study will be analyzed and stored at the Coordinating Center. After the 
study is completed, de-identified data will be stored at the Coordinating Center for use by 
researchers for future analyses with the participant’s approval. The results of these future 
analyses will not be made known to the participant.  
 
 
 

 
 
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Wang Y, Schmeichel AM, Iida H, Schmelzer JD, Low PA. Enhanced inflammatory 
response via activation of NF-kappaB in acute experimental diabetic neuropathy subjected to 
ischemia-reperfusion injury. J Neurol Sci 2006;247:47-52. 
2. 
Cameron NE, Cotter MA. Pro-inflammatory mechanisms in diabetic neuropathy: focus 
on the nuclear factor kappa B pathway. Curr Drug Targets 2008;9:60-7. 
3. 
Cheng HT, Dauch JR, Oh SS, Hayes JM, Hong Y, Feldman EL. p38 mediates 
mechanical allodynia in a mouse model of type 2 diabetes. Mol Pain 2010;6:28. 
4. 
Hur J, Sullivan KA, Pande M, et al. The identification of gene expression profiles 
associated with progression of human diabetic neuropathy. Brain 2011;134:3222-35. 
5. 
Lauria G, Hsieh ST, Johansson O, et al. European Federation of Neurological 
Societies/Peripheral Nerve Society Guideline on the use of skin biopsy in the diagnosis of small 
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Appendix A: Contingency Operation Plan in Case of Reduced Availability of Study 
Medication 
 
The following changes may be implemented and/or adapted in response to disruptions in study 
medications (salsalate and/or placebo) availability.   Information explaining any departures from 
the current protocol due to unavailability, or limited availability of study medication will be 
documented in the subject’s research record.  Changes to study medication dose and visit 
schedules will be documented on the protocol deviation log with a notation that the change in 
dose or schedule is because of change in medication availability.  The notation will permit the 
study team to censor those protocol changes that were made because of medication shortages 
(versus, non-adherence for example) and these events will ultimately be reported separately from 
other protocol deviations.  
 
1. 
New subject enrollment 
Enrollment of new subjects (Screening Visit 1) may be paused until the investigator is notified 
that adequate supplies of study medications are available.  Eligible participants who have not yet 
completed Visit 2 (randomization, assignment, and initiation of study medication) will have Visit 
2 postponed.  The protocol currently allows for Visit 2 to take place up to 180 days after Visit 1.  
If Visit 2 must be delayed for more than 180 days due to unavailability of study medication, then 
baseline laboratory measures performed at Visit 1 will be repeated at Visit 2.  A delay of over 
365 days would require that all Visit 1 screening measures be repeated prior to Visit 2.  
Reconsent of the participant would be required in this case. 
 
2. 
Treatment and visit modifications for enrolled participants (Visits 2 to 7)  
The overarching goal of treatment modifications in the setting of limited supplies of study 
medications will be to keep all participants on at least a minimal daily dose of study medication 
during their remaining time in the study. The study site staff will collaborate with the research 
pharmacy to achieve this goal on a case-by-case basis. 
 
The current protocol indicates that participants will be provided with sufficient study medication 
to last until the next scheduled visit.  If this cannot be achieved based on participants’ current 
maximally tolerated dose of the study medication, then the shipment schedule, or amount of 
medication shipped may be adjusted.  Examples include sending less than the full number of 
tablets (e.g., sending a 2 month versus 3 month supply), or sending fewer tablets to provide a 
lower dose over a period of 3 months.  Similarly, changes to treatment (dosing of study 
medications) and visit schedules will be individualized with consideration to a number of factors 
including: 
• 
the current dose of study medication being taken 
• 
the number of doses of study medication currently in the participant’s possession  
• 
the number of study visits completed/number of study visits remaining 
• 
the expected time to resolution of the study medication supply shortage 
Modifications in the event that supplies of study medication are limited or unavailable include, 
but are not be limited to the following:  
• 
Participants will NOT return unused study medication after each visit/receipt of new 
study medication.  Instead, they will be permitted to continue to use study medications 

 
 
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until the bottles are empty. This instruction will be documented in the participant’s 
research record. 
• 
To avoid gaps in exposure to study medication and to increase the likelihood of 
reaching 12 months of exposure to study medication the dose of study medication may be 
reduced, generally to ½ of the maximally tolerated dose, (up to 6 tablets per day), with 
further reductions as necessary to the minimum of 1 tablet per day).   
• 
When study medication becomes available, the dose will be titrated back to the prior 
maximally tolerated dose according to the current study medication titration schedule 
(beginning with the participants’ current dose and up-titrating to the maximally tolerated 
dose).  Participants will be reminded that study medication can cause changes in blood 
glucose levels and changes to the dose of study medication may require adjustments to 
their insulin doses.  Regular glucose monitoring will be encouraged.  These instructions 
are consistent with the current protocol and consent document.    
• 
If participants are without study medication at any dose for a period of 3 consecutive 
months, at any point between visit 2 and 6, the study team will consult with the DSMB to 
determine whether the participant should be scheduled for an early final visit (visit 7), or 
whether the participant should simply be withdrawn from further participation.  
• 
Participants who have completed visit 6, and who do not have an adequate supply of 
study medication to remain on their maximally tolerated dose until visit 7 will either   
o reduce their daily dose of study medication so that the number of tablets available 
extends to the opening of the Visit 7 window.  Visit 7 should be scheduled as 
early in the window as possible 
o complete visit 7 prior to the opening of the visit 7 window.  This could include 
completing all Visit 7 measures at Visit 6 if it is clear that NO study medication 
will be available to the participant after completing visit 6  
 
3. 
Alternate Source of Study Medication 
The research pharmacy (Belmar) will identify alternate suppliers of study medication.  Once a 
new manufacturer/supplier is identified, appropriate regulatory authorities (FDA/IRB) will be 
notified and the research pharmacy will obtain the study medication. The research pharmacy will 
contact the study team to establish an appropriate shipping schedule (timing, number of doses 
necessary to last until the next scheduled visit) to re-supply participants. All bottles of study 
medication currently include a label indicating that the appearance of medication may be 
different, so a change of manufacturer and possible change in the appearance of the study drug is 
unlikely to raise concerns among the study participants.  Procedures are already in place at the 
site to reduce the risk of inadvertent unblinding of the study team to treatment assignment based 
on appearance of the study medication.   
 
4. 
Safety and Protocol Deviation Reporting 
All current safety and protocol deviation reporting for the study remains in place, per protocol 
requirements.   
 
5. 
 Reporting Study Changes to Participants 
The study staff will be in regular contact with participants to make them aware of changes in the 
study, their dose of study medication, or any changes to their visit follow-up schedule. 

 
 
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Participants have the option to opt out of continued participation in the study.  Any subject 
withdrawals will be reported to the IRB as required.  
 
The study team will be in contact with participants as well, to notify them of expected shipments 
of study medication, alert them to changes in the amount of study medication they may receive, 
provide instruction for reducing (and ultimately increasing) their dose of study medication, and 
review instructions related to dose timing, drug monitoring and return.  Participants who have 
had their dose of study medication reduced will be asked to follow the initial dose titration 
schedule once study medication becomes available, with the goal to return them to their prior 
maximally tolerated dose.   
 
Communications between the study team and participants will generally be accomplished by 
telephone call and documented in the study source documents/case-report forms.  Participants 
have the option to opt out of continued participation in the study.  Communications with 
participants will be summarized in the source documents.  Any subject withdrawals will be 
reported to the IRB as required.