Inforatio Technique to Promote Wound Healing of Diabetic Foot Ulcers

Inforatio Technique to Promote Wound Healing of Diabetic Foot Ulcers: a Parallel-group, Evaluator-blinded, Randomized Clinical Trial

This randomized clinical trial will examine the effect of inforatio technique on healing of diabetic foot ulcers (DFUs). Inforatio technique is a novel procedure developed by the research group. The definition of inforatio technique is application of small cuts in wound beds with punch biopsy tools without involving surrounding epithelia. The aim is to initiate an acute inflammatory response that will promote healing of the ulcers.

BACKGROUND 15-25% of diabetic patients develop chronic foot ulcers - a serious comorbidity that widely affects quality of life and is associated with increased mortality and morbidity. 50-60% of DFUs become infected, and 20% of patients with infected DFUs undergo lower extremity amputation. Inforatio technique is a novel treatment of DFUs where small cuts are made in wound beds with punch biopsy tools without involving epithelia. The aim is to cause controlled bleeding and initiate an acute inflammatory response that may promote healing. The research group has conducted a preceding feasibility trial that showed promising results (see study by Moeini et al. in the reference list). Inforatio technique may become a supplemental treatment option for diabetic patients worldwide if it shows a beneficial effect on healing. TRIAL AIMS This trial aims to assess whether the proportion of ulcers that heal within 20 weeks is higher when DFUs are treated with inforatio technique in addition to usual wound care compared to treatment with usual wound care alone. RECRUITMENT Patients are eligibility-screened and identified by the investigators and wound care staff when patients visit the recruiting wound outpatient clinics. INCLUSION AND FOLLOW-UP Eligible patients are included if they give their oral and written consent to participate. The baseline trial visit takes place on the day that patients give their consent to participate. The trial comprises a 20-week clinical follow-up. Follow-up visits will be at 3, 6, 9, 12, 16 and 20 weeks after baseline (give or take 7 days for midtrial visits and from 20 to 22 weeks after baseline for the last trial visit). Follow-up end whenever the following appears; 20-week follow-up visit, death, amputation, or the outpatient clinic staff observes healing. The trial will be stopped if the intervention shows an unintended effect or potential harm. RANDOMIZATION Allocation of participants is conducted by block randomization with stratification by center and will be generated with Research Electronic Data Capture (REDCap)©. Block sizes will be randomly alternating between two and four. The allocation will immediately be revealed to the participant and wound care staff. Adequate allocation concealment is ensured by the irreversibility of randomization and the inability to predict the next allocation assignment in RedCap. STATISTICAL ANALYSIS PLAN Baseline characteristics will be reported with appropriate descriptive statistics. The analyses of primary and secondary outcomes will be performed on the intention-to-treat population with participants being analyzed as randomized regardless of the treatment received. Supplementary analyses will be performed on the per-protocol population, which excludes participants with one or more serious protocol violations as defined below: Participants not receiving their allocated treatment. Participants with inadequate adherence, which is defined as the following: Participants that miss more than 3 of the 6 follow-up trial visits. Participants in the intervention group that miss more than 2 of the 4 trial visits where inforatio is applied. The primary analysis of the trial will be a logistic regression for proportion of healed ulcers with adjustment for center of recruitment. EQ-5D-5L and Wound-QoL data will be presented descriptively in line with the guidelines for the questionnaires. A multivariate linear regression adjusting for baseline score will be performed for analysis of change in EQ-5D-5L index score and Wound-QoL global score. Safety outcome will be descriptively reported and analyzed on 'as treated' basis where participants are grouped according to whether they received inforatio technique. Mean time and range from the last inforatio applied to an event is detected will be reported for the intervention group. A statistical comparison between groups will be conducted by chi square test and estimates of relative risk with 95% confidence interval. Inforatio-related adverse events will be descriptively reported. An interim analyses will be performed after 50 participant has completed follow-up. Significance is set at p-value <0.05 and p-values are two-tailed. Underlying statistical assumptions for linear and logistic regressions will be assessed graphically. HANDLING OF MISSING DATA Data on Wound-QoL global score that is missing due to death during follow-up, will not be considered in the handling of missing data. Otherwise, missing data on primary and secondary outcomes will be handled according to recommendations by Jakobsen et al. (see reference list). If the proportion of missing data on an outcome is <5%, the primary analysis of missing data will be complete-case analyses. If more than 5% data of an outcome is missing, the primary analysis will include missing data imputed by multiple imputation technique and complete case analysis will be performed as a sensitivity analysis. In both cases, a best-worst and worst-best case analysis will be conducted for missing data on healing. SAMPLE SIZE A meta analysis by Margolis et al. (see reference list) has reported a healing rate of 30.9% (95% CI 26.6-35.1) in 20 weeks for non-infected DFUs that were treated with standard wound care. A healing rate of 60% in 20 weeks was suggested by the preceding feasibility trial when patients receive inforatio technique in addition to usual wound care. When the power is 80% and the alpha-level is 5%, a power calculation gives a sample size of 84 for comparison of two proportions when the outcome proportion is 30% for the control group and the aim is to assess whether the outcome improves with 30 percentage points in the intervention group. To allow an attrition of 20%, the aim is to recruit 100 participants for the trial. ETHICAL CONSIDERATIONS AND RISK ASSESSMENT OF SIDE EFFECTS Inforatio technique is a minimal invasive procedure. Thus, no adverse events are expected as a result of inforatio technique except from a low risk of pain during application. In the preceding feasibility trial, no intervention-related adverse events were observed and participants did not experience pain during application. Furthermore, inforatio technique showed a promising effect on healing. The clinical assessment of the wound care staff, patient preference and patient tolerance is taken into consideration before inforatio technique is applied. If the inforatio technique shows a significant positive effect on healing, it would become of great importance for future patients as a supplemental treatment to standard wound care of DFUs. COMPETING INTERESTS The trial investigators declare that they have no competing interests.

Healing is assessed on digital ulcer images by assessors that are blinded to treatment allocation. The digital images will be blinded by the primary investigator before delivery to the assessors. Data analysts will also be blinded to treatment allocation. It is not possible to blind participants or the staff that perform wound care and clinical assessment of healing because inforatios are visible in the wound bed after application.

Participants that are randomized to the intervention group will receive both inforatio technique and usual care. Inforatio technique will be applied at baseline, 3, 6, 9 and 12 week- follow-up as long as the ulcers have a diameter of minimum four mms and have not developed infection, necrosis, positive probe-to-bone test, exposure of joint or tendon; or underlying osteomyelitis. In addition, inforatio technique will not be applied on ulcers that are covered by scab if the wound care staff assess that the scab should not be removed from the ulcer.

Inforatio technique is a novel minimal invasive procedure where small cuts are made on wound beds with punch biopsy tools near the wound edges but without involving wound edge epithelia. Inforatio technique is applied after the wound has been surgically debrided for slough and devitalized tissue.

Healing is defined as complete epithelialization without any discharge from the site of the index ulcer. The primary outcome is the proportion of ulcers in each allocation group where healing is observed based on blinded assessment of digital images from the participants' last trial visit. The assessment of healing on digital images will be performed separately by two blinded assessors. Digital images where disagreement occurs are discussed between the assessors until agreement is reached. Any inconsistencies between the blinded assessment of healing on images and unblinded clinical assessment at trial visits will be reported. In case of death and amputation the primary outcome is registered as non-healing.

The EQ VAS score is used to assess change in participant-rated health from baseline to end of follow-up. Participants fill out the Danish version of the EQ-5D-5L questionnaire at baseline and the last trial visit or after 20 weeks from the day of amputation if participants undergo amputation of their index limb during follow-up. EQ-5D-5L is a general health patient-reported outcome measure that includes a visual analog scale and a descriptive 5-dimension system. Permission to use EQ5-5D-5L (registration ID 39403) has been obtained. In case of death, the EQ VAS score will be registered as missing.

Wound-QoL global score is used to assess change in disease-specific quality of life from baseline to end of follow-up. Participants fill out the Danish version of the Wound-QoL questionnaire at baseline and the last trial visit. The Wound-QoL is a disease-specific patient-reported outcome measure that consists of 17 items, which each has a score from 0 to 4 - 0 indicates no problem and 4 is the highest score for an ulcer-related problem. The global Wound-QoL score is an average of all item scores. In case of death, the Wound-Qol score will be registered as missing. In case of amputation of the index extremity, the Wound-QoL global score is 4, which represents the worst possible outcome for Wound-QoL. The Wound-QoL response is defined as missing if less than 13 of the 17 items have been completed. Permission to the Danish version of the Wound-QoL has been obtained.

Inforatio-related adverse events will be descriptively reported. Relatedness to inforatio technique will be qualitatively assessed as definitely related, probably related, probably not, or definitely not related. An event is assessed as probably not related to inforatio technique when the event is most likely to be explained by side effects from other treatments; a natural course of DFUs; or the clinical condition of the participant.

Following adverse events related to the index ulcer will be reported: Total wound area increase during follow-up Infection of the ulcer Exposure of bone, tendon or joint in the wound bed Osteomyelitis of underlying bone is diagnosed Surgical wound intervention of the ulcer in an operating theater Hospitalization related to the ulcer Minor amputation of the index extremity (below ankle) Major amputation of the index extremity (above ankle) Mortality related to the ulcer The events will be descriptively reported and analyzed on 'as treated' basis where participants are grouped according to whether they received inforatio technique. Mean time and range from the last inforatio applied to an event is detected will be reported for the intervention group. A statistical comparison between groups will be conducted for proportion of participants experiencing one or more adverse events related to the index ulcer.

Patient-reported adverse events related to index ulcers: Participants are asked at each follow-up visit whether they have experienced onset of an ulcer-related adverse events since their last trial visit. Adverse events related to inforatio technique application: Participants from the intervention group are asked after each inforatio technique application whether they experience any adverse events during or immediately after application.

Serious adverse events are defined according to FDA (U.S. Food and Drug Administration) as death, life-threatening events, hospitalization, disability or permanent damage. The outcome will be descriptively reported and analyzed on 'as treated' basis where participants are grouped according to whether they received inforatio technique. A statistical comparison between groups will be conducted for proportion of serious adverse events.

INCLUSION CRITERIA Age ≥ 18 years Diabetes mellitus Non-surgical ulcer located distal to the malleoli Wound diameter >4 millimeters Patient-reported wound duration ≥ 6 weeks Only one ulcer will be included from each participant. If a patient has more than one eligible ulcer, the largest ulcer is included. In case of equally sized ulcers, the ulcer with the most recent onset is included. EXCLUSION CRITERIA Dementia or other reasons that cause inability to give informed consent Malignant disease Current treatment with systemic immunosuppressive drugs. Exclusion criteria related to the index extremity: Diagnosed with or awaiting evaluation of suspected acute phase Charcot arthropathy or osteomyelitis Non-palpable pulse in both arteria dorsalis pedis and arteria tibialis posterior accompanied by systolic toe pressure <30 mmHg in the index foot. Amputation at midfoot level or proximal to midfoot level Revascularization procedure awaits or has been undertaken within the last 8 weeks or the patient awaits a vascular surgeons' decision on revascularization Gangrene Exclusion criteria related to the index ulcer Infection of the ulcer defined according to IWGDF/IDSA (International Working Group on the Diabetic Foot/ Infectious Diseases Society of America system) classification as presence of at least 2 of the following; 1) local swelling or induration; 2) erythema >0.5 to ≤2 cm around the ulcer; 3) local tenderness or pain; 4) local warmth; or 5) purulent discharge. Current antibiotic treatment due to infection of the index ulcer will also be considered as an ongoing infection regardless of presence of clinical signs of infection. Positive probe-to-bone test Exposed joint or tendon The soft tissue layer covering bone or joint is evaluated to be too thin to allow for inforatio technique anywhere in the wound bed. Interdigital ulcer location

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