In 2017, a group of clinicians and researchers met to discuss the fact that diabetic foot ulcers (DFU) in France may benefit from cellular and/or tissue products (CTPs). However, the DFU that would benefit the most were those that were deep and had recently undergone debridement. The group was aware that most CTP studies addressed relatively easy to heal University of Texas (UT) [1] grade 1 ulcers. This group decided to design a study in which wild caught North Atlantic Cod (Gadus morhua) Fish skin, which supporting the formation of a well-vascularized neodermis, could significantly impact deeper and more difficult to heal DFUs. A protocol was designed that would initially start in France, and eventually become a four Western European country, prospective randomized controlled trial.

Primary endpoint of the complete multinational 255 patient trial has previously been published [2]. However, the unique French data has not been published. The French cohort contributed to 70 % (180/255) of the overall European cohort.

The primary goal of the study was to observe the effect of an Intact fish skin graft (IFSG) being applied to deep diabetic foot wounds healing rates at 16 weeks. The secondary endpoints included; wound area reduction (healing curve), wounds healed greater than 80 % at 16 weeks, percentage of wounds healed at 6 months, mean healed rate in both groups, percentage reduction in wound surface area at 16 weeks, for unhealed wounds assessment of University of Texas grade change over time, number of additional debridements performed in hospital for each group, comparison of pain in the two groups, off-loading assessment, number of planned and unplanned visits to the hospital foot clinic, number of home care visits by the local nurse, number of IFSG applications. In addition, measuring patient and caregiver satisfaction, cost of treatment which is an auxiliary study, and assessment of IFSG safety.

The first patient was enrolled on July 9, 2020 which coincided with France and to some degree the rest of the world undergoing a societal and medical lock down for Covid −19.

The study in France took place in 2 phases, both were affected significantly by the Covid-19 pandemic. The 1st feasibility phase was at 2 pilot centers where a total of 20 patients were included and randomized, the 2nd phase involved the role out to an additional 6 centers. The goal of the trial was to compare the outcomes of patients with UT grade 2 and 3 DFUs treated with IFSG versus the local standard of care.

The study carried out in France had differences in the nature of the cohort and the treatment paradigm. Its originality was to set up a decentralized model with the majority of IFSGs placed in the community care environment. Therefore, we present the unique and robust experience and results of using IFSG in France for the treatment of grade 2 and 3 DFUs , the number of participants (180) included in this study represents a notable sample for a randomized controlled study in the treatment of DFUs.

The French portion of the trial was titled Kerefish. Originally 6 French centers were identified for this open-label, randomized, parallel group, clinical trial. The primary endpoint, percentage of wounds healed at 16 weeks was assessed, with follow-up continuing through 24 weeks. The trial protocol was approved by the relevant ethics committees at each center and followed the tenets of the Declaration of Helsinki. The trial was registered on clinialtrials.gov (NCT04257370). All patients provided written informed consent. A total of 6 French sites enrolled a total of 180 patients. The first patient was enrolled on July 9th, 2020, and the last patient visit was on December 16th 2022.

The product being evaluated is intact fish skin graft (IFSG) (Kerecis Limited, Ísafjörður, Iceland) which is a (CE Mark, FDA cleared) medical device sourced from Atlantic cod (Gadus morhua). The intact fish skin graft is decellularized, lyophilized and supplied as a sterile, intact, sheet in the EU, at the time of the study.

Inclusion criteria were 18 years of age or older with diabetes mellitus and a lower limb wound below the malleolus penetrating to tendon or capsule (UT grade 2) or to the bone or joint (UT grade 3) ) [2] that had been present for at least one month, or, with open (i.e. not closed or dehiscent) amputation wounds at or below the malleolus. Non-ischemic to moderately ischemic wounds were included as assessed by an ankle to brachial systolic pressure index (ABPI) of at least 0.6. Patients were excluded if they had active, unmanaged osteomyelitis, immune deficiency or autoimmune disease, had undergone arterial reconstruction within the previous month, were under treatment with systemic glucocorticoids or other treatments known to delay wound healing, or had a known allergy to fish. Pregnant or breastfeeding women, or women planning to be pregnant were excluded. Almost all patients in France (97.2 %, n = 175) had at least one complication of diabetes or co-morbidity.

Eligible patients were randomly assigned 1:1 to either intact fish skin graft (ISFG) or standard of care (SOC). Randomization was stratified by wound type (non-amputation or amputation) and by ABPI (≤ 0.9 and > 0.9). Wound care staff that applied the trial treatments were unblinded due to the differences in appearance of the intact fish skin graft product and standard of care dressings.

Beginning at Week 1, all patients underwent aggressive surgical wound debridement. In most to all cases this was fundamentally performed in a operating theater environment. In the treatment group, IFSG was then applied to the still bleeding wound, and covered with a secondary foam dressing (e.g., Allevyn non-adhesive, Smith & Nephew, Watford, UK). Dressings were not to be changed for a minimum of 48 h. The control arm (SOC) patients underwent surgical debridement in a similar environment and the application of a suitable dressing chosen by the investigator including hydrocolloid, alginate, hydrocellular, charcoal, sucrose octasulfate, silver, petrolatum gauze, or pressure dressings, the dressing regime as well as the offloading conformed to the International Working Group on the Diabetic Foot’s Guidelines (IWGDF) [2]. The quality of off-loading was confirmed by wound care nurses on a weekly basis. Patients were actively treated for up to 14 weeks and followed until week 24 or until wound closure. IFSG was applied weekly at visits 1 through 6, and then every other week through week 14, for a maximum number of 10 applications. In this French cohort, wounds were treated by both physicians and nurses trained in wound care, according to standard guidelines. The hospital investigator assessed the wound status of both treatment groups during hospital visits at weeks 7, 16, 20 and 24. Additional visits with the hospital investigator were scheduled as needed. The de-centralized model allowed for a study wound nurse who trained and oversaw skilled regional wound nurses who further monitored and directed community nurses. These nurses were extensively trained on the application of the IFSG. The initial applications were performed by wound care nurses, with subsequent applications of IFSG performed by community nurses under the supervision of the wound care nurse. During the hospital stay, wound care nurses were responsible for wound changes, and once discharged home, community nurses were responsible for secondary dressing changes, under the supervision of the wound care nurse and investigator.

The primary efficacy endpoint was the percentage of wounds achieving complete closure by 16 weeks (2 weeks after the last application of intact fish skin graft or standard of care). Complete closure was defined as re-epithelialization without drainage or need for dressing. Wound closure was confirmed by the investigator, and adjudicated, through photographs, by a blinded adjudication committee. Safety was assessed through incidence of adverse events.

Secondary endpoints included in this report were the percentage of wounds healed at 20 and 24 weeks, time to healing, and wound area reduction.

The primary endpoint was analyzed using Fisher’s exact test. This test was chosen to remain conservative and to avoid distributional assumptions. To provide a more accurate estimate of the odds ratio (OR) for wound healing, we also performed a logistic regression adjusting for UT score at baseline. The secondary endpoints, which include the percentage of wounds healed at 20 and 24 weeks, were analyzed in the same manner as the primary endpoint.

The modified intention-to-treat population (mITT), which included all randomized patients in France meeting eligibility criteria and with at least one post-randomization evaluation of the wound surface, was used for all efficacy analyses. One patient, randomized to the IFSG group, did not receive the treatment due to premature discontinuation of follow-up, caused by an increase of >25 % in wound area.

For the analysis of time to healing, we utilized the Kaplan-Meier method, with differences assessed using the log-rank test, based on the subgroup of mITT patients with available wound area measurements in the screening visit. This was to ensure that follow-up was started at week 0 for all patients. Patients documented as having achieved full wound closure were considered healed at all subsequent timepoints for statistical analysis. We compared the mean time to healing between the two groups using median survival times and restricted mean survival times up to 24 weeks, which is the end of the follow-up period in the trial. We also performed a Cox proportional hazards analysis, adjusted for sex, age, and country—the same variables used in the analyses of the primary and secondary endpoints—to calculate the hazard ratio for the difference in the rate of wound healing between the two groups.