# Intact Fish Skin Graft for Deep Diabetic Foot Ulcers: results from the KEREFISH randomized controlled trial

Dured Dardari, Louis Potier, Ariane Sultan, Maud Francois, Jocelyne M'bemba, Benjamin Bouillet, Lucy Chaillous, Laurence Kessler, Aurélie Carlier, Abdulkader Jalek, et al.

<figure><img src="image_1.png" /></figure>

## To cite this version:

Dured Dardari, Louis Potier, Ariane Sultan, Maud Francois, Jocelyne M'bemba, et al. Intact Fish Skin Graft for Deep Diabetic Foot Ulcers: results from the KEREFISH randomized controlled trial. Diabetes & Metabolism, 2025, 51 (6), pp.101691. 10.1016/j.diabet.2025.101691. hal-05190975

HAL Id: hal-05190975

https://hal.science/hal-05190975v1

Submitted on 31 Jan 2026

HAL is a multi-disciplinary open access archive for the deposit and dissemination of scientific research documents, whether they are published or not. The documents may come from teaching and research institutions in France or abroad, or from public or private research centers.

L'archive ouverte pluridisciplinaire HAL, est destinée au dépôt et à la diffusion de documents scientifiques de niveau recherche, publiés ou non, émanant des établissements d'enseignement et de recherche français ou étrangers, des laboratoires publics ou privés.

<figure><img src="image_2.png" /></figure>
# Intact fish skin graft for deep diabetic foot ulcers: results from the KEREFISH randomized controlled trial

Dured Dardari ¹, Louis Potier ², Ariane Sultan ³, Maud Francois ⁴, Jocelyne M'Bemba ⁵,
Benjamin Bouillet ⁶, Lucy Chaillous ⁷, Laurence Kessler ⁸, Aurélie Carlier ⁹, Abdulkader
Jalek ¹⁰, Ayoub Sbaa ⁸, Hilmar Kjartansson ¹¹, Baldur Tumi Baldursson ¹¹, Guillaume
Charpentier ¹², John C Lantis ¹³

¹ Diabetic Foot Unit, Centre Hospitalier sud Francilien Corbeil Essonnes, 91100 Corbeil-Essonnes, France; LBEPS, IRBA, Université Evry Paris Saclay, 91025 Evry, France. Electronic address: dured.dardari@chsf.fr.

² Diabetology Department, CHU Bichat-Claude Bernard, AP-HP, 75018 Paris, France; Institut Necker-Enfants Malades, Université Paris Cité, INSERM UMR-S1151, CNRS UMR-S8253,75006 Paris, France.

³ Diabetology Nutrition Department, CHU Montpelier, Université de Montpellier, 34090 Montpellier, France; Inserm, CNRS, Phymedexp, CHU de Montpellier, 34090 Montpellier, France.

⁴ Diabetology Department, CHU Reims, 51100 Reims, France.

⁵ Department of Diabetology, CHU Cochin, 75000 Paris, France.

⁶ Department of Endocrinology-Diabetology, Dijon University Hospital, 21000 Dijon, France; INSERM Unit, LNC-UMR 1231, University of Burgundy, 21078 Dijon, France.

⁷ Department of Endocrinology, Diabetology and Nutrition, Institut du Thorax, University Hospital of Nantes, 44000 Nantes, France.

⁸ Department of Diabetology, CHU Strasbourg, 67000 Strasbourg, France.

⁹ Diabetology Department, CHU Bichat-Claude Bernard, AP-HP, 75018 Paris, France.

¹⁰ Diabetology Nutrition Department, CHU Montpelier, Université de Montpellier, 34090 Montpellier, France.

¹¹ National University Hospital, Reykjavik, Iceland.

¹² CERITD (Center for Study and Research for Improvement of the Treatment of Diabetes), Bioparc-Genopole Evry-Corbeil, 91042 Evry, France.

¹³ Mount Sinai Hospital, NY, NY, USA.

**Keywords:**

Diabetic foot; Intact fish skin graft; Multinational randomized trial.
# Abstract

**Introduction:**

The main objective : To assess the efficacy of Intact fish skin graft (IFSG) for the closure of University of Texas (UT) Grade 2 and 3 Diabetic foot ulcers (DFUs) versus local standard of care (SOC).

**Methods:**

In the French subgroup of a multinational randomized trial, 180 (179 in primary endpoint analysis) patients with UT grade 2 and 3 DFUs (8 centers) were randomized to receive IFSG or SOC, that adhered to the International Working Group on the Diabetic Foot (IWGDF) guidelines. Primary endpoint was complete epithelialization at 16 weeks. Secondary endpoints were healing curve, percentage of wounds healed to 80 % or more in an average of 16 weeks, percentage healed at 20 weeks.

**Results:**

The primary endpoint was 41.6 % closure rate in IFSG group versus 22.2 % in SOC group (P = 0.0053). In the intent to treat analysis (ITT), there was a statistically significant difference (P < 0.05) in mean relative wound area between the IFSG and SOC arms at weeks 6. The proportion of patients with complete epithelialization at 20 weeks was 2.11 times higher in the IFSG group than in the SOC group. For those patients that healed a median of 7 graft applications was required.

**Conclusions:**

In France, the addition of IFSG to the care plan of patients with deep diabetic foot wounds improved the closure rate by 41.6 % in IFSG group versus 22.2 % in SOC.
# Introduction

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 neo- dermis, 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.

## Research design and methods

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 clinicaltrials.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, Isafjörður, 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.

## Patients

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.

## Procedures

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 Inter-national 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.

## Statistical analysis

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 UTI 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.

# Results

## Patients

Between July 2020 and October 2022, a total of 180 patients, randomized equally between the experimental group receiving IFSG (n = 90) and the standard of care (SOC) group (n = 90). One patient randomized to the IFSG but never received IFSG. Baseline characteristics of the participants and DFUs of each group were presented in Table I and Table II. The overall mean age was 67.2 ± 11.5 years, with a median of 68.5 years. Age distribution was similar between the two groups, with the IFSG group having a mean age of 66.4 ± 11.7 years and the SOC group 68.0 ± 11.4 years. Race and ethnicity reporting is prohibited for European populations.

<table><caption>**Table I**

Characteristics of participants at baseline.</caption><thead><tr><th>Characteristic</th><th>IFSG Group (n = 90)</th><th>SOC Group (n = 90)</th><th>Total (n = 180)</th></tr></thead><tbody><tr><td>Sex, n (%)</td><td></td><td></td><td></td></tr><tr><td>Male</td><td>75 (83.3 %)</td><td>68 (75.6 %)</td><td>143 (79.4 %)</td></tr><tr><td>Female</td><td>15 (16.7 %)</td><td>22 (24.4 %)</td><td>37 (20.6 %)</td></tr><tr><td>Age, years</td><td></td><td></td><td></td></tr><tr><td>Mean ± SD</td><td>66.4 ± 11.7</td><td>68.0 ± 11.4</td><td>67.2 ± 11.5</td></tr><tr><td>Median (Q1-Q3)</td><td>67.0 (58.0-74.0)</td><td>69.0 (60.0-75.0)</td><td>68.5 (59.5-75.0)</td></tr><tr><td>Range</td><td>40-92</td><td>38-93</td><td>38-93</td></tr><tr><td>BMI, kg/m²</td><td></td><td></td><td></td></tr><tr><td>Mean ± SD</td><td>27.4 ± 6.2</td><td>29.3 ± 6.2</td><td>28.3 ± 6.2</td></tr><tr><td>Underweight (&lt;18.5), n (%)</td><td>4 (4.7 %)</td><td>3 (3.5 %)</td><td>7 (4.1 %)</td></tr><tr><td>Normal weight (18.5-24.9), n (%)</td><td>31 (36.5 %)</td><td>17 (20.0 %)</td><td>48 (28.2 %)</td></tr><tr><td>Overweight (25-29.9), n (%)</td><td>26 (30.6 %)</td><td>24 (28.2 %)</td><td>50 (29.4 %)</td></tr><tr><td>Obese (≥ 30), n (%)</td><td>24 (28.2 %)</td><td>41 (48.2 %)</td><td>65 (38.2 %)</td></tr><tr><td>Patient Setting, n (%)</td><td></td><td></td><td></td></tr><tr><td>Hospitalized</td><td>38 (42.2 %)</td><td>46 (51.1 %)</td><td>84 (46.7 %)</td></tr><tr><td>Outpatient</td><td>52 (57.8 %)</td><td>44 (48.9 %)</td><td>96 (53.3 %)</td></tr></tbody></table>

Note: Values are presented as mean ± standard deviation (SD), median (first quartile-third quartile), or number (percentage) as appropriate.
<table><caption>**Table II**

Characteristics of participants and DFU at baseline.</caption><thead><tr><th>Characteristic</th><th>Experimental Group - n = 90</th><th>Conventional Group - SOC n = 90</th><th>Total</th></tr></thead><tbody><tr><td>UT score 2: tendon or capsule involvement [n (%)]</td><td>43 (47.2)</td><td>47 (51.1)</td><td>88 (49.2)</td></tr><tr><td>UT score 3: bone or joint involvement [n (%)]</td><td>46 (51.7)</td><td>44 (48.9)</td><td>90 (50.3)</td></tr><tr><td>ABPI</td><td>0.9 (0.2)</td><td>0.9 (0.2)</td><td>0.9 (0.2)</td></tr><tr><td>&lt;= 0.9 [n (%)]</td><td>40 (44.4)</td><td>42 (46.7)</td><td>82 (45.6)</td></tr><tr><td>&gt; 0.90 [n (%)]</td><td>50 (55.6)</td><td>48 (53.3)</td><td>98 (54.4)</td></tr></tbody></table>

Note: Values are presented as mean ± standard deviation (SD), median (first quartile–third quartile), or number (percentage) as appropriate.

Eighty two percent of the IFSG group completed the trial while 74.4 % of the SOC group completed the trial. Almost all patients (97.2 %; n = 175) had at least one diabetic complication or comorbidity. Half (50 %, n = 86) had retinopathy, 11.2 % (n = 15) had associated maculopathy, 53.3 % of patients (n = 90) had abnormal renal function, 90.3 % (n=158) had neuropathy and 73.6 % (n = 128) had macroangiopathy.

Most patients were male (79.4 %), with a slightly higher proportion in the IFSG group compared to the SOC group (83.3 % vs. 75.6 %). BMI categorization revealed that 67.6 % of patients were overweight or obese (BMI ≥ 25 kg/m²). Specifically, 28.2 % of patients in the IFSG group and 48.2 % in the SOC group were classified as obese (BMI ≥ 30 kg/m²). Underweight patients (BMI < 18.5 kg/m²) constituted 4.1 % of the total population.

At baseline, 46.7 % of patients (n = 84) were hospitalized. A higher proportion of patients in the SOC group were hospitalized compared to the IFSG group (51.1 % vs. 42.2 %). The remaining 53.3 % of patients were managed in an outpatient setting, with 57.8 % in the IFSG group and 48.9 % in the SOC group.

The mean HbA1c in IFSG is 8.0 % (1.7) versus 8.0 % (1.5) in SOC, the duration of diabetes (years) in years was 19.6 (11.5) for the IFSG group versus 21.4 (13.8) for the SOC.

Offloading assessment: In the mITT population, 70.3 % of patients in the IFSG arm and 65.1 % in the SOC arm always or often used discharge at W1. At W16, these proportions were
83.9 % and 81.1 %, and at W24 they were 88.9 % and 81.5 % for the IFSG and SOC arms respectively.

An analysis of the common risk factors, (BMI, amputation status, ABPI) showed that there was no significant difference between IFSG and SOC. Center effect on results was analyzed using a logistic regression model (center 1 vs other centers). The adjusted odds ratio was 2.61 [95 % CI=1.088;6.21) in favor of the IFSG group (P = 0.0317). The center effect and interaction between the randomization arm and the center were not significant (P = 0.6409 and 0.933 respectively). Therefore, there was no center effect.

## Primary outcomes

The primary efficacy endpoint of the study was the proportion of patients achieving complete wound healing at week 16. Complete epithelialization was confirmed by examination of the last two photographs taken by the local nurse (at the time of declaration of healing and 10±3 days later) in a blinded review by Adjudication Committee (an independent assessment committee). In the intention-to-treat (ITT) population, the IFSG group demonstrated a significantly higher healing rate compared to the SOC group (Fig. 1). Specifically, 41.6 % of patients treated with IFSG achieved complete wound healing at week 16, compared to 22.2 % in the SOC group. The unadjusted odds ratio (OR) was 2.546 (95 % CI, 1.319 to 4.915, P = 0.006), indicating that patients receiving IFSG were more than twice as likely to achieve complete healing by week 16 compared to those receiving SOC. Not surprisingly in the per protocol cohort both groups did better with the IFSG treated patients having a closure rate of 48 % at week 16 versus 26.8 % (P =0.010).
<figure><img src="image_3.png" /><figcaption>Fig. 1. Percentage of patients with complete wound epithelialization at 16 weeks.</figcaption></figure>

## Secondary outcomes

At week 20, the healing rate increased to 47.2 % in the IFSG group, while the SOC group had a healing rate of 30.0 % (P = 0.022). At week 24, 52.8 % of patients in the IFSG group achieved complete wound healing compared to 37.8 % (P = 0.051) in the SOC group. Wound Area Reduction: At the beginning of the study the wound area for the IFSG cohort was larger (mean 1048.6 mm2) than the SOC arm (mean 976.1 mm2). At week 24, the median wound area was 130.0 mm2 (30.7) and 161.1 mm2 (39.7) for the IFSG and SOC arms respectively. In the mITT population, there was a statistically significant difference (P < 0.05) in mean relative wound area between the IFSG and the SOC arms at weeks 6, 7, 8, 9, 10, 11 and 12 after adjustment; wound area at baseline in mm2 ; P = 0.8614 (Fig. 2).

Time to Healing: was compared with Kaplan-Meier survival analysis. During the follow-up period through 6 months: 47 patients had wounds healed (52.8 %) in the IFSG group and 34 patients (37.8 %) in the SOC group. The median time to complete healing was 20.4 weeks (95 % CI, 15 weeks – not estimable) for the IFSG group, compared to 24 weeks (95 % CI, 23.4 weeks – not estimable) for the SOC group. Cox proportional hazards modeling demonstrated that patients treated with IFSG experienced a significantly higher probability of wound healing than those receiving SOC. For those that healed, those who were treated with IFSG healed 15 % or 25 days sooner than those in SOC.
<figure><img src="image_4.png" /><figcaption>Fig. 2. Wound area reduction over time.</figcaption></figure>

**Quality of Life:** In France, the 5 dimensions of the EQ-5D-5 L questionnaire (mobility, ability to wash and dress oneself, daily activities, discomfort and pain and anxiety), and Wound-QoL-17 quality-of-life where the levels range from 1 ("no health problems") to 5 ("extreme health problems") were collected. There was no meaningful statistical difference.

**Pain:** In the mITT population, the median pain score measured with a visual analog scale (in a 0–100 mm range) was 0 mm in both IFSG and SOC arms, at each follow-up visit.

In the IFSG cohort patients received a median of 9 grafts, however for those patients that healed by week 16 the median number was 7 graft applications.

During the study, a total of 125 serious adverse events (SAEs) were reported, all of which were anticipated and associated with either the underlying condition being treated or concomitant diseases. These SAEs included: 9 deaths, 94 hospitalizations (50 due to aggravation of lower limb conditions and 44 related to a deterioration in general health or other comorbidities), 12 medically significant events as determined by investigators, and 13 lower limb amputations.
In the IFSG group (n = 89), 61 patients (68.5 %) experienced at least one adverse event (AE), and 43 patients (48.3 %) had at least one SAE. Wound-related amputations occurred in 5 patients (5.6 %), and 7 patients (7.9 %) developed bone infection within four weeks of initiating treatment. There were 3 deaths (3.3 %) in this group: one patient died of diabetic foot ischemia, and two patients died from unknown causes reported as "faintness" and "patient death" by the investigators.

In the SOC group (n = 90), 57 patients (63.3 %) experienced at least one AE, and 42 patients (46.7 %) had at least one SAE. Wound-related amputations were reported in 8 patients (8.8 %), and osteitis occurred in 5 patients (5.6 %) within four weeks of treatment initiation. There were 6 deaths (6.6 %) in this group: none related to the study.

Overall, the incidence of adverse events was comparable between the two groups, with almost twice as many wound-related amputations (5 v 8) and fatal events (3 v 6) in the SOC group. No unexpected safety concerns related to the IFSG treatment emerged during the study, suggesting that IFSG is as safe as the standard of care for patients with chronic wounds.

## Other secondary endpoints

Medical economic analyses are currently being carried out and will be published separately.

# Discussion

This prospective, multicenter randomized control trial (RCT) is, to our knowledge, the first such completed study to assess safety and efficacy of a topically applied skin substitute in the most severe DFU types.

The use of biologic tissue grafts from various sources has been studied in chronic wounds including venous leg ulcers and DFU, and biologic tissue grafts have generally shown better outcomes than standard care [3]. The potential advantages of such products are thought to include the provision of an extracellular matrix(ECM) scaffold, the major component of the
dermis, promoting cellular and vascular infiltration, biologically active components including binding sites for cell surface receptors involved in signal transduction, binding sites for diffusible growth factors, and matricellular proteins with the ability to sequester or modulate growth factor activity [4]. In contrast to other ECM products, the gentle processing of IFSG preserves native skin components and fat content including polyunsaturated fatty acids [4].

Our findings consistently demonstrate the efficacy of IFSG over SOC in promoting wound healing among patients. The higher proportion of healed wounds, at each assessed time point, in the IFSG group underscores its efficacy. The statistically significant differences at Weeks 16 and 20, alongside the almost statistically significant result at Week 24, demonstrate the efficacy of IFSG over SOC, which provides valuable information for understanding time to wound closure and healthcare resource utilization for IFSG compared to SOC.

Local DFU therapy is a new approach recommended by the Inter-national Working Group on the Diabetic Foot [3]. Several randomized controlled studies have shown the benefits of this treatment in accelerating DFU healing [3]. However, the common characteristic of the participants included in these trials is that they have non-ischemic, non-infected DFUs. In Explorer trial [5] the use of sucrose octasulfate dressings, allowed healing at week 20 (in our study, the end point was week 16) 48% in the sucrose octasulfate dressing group and 34 patients (30%) in the control dressing group (18 percentage points difference, 95% CI 5-30; adjusted odds ratio 2.60, 95% CI 1.43-4.73; P = 0.002). The lesions included in this study were grade Ic-IIc of UT versus II and III in our study. To be eligible, a patient's Ankle Brachial Pressure Index (ABPI) score had to be 0.9 or less [4].

Another clinical trial used autologous leukocytes, platelets and fibrin placed on the wound. A randomized clinical trial [6] involving 269 patients reported improved healing at 20 weeks with the patch compared to standard care consisting of adequate off-loading, wound debridement and moist dressings (34% versus 22% healing; odds ratio, 1.58; 95% CI, 1.04-2.40). Infected DFUs were not included in this trial.

The use of placenta-derived products containing growth factors and a collagen-rich extracellular matrix induces complete healing at 12-16 weeks compared to the control dressing (relative risk, 2.0; 95% CI, 1.67-2.39; P < 0.001). Infected wounds were excluded from this study and the ABPI should be greater than 0.7 [7].

Facts that until now have been kept quiet, the use of the IFSG has been cited in literature on small series of patients or on superficial wounds [8-11]. The strengths of our study include the
randomized, multi-center design, prospectively defined endpoints, and adequate sample size
with good statistical power. The utilization of the community nurses was a significant strength
of this study as well. Moreover, in contrast to our study, DFU studies typically limit
enrollment to superficial ulcers with good perfusion ignoring the reality that many DFU are
more severe and on poorly perfused limbs which limits generalizability.

We also purposely allowed investigators flexibility in their selection of appropriate standard
of care dressings, recognizing the idiosyncratic nature of each DFU in a setting where no
universally accepted “standard” exists and wound care provider selection of a dressing is
influenced by their clinical experience and medical judgment. While this may have introduced
some variability in the SOC treatment group, we believe it likely gave patients randomized to
SOC the best chance for healing, and puts the results obtained with IFSG in a more realistic
context. As noted previously the absence of a center affect supports the fact that this local
standard of care was not detrimental to the study. We set the trial endpoint at 16 weeks
instead of the more usual 12 weeks to better assess the healing trajectory and the ultimate
proportion achieving healing in this population of chronic, severe, difficult to heal DFU.

Limitations

This French study was conducted during Covid-19 with a decentralized model. Three days
after the first patient was enrolled the French healthcare system went into a “lockdown.” This
decentralized model allowed for the patients to be enrolled and to remain in the study. The
data presented represent a more decentralized model which in- corporates community care
over a physician based model. Another limitation was we set the maximum weeks of therapy
to 14 with a maximum number of applications of IFSG set at 10. It is possible that further
intact fish skin graft on some of the patients who were healing but had not healed would have
impacted the healing rate even further.

Limited ABPI to 0.6 is one of the limitations of this study; wounds with severe ischemia
remain a major challenge for local therapy. The non- publication of treatment costs at this
moment is also an additional limitation.
**Data sharing statement**

The datasets generated during and/or analyzed in the current study are available from the
corresponding author upon reasonable request.

CRediT authorship contribution statement

**Dured Dardari:** Writing – original draft, Methodology, Investigation, Funding acquisition, **Formal analysis, Conceptualization.**

LouisPotier: Investigation. Ariane Sultan: Investigation.

Maud Francois: Investigation.

Jocelyne M'Bemba: Investigation.

Benjamin Bouillet: Investigation.

Lucy Chaillous: Investigation.

Laurence Kessler: Investigation.

Aurélie Carlier: Investigation.

Abdulkader Jalek: Investigation.

Ayoub Sbaa: Investigation.

Hilmar Kjartansson: Methodology, Investigation.

Baldur Tumi Baldursson: Methodology, Investigation, Data curation.

Guillaume Charpentier: Investigation, Funding acquisition.

John C Lantis: Funding acquisition, Data curation, Conceptualization.

Declaration of competing interest

All authors declare that they have no conflict of interest with the study.
## Acknowledgements

This paper has been produced with the assistance of the European union. The contents of the article or the sole responsibility the authors and can no way be taken to reflect the views of the EU or the INEA Agency.

## References

[1] Santema TB, Lenselink EA, Balm R, Ubbink DT. Comparing the Meggitt-Wagner and the University of Texas wound classification systems for diabetic foot ulcers: inter-observer analyses. Int Wound J 2016;13:1137-41. https://doi.org/10.1111/iwj.12429.

[2] Dardari D, Piaggesi A, Potier L, Sultan A, Diener H, Francois M, et al. Intact fish skin graft to treat deep diabetic foot ulcers. NEJM Evid 2024;3:EVIDoa2400171. https://doi.org/10.1056/EVIDoa2400171. Erratum in: NEJM Evid 2025;4: EVIDx2500032.

[3] Schaper NC, van Netten JJ, Apelqvist J, Bus SA, Fitridge R, Game F, et al. Practical guidelines on the prevention and management of diabetes-related foot disease (IWGDF 2023 update). Diabetes Metab Res Rev 2024;40(3):e3657. https://doi.org/10.1002/dmrr.3657. MarEpub 2023 May 27.

[4] Cazzell SM, Lange DL, Dickerson Jr JE, Slade HB. The management of diabetic foot ulcers with porcine small intestine submucosa tri-layer matrix: a randomized controlled trial. Adv Wound Care (New Rochelle) 2015;4:711-8. https://doi.org/10.1089/wound.2015.0645.

[5] Roberts DD. Emerging functions of matricellular proteins. Cell Mol Life Sci 2011; 68:3133-6.
[6] Edmonds M, Lázaro-Martínez JL, Alfayate-García JM, Martini J, Petit JM, Rayman G, et al. Sucrose octasulfate dressing versus control dressing in patients with neuroischaemic diabetic foot ulcers (Explorer): an international, multicentre, double-blind, randomised, controlled trial. Lancet Diabetes Endocrinol 2018;6: 186–96. https://doi.org/10.1016/S2213-8587(17)30438-2. Erratum in: Lancet Diabetes Endocrinol. 2018 Apr;6(4):e4. 10.1016/S2213-8587(18)30082-2.

[7] Game F, Jeffcoate W, Tarnow L, Jacobsen JL, Whitham DJ, Harrison EF, et al. LeucoPatch II Trial Team. LeucoPatch system for the management of hard-to-heal diabetic foot ulcers in the UK, Denmark, and Sweden: an observer-masked, randomised controlled trial. Lancet Diabetes Endocrinol 2018;6:870–8. https://doi.org/10.1016/S2213-8587(18)30240-7.

[8] Lavery LA, Fulmer J, Shebetka KA, Regulski M, Vayser D, Fried D, et al. Grafix Diabetic Foot Ulcer Study Group. The efficacy and safety of Grafix for the treatment of chronic diabetic foot ulcers: results of a multi-centre, controlled, randomised, blinded, clinical trial. Int Wound J 2014;11:554–60. https://doi.org/10.1111/iwj.12329.

[9] Dardari D, Lequint C, Jugnet AC, Bénard T, Bouly M, Penfornis A. curing necrotic angiodermatitis with an intact fish skin graft in a patient living with diabetes. Medicina (Kaunas) 2022;58:292. https://doi.org/10.3390/medicina58020292.

[10] Jugnet AC, Benard T, Lequint C, Bobony E, Pieheiro AR, Winther T, et al. Intact fish skin graft a new hope for the treatment of diabetic foot ulcers: a case report. World J Diabetes 2024;15:2353–9. https://doi.org/10.4239/wjd.v15.i12.2353.

[11] Ciprandi G, Kjartansson H, Grussu F, Baldursson BT, Frattaroli J, Urbani U, Zama M. Use of acellular intact fish skin grafts in treating acute paediatric wounds during the Covid-19 pandemic: a case series. J Wound Care 2022;31:824–31
<figure><img src="image_5.png" /></figure>