Objective. Fifty-five subjects with diabetes mellitus type 1 or 2 and a neuropathic, nonischemic foot ulcer were enrolled into this randomized, controlled, multicenter trial designed to examine the effects of debridement with clostridial collagenase ointment (CCO) used in conjunction with serial sharp debridement for a period of 6 weeks. Methods. Serial sharp debridement without adjunctive CCO was used in the control group. Various standard care therapies thought to support debridement by endogenous proteases were selected at the discretion of the investigators for use in the control group. The primary outcome measure of this trial was the percent change in ulcer area from baseline at the end of the debridement/treatment period (EOT) and at the end of an additional 6 weeks of follow-up (EOS). Secondary objectives were to assess wound status at EOT and EOS using a standardized wound assessment tool, and to compare the average time to closure for ulcers debrided with serial sharp debridement with and without adjunctive CCO. Results. Wound area decreased relative to baseline for both the CCO group (-68%, -61%) and the control group (-36%, -46%) at EOT and EOS, respectively. While the inter-group differences did not reach statistical significance, wound area was significantly decreased from baseline at both EOT and EOS for the CCO (P < 0.001) but not for the control group. Wound status scores (scale range 8 to 40) improved for both groups during treatment (CCO: -3.5, control: -3.2) and follow-up (CCO: -5.3, control: -6.4). No differences were observed in the number of sharp debridements (CCO: 3.7, control: 4.0). Median time to closure for wounds that healed was 6 weeks for CCO and 8 weeks for control. On average, ulcers treated with serial sharp debridement plus adjunctive CCO decreased in size more rapidly than ulcers treated without adjunctive CCO debridement. No safety issues were identified based on a review of reported adverse events. Conclusion. These results suggest there is more to wound debridement than meets the eye, and establish a foundation for larger, confirmatory studies.

  Debridement of diabetic foot ulcers (DFU) is accepted as an essential component of appropriate treatment.¹ Active methods include mechanical, surgical, biological, and enzymatic debridement, while the passive maintenance of a moist wound environment is thought by some to promote debridement by endogenous proteases. The word debridement has its origin in middle French and literally means “unbridling.” The generally accepted immediate goal of all of these methods is the removal of debris and nonviable tissue as a means to the end goal of unleashing the capacity of the wound to heal itself. In a 2006 Guidance for Industry, the US Food and Drug Administration noted “debridement of necrotic tissue generally is considered part of standard care for most ulcers and burns” and that in the assessment of the efficacy of debriding products, improved wound healing (ie, increased incidence or acceleration of complete closure) is a clinically relevant endpoint.²

Healing is a cellular process dependent not only on the signals provided by the complex molecular milieu present in the wound bed, but equally dependent on the ability of the resident cells to respond appropriately to those signals.3 With that in mind, the way in which a wound is debrided could be of greater importance than the simplistic goal of removing visible material until the wound bed appears to be clean; how that clean wound bed is achieved may matter.

Given that the various debridement modalities are fundamentally different mechanistically, it is reasonable to hypothesize the choice of debridement method or combination of methods may have clinically significant consequences that affect the progress and rate of healing.

  The present study is one of a series of exploratory, randomized clinical trials designed to provide descriptive data and generate testable hypotheses relevant to the clinical outcomes associated with debridement of DFU with clostridial collagenase ointment (CCO). Unlike other modalities of debridement, CCO debridement can be used in most patients and can be performed in any care setting and by any caregiver. This makes CCO an ideal choice not only for first-line debridement, but also for adjunctive debridement in those patients where initial sharp and/or serial sharp debridement is necessary.

Study Participants. This study was registered at ClinicalTrials.gov as NCT01408277. Eligible patients were 18 years or older with a diagnosis of type 1 or 2 diabetes mellitus requiring medications to control blood glucose levels. Key inclusion criteria were: neuropathic foot ulcers of at least 1 month’s duration measuring 0.5 cm2 to 10 cm2; ankle brachial index (ABI) > 0.7; no clinical signs or symptoms of ulcer infection; serum albumin ≥ 2.0 g/dL and serum pre-albumin ≥ 15 mg/dL; and HbA1c ≤ 12%. Patients were excluded for any of the following reasons: ulcer did not require debridement, uncontrolled bleeding disorder, infection with systemic toxicity, ulcer could not be offloaded, current osteomyelitis of the target foot, or sub-dermal tissue involvement. This study was performed in compliance with the ethical principles of the Declaration of Helsinki and Good Clinical Practice. The study protocol, investigators, and consent documents were reviewed and approved by accredited Institutional Review Boards, and all patients provided written informed consent before taking part in the study.

  Study Design. This was a randomized, parallel group, open-label, multicenter, 12-week clinical study (Figure 1), carried out at 7 sites in Arizona, California, Michigan, Texas, and Virginia. The study participants were recruited betweenAugust 2011 and October 2012. Randomization to treatment was centralized using a computer-generated sequence. Patients were evaluated for eligibility at a screening visit that included a blood draw for assessment of hematology and blood chemistry. Eligible patients reported for the baseline/randomization visit from 1 to 5 days later. The baseline wound bed assessment was carried out followed by sharp surgical debridement and baseline wound area measurement for all patients. Patients were then randomly assigned to treatment in an equal allocation ratio to either the CCO (Santyl, Smith & Nephew, Hull, UK) group or to the control group (serial sharp debridement without adjunctive CCO). Clostridial collagenase ointment was applied once daily at a thickness of ~2 mm on the DFU of patients in the CCO group. Patients randomized to the control group received daily wound care/dressings as deemed appropriate by the investigator with the exception of CCO. Hyperbaric or negative pressure therapies were not allowed for any study participants as inclusion would have complicated interpretation of the results. A categorical listing of the care provided in the control group is given in Table 1.

  In either case, treatment was given for 6 weeks and patients were followed for up to an additional 6 weeks or to complete wound closure, whichever came first. During the 6-week follow-up phase, all ulcers which had not closed received daily dressing changes consisting of a foam primary dressing (Allevyn, Smith & Nephew, Hull, UK), and a single layer of cast padding held in place with a self-adherent bandage (Coban, 3M, St. Paul, MN). At the investigators’ discretion, a hydrogel could be used if deemed necessary to maintain a moist wound environment. Sharp debridement was required for both treatment groups throughout the treatment and follow-up periods whenever the ulcer was rated ≥ 3 on any one of several designated wound assessment subscales (eg, edges, undermining, necrotic tissue type, necrotic tissue amount, see Table 2). All patients agreed to wear an offloading boot or other appropriate device.

  Patients were seen every week during the treatment and follow-up phases. Patients were instructed in daily wound cleansing with sterile saline, dressing changes, and application of either CCO or any assigned 

nonenzymatic treatment.

  Study Assessments. Wound area was measured at each study visit using a digital image capture and wound measurement system (Silhouette, ARANZ Medical, Christchurch, New Zealand). Wound appearance was assessed at each study visit using a modified version of the Bates-Jensen wound assessment tool4 which included separate assessments for wound edge appearance, undermining, necrotic tissue type, necrotic tissue amount, exudate type, exudate amount, periwound skin color, and granulation tissue appearance (Table 2).

  Neuropathy was confirmed by subject inability to perceive 10 g pressure in the periwound area using a nylon monofilament test. Adequate perfusion to the affected foot was confirmed by ABI > 0.7 or, alternatively, a TcPO2 > 40 mm Hg, a great toe pressure of > 40 mm Hg, or a Doppler waveform consistent with adequate blood flow.

  Safety was assessed through analysis of adverse events. Adverse events were collected from the date a subject provided informed consent through follow-up and exit from the study. 
 

  Between-group comparisons of the average percent change in wound area from baseline at both time points (ie, end of treatment [EOT] and end of study [EOS]) and in wound status total score was carried out using a 2-way analysis of covariance (ANCOVA) model. Paired t tests were employed to evaluate the significance of changes from baseline in wound area and wound status total score within each treatment group. Chi-square tests were used to compare demographic and baseline ulcer characteristics for the 2 treatment groups. Missing values for wound area measurements, due to missed visits or early study discontinuation, were imputed using the population mean value for a given study visit. For missing values due to complete wound closure, the missing values were imputed as 0. For scale measures (eg, wound assessment scores) missing data were imputed with the mean score for that assessment (at that visit) and rounded up to the nearest integer provided that the total number of missing subscale values did not exceed 20% of the total item number. Otherwise the assessment score was set to “missing.” All efficacy data were analyzed in the intention-to-treat population, defined as all randomized patients.

  A significance level of 0.05 was used in all statistical analyses.

 Sample size. The purpose of this study was to provide descriptive data in regard to the outcomes achieved following 6 weeks of serial sharp debridement with or without CCO debridement in the treatment of chronic DFU. Because theintent of the study was to generate rather than to test hypotheses, sample size was not based on statistical power calculations. Sample size was set at 48 so that each study site could enroll a minimum of 2 or more patients into each treatment group.

  A summary of the demographic and wound characteristics for the study population appear in Table 3. The majority of subjects were < 65 years of age (76%), male (75%), white (86%), and non-Hispanic/Latino (84%). Study wounds were generally small, with a median area of 1.5 cm2. The majority were plantar (78%), or with a plantar component (11%), and evenly distributed between the left and right side. There were no significant differences between treatment groups for any of these parameters.

 Wound area. Diabetic foot ulcers enzymatically debrided with CCO in conjunction with serial sharp debridement decreased in area from an average of 1.94 cm2 at baseline to 0.56 cm2 at EOT (P < 0.001). Over the same period, the controlgroup went from an average wound area of 1.77 cm2 to 1.17 cm2, which was not statistically significant (Figure 2). When CCO debridement was stopped at EOT, the decrease in wound area observed for the CCO group was generally maintained. Average wound area in the CCO group at EOS was 0.75 cm2 (P < 0.001). The corresponding value for the control group was 1.20 cm2. The differences in average percentage change in wound area from baseline between the CCO and control groups (between-group comparisons) at both EOT (68% vs 36%) and EOS (61% vs 46%) did not reach statistical significance.

  Wound appearance. The wound assessment tool has a score range of 8 (healthy skin) to 40. At baseline, average total scores were similar for the CCO and control groups (18.4 vs 19.1). Debridement in both groups resulted in improvement in the appearance of the wound bed. The decreases from baseline in total score were clinically relevant (approximately 0.75 units on average for each of the 8 subscales by the end of the study) and statistically significant for both groups at both EOT (-3.5 for CCO, P = 0.0028; -3.2 for control, P = 0.0444) and EOS (-5.3 for CCO, P < 0.0001; -6.4 for control, P = 0.0011). There were no statistically significant differences between the 2 groups for total score or individual subscales. The mean wound appearance scores at each assessment are shown in Figure 3.

  Sharp debridement. No statistically significant differences were noted in the number of sharp debridements at EOT (3.7 for CCO; 4.0 for control) or the study as a whole (6.7 for CCO; 6.8 for control).

  Time to closure. Median time to closure for all DFU was 9 weeks for the CCO group compared to 11 weeks for the control group.

  Considering only those DFU that achieved closure, median time to closure was 6 weeks for the CCO group and 8 weeks for the control group.

  Adverse events. A total of 38 adverse events were recorded for the 55 patients over the course of the study. These were evenly distributed between the 2 treatment groups (20 for CCO; 18 for control), and none were considered by the investigators to be related to treatment.

  Wound debridement is essentially a therapeutic intervention aimed at clearing away impediments to healing more rapidly or effectively than can be accomplished by the tissue itself. Consequently, there is general agreement that debridement involves the removal of foreign bodies, nonviable tissue, and callus.⁵ More expansive definitions have included infection control, biofilm removal,⁶inflammation reduction,^7,8 visualization and probing for tunneling and undermining, and removal of dysfunctional or senescent cells.^5,9,10 While most debridement modalities can, with varying efficiency, accomplish the removal of nonviable tissue, and rapidly so when used in conjunction with sharp methods, the accomplishment of other debridement goals is less obvious, particularly to the unaided eye. This is especially true when considering provision of the conditions for inflammation resolution or in changing the balance between senescent and nonsenescent cells. Debridement efficacy relevant to these less visible effects could be measured by tissue biopsy and sophisticated histopathological and biochemical tests but, in the absence of such testing, should also be apparent in the accelerated progress toward healing that would be expected under such conditions.

  The present study explores and contrasts the fate of DFU treated with a daily application of an enzymatic debrider, CCO, along with serial sharp debridement for 6 weeks against DFU treated with serial sharp debridement with commonly used care, including those generally employed to maintain a moist wound environment and thought to promote autolysis. Wound appearance, as assessed using a standard wound assessment tool, could not differentiate between the 2 treatment modalities. However, the significant progress toward healing that was observed for the CCO group, but not the control group, clearly implies there may be more to debridement than meets the eye (at least the unaided eye). It is reasonable to speculate that biochemical differences associated with the mode of debridement are responsible for the different outcomes. So called “autolytic” methods depend on endogenous proteases, the expression of which is a hallmark of an inflammatory state. In contrast, debridement by CCO is accomplished in part through the action of bacterial enzymes which cleave the collagen triple helix at specific sites characteristic of these enzymes and not shared by endogenous proteases.

  Enzymatic debridement with CCO in conjunction with serial sharp debridement appeared to provide a benefit beyond what can be achieved using serial sharp debridement with standard care alone. In addition, it seems desired outcomes may be achieved more rapidly when using CCO with sharp debridement than when using CCO alone. In another study, DFU debrided with CCO alone without sharp debridement for 4 weeks had a mean percent area decrease from baseline of 45% at 4 weeks11 contrasted with 68% at 6 weeks in the present study.
 

  A limitation of the present study is the unblinded design. This was chosen intentionally to allow clinically relevant standard care in the control group, with bias controlled through randomization and the use of a sophisticated wound measurement device.

  Studies of similar design, larger scale studies, and studies of other chronic wound types will be important to further examine potential differences in the various approaches to debridement.

  The authors are grateful to Dr. Yuxin Zhang for biostatistical consultation and analysis, to Dr. Craig Simpson for valuable discussion and suggestions, and to The Collagenase Plus Sharp Debridement Study Group: Patrick Agnew, DPM; Travis Motley, DPM; Neal Mozen, DPM; Russell Stanley, DPM; Arthur Tallis, DPM; Dean Vayser, DPM; Robert Wunderlich, DPM, for their contribution in the recruitment of patients and the conduct of the study.

Travis A. Motley, DPM is from the Department of Orthopaedics, University of North Texas Health Science Center, Fort Worth, TX. Darrell L. Lange, PhD is from Smith & Nephew Biotherapeutics, Fort Worth, TX. Jaime E. Dickerson, Jr, PhD is from Smith & Nephew Biotherapeutics, Fort Worth, TX; and Department of Cell Biology and Anatomy,University of North Texas Health Science Center, Fort Worth, TX. Herbert B. Slade, MD is from Smith & Nephew Biotherapeutics, Fort Worth, TX; and Department of Pediatrics, University of North Texas Health Science Center, Fort Worth, TX.

Address correspondence to:
Jaime E. Dickerson, Jr, PhD
Smith & Nephew Biotherapeutics
Medical & Clinical Affairs
3909 Hulen Street
Fort Worth, TX 76107
jaime.dickerson@smith-nephew.com

Disclosure: The authors disclose that Dr. Lange, Dr. Dickerson, and Dr. Slade are employed by Smith and Nephew Biotherapeutics. Portions of this work were previously presented at the Symposium on Advanced Wound Care Fall 2013, Las Vegas, NV, and at the 28th Annual Clinical Symposium on Advances in Skin and Wound Care, Orlando, FL.