A Retrospective Crossover Study of the Use of Aseptically Processed Placental Membrane in the Treatment of Chronic Diabetic Foot Ulcers

Background. In a recently published, prospective randomized controlled trial (RCT) comparing aseptically processed dehydrated human amnion/chorion allograft (dHACA) to standard of care (SOC), 85% wound closure rates were reported in the dHACA arm while only 25% of patients in the SOC arm healed. Objective. The purpose of this retrospective crossover study is to evaluate the effectiveness of dHACA in those patients that failed to respond to the SOC treatments and who exited the original study after failing up to 12 weeks of SOC treatment. Materials and Methods. Patients with nonhealing wounds from the SOC arm after exit from the original study were offered weekly adjunctive applications of dHACA (AmnioBand; Musculoskeletal Transplant Foundation, Edison, NJ) for up to 12 weeks. The primary endpoint was the proportion of wounds completely healed at 12 weeks. Secondary endpoints included the difference in wound area from baseline to the end of study and the percentage area reduction (PAR). Results. Eleven patients were eligible to participate; wounds for 9 of the 11 patients healed (82%). The mean wound area decreased from 1.7 cm² to 0.2 cm² (P = .0005), with a corresponding mean PAR of 92%. Of the 2 wounds that failed to heal, 1 diabetic foot ulcer (DFU) decreased in area by 91% and the other by 26%. Conclusion. The results of this crossover study support the conclusions of the original RCT, which determined that aseptically processed dHACA is an effective means to treat recalcitrant DFUs. Further studies, including comparative clinical trials, may offer additional information on this unique aseptically processed graft in the healing of indolent wounds.

A recently published, prospective randomized clinical trial (RCT)¹ showed that an aseptically processed dehydrated human amnion/chorion allograft (dHACA) (AmnioBand; Musculoskeletal Transplant Foundation [MTF], Edison, NJ) significantly increased the healing response in diabetic foot ulcers (DFUs). In this study of 40 patients, 85% (17/20) of patients healed with dHACA treatments, whereas only 25% (5/20) healed with standard of care (SOC), which included weekly debridement, daily application of a calcium alginate follow by a 3-layer dressing, and an offloading camboot. 

The purpose of the present study is to crossover eligible nonhealing patients who received SOC and track whether introduction and weekly applications of dHACA could facilitate complete healing of the wound. 

Patients from the original RCT¹ were eligible to participate in this crossover study if 1 of 2 circumstances occurred: (1) they were in the SOC arm and withdrew from the original trial, per protocol, at 6 weeks because the area of their DFU did not decrease by at least 50%; or (2) if their wound had not healed by 12 weeks.

Of the 15 patients who failed to heal in the SOC arm, 11 eligible patients participated in the crossover study and returned to the 5 outpatient wound care centers across the United States that served as the study sites. Four patients were excluded: 3 had prior adverse events (namely infections) that prevented enrollment and 1 chose not to participate. All patient information and photographs were kept in the patient’s source file or regular medical file. Once all patients who failed out of SOC in the prospective study completed their dHACA treatment in the crossover, the Western Institutional Review Board retrospectively approved the additional study protocol on June 20, 2016 (#20161369). Patients then returned to their clinic and provided written informed consent to participate in this crossover study analysis. This study adhered to the Declaration of Helsinki and Good Clinical Practice regulatory requirements.

During the crossover period, wounds were cleansed, debrided, and digitally photographed at each visit and treated in an identical manner as the original experimental group of patients in the RCT.¹ Wounds were assessed for infection based on the guidelines of Woo and Sibbald.² Patients were offered to receive the gifted graft immediately upon exit from the prospective study, and therefore, their choice to enter the crossover occurred immediately following original prospective study exit.

The aseptically processed dHACA used in this study was donated by MTF (Edison, NJ; a 501c3 nonprofit corporation) and was sized from 1-cm disks to sheets of 4 cm x 6 cm. An appropriately sized graft was chosen for each application based on the wound surface area. The graft was cut to size with a number 15 blade and pie-crusted, as needed, to a ratio < 1:5:1.0. The graft was rinsed with sterile saline and applied over the entire wound surface. A nonadherent dressing (ADAPTIC TOUCH; Acelity, San Antonio, TX) was the primary backing, which was covered with a moisture-retentive dressing (hydrogel bolster) and a padded 3-layer dressing (DYNA-FLEX; Acelity). A removable controlled ankle movement (CAM) walker (Royce Medical, Camarillo, CA) was used to offload the wound, and a total contact cast (TCC) was used when the removable CAM walker would not fit appropriately. Only 1 patient in the entire cohort required a TCC due to an ill-fitting CAM boot. Patients were seen once weekly and received a dHACA application at each visit until complete wound closure occurred or after 12 weeks of treatment. At the conclusion of the study, patients were provided with diabetic shoes and insoles at no cost.

The primary endpoint of this study was the proportion of wounds completely healed at 12 weeks. Secondary endpoints included the difference in wound area from baseline to end-of-study values and percentage wound area reduction (PAR). The primary endpoint was calculated using the Wilcoxon signed-rank test; the secondary endpoint PAR was calculated as PAR = ([A_I – A_12w]/A_I)*100, where A_Iwas wound area at study entry and A_12w was the wound area at 12 weeks.

Missing data would be handled using the last observation carried forward. Continuous variables were summarized as means, medians, and standard deviations (SDs). Categorical variables were summarized as a number and percentage. Statistical analysis was performed using SPSS Statistics, Version 19 (IBM Corp, Armonk, NY).

This study is registered under Central IRB Registration: WIRB PRO NUM: 20161369.

The original RCT¹ commenced on May 5, 2015, and concluded on January 19, 2016. Of the 11 patients eligible for the crossover study, 8 had exited from the RCT at 6 weeks because their wounds had not reduced in area by 50% when treated with SOC. The remaining 3 patients exited after 12 weeks of unsuccessful SOC treatment.

At study entry, patient characteristics (N = 11) were similar to the patients as a whole in the SOC arm of the RCT¹ (N = 20; Table). Wounds were much smaller in area at study entry (1.7 cm²) than the SOC group at the beginning of the RCT (3.3 cm²) due to some reduction in wound area using SOC during the RCT. 

By the end of the 12-week crossover period, 9 of 11 wounds closed (82%; Figure 1), and the mean area of the DFUs decreased from 1.7 cm² to 0.2 cm² (P = .0005). Two patients failed to heal by 12 weeks; however, 1 decreased in area by 91% and the other by 26%. At the end of the study period, the mean PAR was 92% (SD: 21; Figure 2). There were no adverse events or serious adverse events experienced during the crossover study. The significant improvement seen with the application of dHACA is further illustrated with 2 cases. 

Case 1
A 68-year-old man with a history of a plantar medial midfoot ulcer (5.4 cm²) of 28 weeks’ duration (hemoglobin A_1c [HbA_1c] 7.3; creatinine 1.0 mg/dL) entered the SOC arm of the original trial (Figure 3A). After 12 weeks of SOC treatment, he exited the study with a HbA_1c of 6.8 and agreed to join crossover (Figure 3B). He received 1 application of dHACA per week for 2 weeks, and by week 3 of the crossover study the wound had healed (Figure 3C). 

Case 2
A 67-year-old man with a 50-pack-year history and a plantar forefoot ulcer (1.95 cm²) of 12 weeks duration (HbA_1c 9.8; creatinine 2.0 mg/dL) entered the SOC arm of the original trial (Figure 4A). After 6 weeks of SOC treatment, the wound failed to heal by 50%; the patient exited per study protocol with a HbA_1c of 10.5 and agreed to join the crossover (Figure 4B). He received 1 dHACA graft per week for 5 weeks and healed completely by week 6 (Figure 4C). 

Aseptically processed dHACA does not undergo terminal sterilization and is regulated as a human cell, tissue, and cellular- and tissue-based product under the US Food and Drug Administration Code of Federal Regulations Title 21 guidelines and Section 361 of the Public Health Service Act based on homologous use and minimal manipulation of human tissue. The aseptic processing helps maintain the structural integrity of the amnion and chorion layers.  

The efficacy of this graft has been detailed by the results of an RCT¹ that compared the application of aseptically processed dHACA with SOC alone on 40 DFUs. Wounds treated with dHACA had superior healing rates at 12 weeks (85%) compared with SOC alone (25%) as well as faster healing rates with a mean time to heal in dHACA-treated wounds of 36 days versus 70 days in SOC-treated wounds.¹ 

The SOC arm in the previous study¹ left many patients unhealed who would have regretfully had to seek outside treatment; however, gifted dHACA graft was made available for this group and they were invited to the crossover study to receive weekly application of dHACA for up to 12 weeks.

Eleven patients received dHACA as an adjunct treatment to SOC in this retrospective crossover study. Although the crossover group was smaller than the dHACA arm (20 patients) in the RCT,¹ the proportion of healed wounds was similar in both cohorts: (82% and 85%, respectively). This further validates the high healing rate and efficacy of dHACA over SOC alone.  

The mean initial area of the participating crossover group (1.7 cm²) was less than the original SOC group of the RCT (3.3 cm²), which would be expected as some of the wounds did gradually improve with 25% of patients healing completely in the RCT.¹ This statistic notably aligns with previously reported healing rates for DFU treatments of SOC.³

Over 12 weeks, PAR was also tracked in the original RCT. The extracted PAR data of the 11 patients who failed to heal with SOC and were eligible for this crossover study reveal a compelling difference in PAR progression when their mean PAR during SOC is compared with their mean PAR during dHACA treatment (Figure 2). In particular, the PAR of this group during the original RCT¹ remained stagnant over the course of 12 weeks, even after an initial increase between weeks 1 and 2; the group displayed radical improvement in PAR over the 12-week period of the present study of weekly dHACA treatment. In fact, after only 2 weeks of treatment in the crossover, this indolent subset of wounds had already reduced in area by greater than 60%. This finding also validates the efficacy of dHACA over SOC alone.

An advantage of using dHACA over other comparable grafts is avoiding terminal sterilization methodologies in order to reduce the potential of both macro- and microstructural alterations as well as biochemical component damage in dHACA.^4-9 

Another benefit of dHACA is its availability as a disk as small as 1 cm in diameter, the smallest size offered in the marketplace, to more precisely match the size of the graft to the size of the wound. As the processing of these materials is relatively expensive, reducing graft waste is a simple way to reduce cost. In fact, when discussing cost to closure using bioengineered tissue or allogenic grafts, some of the lowest cost to closure that has ever been reported was with dHACA at $1400 per healed wound in the original RCT¹ and a comparable RCT¹⁰ looking at a unique human reticular allogenic dermal matrix (Allopatch; MTF) with a cost to closure of $1475. 

The main limitations of this study were its retrospective nature, small sample size, and the fact that patients were not required to follow-up since they were seen as regular wound patients in the clinic and were under no obligation to return and receive the complimentary graft. In addition, a comprehensive economic analysis was also beyond the scope of this study.

In conclusion, this crossover study demonstrated high healing rates of 82% among patients with DFUs who were treated with dHACA after failing to improve with SOC alone during the original RCT.¹ The results further support the conclusion that dHACA is a beneficial modality for treating the indolent DFU. Further comparative investigations and studies of more complex wounds will further clarify which patients will most benefit from this technology. 

Affiliations: Lower Extremity Institute for Research and Therapy (LEIRT), Youngstown, OH; Division of Plastic Surgery, Brigham and Women’s Hospital, Boston, MA; Division of Plastic Surgery, Feinberg School of Medicine, Northwestern University, Chicago, IL; SerenaGroup, Cambridge, MA; Strategic Solutions, Inc, Cody, WY; General Surgery, Premier Surgical, Brick, NJ; Martinsville Research Institute, Martinsville, VA; and the Professional Education and Research Institute, Roanoke, VA

Correspondence:
Charles M. Zelen, DPM
Professional Education and Research Institute
222 Walnut Avenue
Roanoke, VA 24016
cmzelen@periedu.com

Disclosure: Dr. DiDomenico is Medical Director of LEIRT and received research funds from the Musculoskeletal Transplant Foundation (MTF; Edison, NJ) to conduct this study. Dr. Orgill is a consultant for MTF and receives research funding through grants to Brigham and Women’s Hospital. Dr. Galiano is a consultant for MTF and receives research funding through grants to Northwestern University School of Medicine. Dr. Serena is CEO and Medical Director of SerenaGroup and received research funds from MTF to conduct this study. Dr. Carter has received funds as a consultant for MTF. Dr. Kaufman is a consultant for MTF and has received funds for speaking engagements. Dr. Zelen is Medical Director and CEO of the Professional Education and Research Institute and has received funds from MTF to conduct clinical trials. This study was funded by MTF.