Effect of a Novel Macrophage-Regulating Drug on Wound Healing in Patients With Diabetic Foot Ulcers: A Randomized Clinical Trial | Dermatology | JAMA Network Open | JAMA Network
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Visual Abstract. RCT: Effect of a novel macrophage-regulating drug on wound healing in patients with diabetic foot ulcers
RCT: Effect of a novel macrophage-regulating drug on wound healing in patients with diabetic foot ulcers
Figure 1.  CONSORT Diagram of Study Flow
CONSORT Diagram of Study Flow

A total of 236 patients were randomized. Absorbent dressing was Hydrofiber (ConvaTec Ltd). To convert glycated hemoglobin (HbA1c) to proportion of total hemoglobin, multiply by 0.01. ABI indicates ankle-brachial index; FAS, full-analysis set; and mITT, modified intention to treat.

aJudged by the investigator to be unsuitable for the study for any other reason.

Figure 2.  Kaplan-Meier Plot of Time to Complete Healing in the Full-Analysis Set Population
Kaplan-Meier Plot of Time to Complete Healing in the Full-Analysis Set Population

The survival curve indicates the incidence of ulcers healed at each visit in the full-analysis set population. Complete healing was defined as epithelialization without drainage observed at 2 consecutive visits. A full-analysis set cohort randomly assigned to the absorbent dressing (Hydrofiber; ConvaTec Ltd) group (n = 114) or ON101 group (n = 122) was used for Kaplan-Meier analysis.

Table 1.  Baseline Patient Characteristics and Intervention During the Study
Baseline Patient Characteristics and Intervention During the Study
Table 2.  Primary and Secondary Outcomesa
Primary and Secondary Outcomesa
1.
Edmonds  M.  A renaissance in diabetic foot care: new evidence-based treatments.   Lancet Diabetes Endocrinol. 2018;6(11):837-838. doi:10.1016/S2213-8587(18)30262-6 PubMedGoogle ScholarCrossref
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Hingorani  A, LaMuraglia  GM, Henke  P,  et al.  The management of diabetic foot: a clinical practice guideline by the Society for Vascular Surgery in collaboration with the American Podiatric Medical Association and the Society for Vascular Medicine.   J Vasc Surg. 2016;63(2)(suppl):3S-21S. doi:10.1016/j.jvs.2015.10.003 PubMedGoogle ScholarCrossref
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Everett  E, Mathioudakis  N.  Update on management of diabetic foot ulcers.   Ann N Y Acad Sci. 2018;1411(1):153-165. doi:10.1111/nyas.13569 PubMedGoogle ScholarCrossref
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Jeffcoate  WJ, Vileikyte  L, Boyko  EJ, Armstrong  DG, Boulton  AJM.  Current challenges and opportunities in the prevention and management of diabetic foot ulcers.   Diabetes Care. 2018;41(4):645-652. doi:10.2337/dc17-1836 PubMedGoogle ScholarCrossref
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Bus  SA, Van Netten  JJ, Hinchliffe  RJ, Apelqvist  J, Lipsky  BA, Schaper  NC; IWGDF Editorial Board.  Standards for the development and methodology of the 2019 International Working Group on the Diabetic Foot guidelines.   Diabetes Metab Res Rev. 2020;36(suppl 1):e3267. doi:10.1002/dmrr.3267 PubMedGoogle Scholar
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Geiss  LS, Li  Y, Hora  I, Albright  A, Rolka  D, Gregg  EW.  Resurgence of diabetes-related nontraumatic lower-extremity amputation in the young and middle-aged adult US population.   Diabetes Care. 2019;42(1):50-54. doi:10.2337/dc18-1380 PubMedGoogle ScholarCrossref
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Baltzis  D, Eleftheriadou  I, Veves  A.  Pathogenesis and treatment of impaired wound healing in diabetes mellitus: new insights.   Adv Ther. 2014;31(8):817-836. doi:10.1007/s12325-014-0140-x PubMedGoogle ScholarCrossref
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Jalilian  M, Ahmadi Sarbarzeh  P, Oubari  S.  Factors related to severity of diabetic foot ulcer: a systematic review.   Diabetes Metab Syndr Obes. 2020;13:1835-1842. doi:10.2147/DMSO.S256243 PubMedGoogle ScholarCrossref
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Kuo  YS, Chien  HF, Lu  W.  Plectranthus amboinicus and Centella asiatica cream for the treatment of diabetic foot ulcers.   Evid Based Complement Alternat Med. 2012;2012:418679. doi:10.1155/2012/418679 PubMedGoogle Scholar
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Arumugam  G, Swamy  MK, Sinniah  UR.  Plectranthus amboinicus (Lour) Spreng: botanical, phytochemical, pharmacological and nutritional significance.   Molecules. 2016;21(4):369. doi:10.3390/molecules21040369 PubMedGoogle ScholarCrossref
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Nagoor Meeran  MF, Goyal  SN, Suchal  K, Sharma  C, Patil  CR, Ojha  SK.  Pharmacological properties, molecular mechanisms, and pharmaceutical development of Asiatic acid: a pentacyclic triterpenoid of therapeutic promise.   Front Pharmacol. 2018;9:892. doi:10.3389/fphar.2018.00892 PubMedGoogle ScholarCrossref
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Leu  WJ, Chen  JC, Guh  JH.  Extract from Plectranthus amboinicus inhibit maturation and release of interleukin 1β through inhibition of NF-κB nuclear translocation and NLRP3 inflammasome activation.   Front Pharmacol. 2019;10:573. doi:10.3389/fphar.2019.00573 PubMedGoogle ScholarCrossref
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Sawatdee  S, Choochuay  K, Chanthorn  W, Srichana  T.  Evaluation of the topical spray containing Centella asiatica extract and its efficacy on excision wounds in rats.   Acta Pharm. 2016;66(2):233-244. doi:10.1515/acph-2016-0018 PubMedGoogle ScholarCrossref
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Bylka  W, Znajdek-Awiżeń  P, Studzińska-Sroka  E, Dańczak-Pazdrowska  A, Brzezińska  M.  Centella asiatica in dermatology: an overview.   Phytother Res. 2014;28(8):1117-1124. doi:10.1002/ptr.5110 PubMedGoogle ScholarCrossref
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Maderal  AD, Vivas  AC, Eaglstein  WH, Kirsner  RS.  The FDA and designing clinical trials for chronic cutaneous ulcers.   Semin Cell Dev Biol. 2012;23(9):993-999. doi:10.1016/j.semcdb.2012.09.014 PubMedGoogle ScholarCrossref
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International Council for Harmonisation of Technical Requirements for Pharmaceuticals for Human Use. ICH guidelines. Accessed August 11, 2021. https://www.ich.org/page/ich-guidelines
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International Working Group on the Diabetic Foot. IWGDF guidelines. Accessed August 11, 2021. https://iwgdfguidelines.org/
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Strotman  PK, Reif  TJ, Pinzur  MS.  Charcot arthropathy of the foot and ankle.   Foot Ankle Int. 2016;37(11):1255-1263. doi:10.1177/1071100716674434 PubMedGoogle ScholarCrossref
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Musa  HG, Ahmed  ME.  Associated risk factors and management of chronic diabetic foot ulcers exceeding 6 months’ duration.   Diabet Foot Ankle. 2012;3:3. doi:10.3402/dfa.v3io.18980PubMedGoogle Scholar
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Khunkaew  S, Fernandez  R, Sim  J.  Health-related quality of life among adults living with diabetic foot ulcers: a meta-analysis.   Qual Life Res. 2019;28(6):1413-1427. doi:10.1007/s11136-018-2082-2 PubMedGoogle ScholarCrossref
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DeMets  DL, Lan  KK.  Interim analysis: the alpha spending function approach.   Stat Med. 1994;13(13-14):1341-1352. doi:10.1002/sim.4780131308PubMedGoogle ScholarCrossref
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Monteiro-Soares  M, Boyko  EJ, Ribeiro  J, Ribeiro  I, Dinis-Ribeiro  M.  Predictive factors for diabetic foot ulceration: a systematic review.   Diabetes Metab Res Rev. 2012;28(7):574-600. doi:10.1002/dmrr.2319 PubMedGoogle ScholarCrossref
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Moganti  K, Li  F, Schmuttermaier  C,  et al.  Hyperglycemia induces mixed M1/M2 cytokine profile in primary human monocyte-derived macrophages.   Immunobiology. 2017;222(10):952-959. doi:10.1016/j.imbio.2016.07.006 PubMedGoogle ScholarCrossref
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Mirza  RE, Fang  MM, Weinheimer-Haus  EM, Ennis  WJ, Koh  TJ.  Sustained inflammasome activity in macrophages impairs wound healing in type 2 diabetic humans and mice.   Diabetes. 2014;63(3):1103-1114. doi:10.2337/db13-0927 PubMedGoogle ScholarCrossref
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Jeffcoate  WJ, Price  PE, Phillips  CJ,  et al.  Randomised controlled trial of the use of three dressing preparations in the management of chronic ulceration of the foot in diabetes.   Health Technol Assess. 2009;13(54):1-86. doi:10.3310/hta13540 PubMedGoogle ScholarCrossref
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    Original Investigation
    Diabetes and Endocrinology
    September 3, 2021

    Effect of a Novel Macrophage-Regulating Drug on Wound Healing in Patients With Diabetic Foot Ulcers: A Randomized Clinical Trial

    Author Affiliations
    • 1Division of Endocrinology and Metabolism, Department of Internal Medicine, Chang Gung Memorial Hospital, Taoyuan City, Taiwan
    • 2Department of Medical Nutritional Therapy, Chang Gung Memorial Hospital, Taoyuan City, Taiwan
    • 3Oneness Biotech Company Limited, Taipei, Taiwan
    • 4Division of Plastic Surgery, Department of Surgery, National Taiwan University Hospital and College of Medicine, Taipei, Taiwan
    • 5Limb Preservation Platform Inc, Fresno, California
    • 6Plastic and Reconstruction Surgery, China Medical University Hospital, Taichung, Taiwan
    • 7Plastic Surgery Center, Chi-Mei Medical Center, Tainan, Taiwan
    • 8Department of Surgery, Mackay Memorial Hospital, Taipei, Taiwan
    • 9Division of Cardiology, Buddhist Tzu Chi General Hospital, Taipei, Taiwan
    • 10Department of Plastic and Reconstructive Surgery, Chang Gung Memorial Hospital, Kaohsiung, Taiwan
    • 11Division of Plastic and Reconstructive Surgery, Taipei Veterans General Hospital, Taipei, Taiwan
    • 12Department of Surgery, National Yang Ming Chiao Tung University School of Medicine, Taipei, Taiwan
    • 13Department of Endocrinology, The First Affiliated Hospital of Soochow University, Jiangsu, China
    • 14Department of Burn and Plastic Surgery, Affiliated Hospital of Jiangsu University, Jiangsu, China
    • 15Department of Endocrinology, The First Affiliated Hospital of Henan Science and Technology University, Henan, China
    • 16Department of Vascular Anomalies, Shanghai TCM (Traditional Chinese Medicine)–Integrated Hospital, Shanghai, China
    • 17Department of Endocrinology, The First Affiliated Hospital of Sun Yat-sen University, Guangdong, China
    • 18Department of Endocrinology, Nanfang Hospital of Southern Medical University, Guangdong, China
    • 19Department of Endocrinology, Sun Yat-sen Memorial Hospital of Sun Yat-sen University, Guangdong, China
    • 20Department of Endocrinology, Shandong Provincial Hospital Affiliated to Shandong First Medical University, Shandong, China
    • 21Shanghai National Clinical Research Center for Endocrine and Metabolic Diseases, Shanghai, China
    • 22Key Laboratory for Endocrine and Metabolic Diseases of the National Health Commission of the People’s Republic of China, Shanghai, China
    • 23Shanghai Institute of Endocrine and Metabolic Diseases, Ruijin Hospital, Shanghai, China
    • 24Shanghai Jiao Tong University School of Medicine, Shanghai, China
    • 25Division of Plastic Surgery, Department of Surgery, Integrated Burn and Wound Care Center, Taipei Medical University-Shuang Ho Hospital, Ministry of Health and Welfare, New Taipei City, Taiwan
    • 26Division of Plastic Surgery, Department of Surgery, School of Medicine, College of Medicine, Taipei Medical University, Taipei, Taiwan
    JAMA Netw Open. 2021;4(9):e2122607. doi:10.1001/jamanetworkopen.2021.22607
    Key Points

    Question  Can the topical application of ON101 cream demonstrate a superior therapeutic benefit in wound healing among patients with diabetic foot ulcers (DFUs) compared with standard care?

    Findings  In this randomized phase 3 clinical trial of 236 patients with DFUs, topical application of ON101 with gauze immediately after debridement demonstrated significant healing efficacy compared with an absorbent dressing in all patients, including those with DFU-related risk factors.

    Meaning  Topical treatment with ON101 resulted in improved healing of DFUs.

    Abstract

    Importance  Delayed healing of diabetic foot ulcers (DFUs) is known to be caused by dysregulated M1/M2-type macrophages, and restoring the balance between these macrophage types plays a critical role in healing. However, drugs used to regulate M1/M2 macrophages have not yet been studied in large randomized clinical trials.

    Objective  To compare the topical application of ON101 cream with use of an absorbent dressing (Hydrofiber; ConvaTec Ltd) when treating DFUs.

    Design, Setting, and Participants  This multicenter, evaluator-blinded, phase 3 randomized clinical trial was performed in 21 clinical and medical centers across the US, China, and Taiwan from November 23, 2012, to May 11, 2020. Eligible patients with debrided DFUs of 1 to 25 cm2 present for at least 4 weeks and with Wagner grade 1 or 2 were randomized 1:1 to receive ON101 or control absorbent dressings.

    Interventions  Twice-daily applications of ON101 or a absorbent dressing changed once daily or 2 to 3 times a week for 16 weeks, with a 12-week follow-up.

    Main Outcomes and Measures  The primary outcome was the incidence of complete healing, defined as complete re-epithelialization at 2 consecutive visits during the treatment period assessed on the full-analysis set (FAS) of all participants with postrandomization data collected. Safety outcomes included assessment of the incidences of adverse events, clinical laboratory values, and vital signs.

    Results  In the FAS, 236 eligible patients (175 men [74.2%]; mean [SD] age, 57.0 [10.9] years; mean [SD] glycated hemoglobin level, 8.1% [1.6%]) with DFUs classified as Wagner grade 1 or 2 (mean [SD] ulcer area, 4.8 [4.4] cm2) were randomized to receive either the ON101 cream (n = 122) or the absorbent dressing (n = 114) for as long as 16 weeks. The incidence of complete healing in the FAS included 74 patients (60.7%) in the ON101 group and 40 (35.1%) in the comparator group during the 16-week treatment period (difference, 25.6 percentage points; odds ratio, 2.84; 95% CI, 1.66-4.84; P < .001). A total of 7 (5.7%) treatment-emergent adverse events occurred in the ON101 group vs 5 (4.4%) in the comparator group. No treatment-related serious adverse events occurred in the ON101 group vs 1 (0.9%) in the comparator group.

    Conclusions and Relevance  In this multicenter randomized clinical trial, ON101 exhibited better healing efficacy than absorbent dressing alone in the treatment of DFUs and showed consistent efficacy among all patients, including those with DFU-related risk factors (glycated hemoglobin level, ≥9%; ulcer area, >5 cm2; and DFU duration, ≥6 months).

    Trial Registration  ClinicalTrials.gov Identifier: NCT01898923

    Introduction

    Approximately 80% of lower limb amputations are preceded by chronic diabetic foot ulcers (DFUs), resulting in a heavy burden of medical care and expenditure.1,2 The current treatment for DFUs in clinical practice focuses primarily on local wound care, including debridement, off-loading, infection control, and maintaining a moist environment with dressings,3,4 whereas adjunctive therapies such as the use of growth factors, tissue engineering products, hyperbaric oxygen, and negative pressure wound therapies are applied if the DFUs worsen.5 Although current treatments featuring tissue repair or the use of anti-inflammatory agents might help in closing or controlling the progression of DFUs, most of these treatments are not well supported by clinical evidence or are not recommended for routine care by the International Working Group on the Diabetic Foot.6 In addition, the annual increase in amputations also suggests that treatment improvement is needed.7 Diabetic foot ulcers are pathologically complex mostly because the ulceration is undermined by the existence of multiple risk factors, such as poor patient adherence to treatment, severity of the ulcer, ulcer location and duration, vascular condition, control of glycated hemoglobin (HbA1c) levels, smoking habits, and kidney dysfunction.8,9 These factors impose a significant clinical need for novel and effective interventions to tackle this life-debilitating and life-threatening disease.

    Accumulating scientific evidence has revealed that targeting macrophage phenotypes might be a potentially effective therapy in DFUs because hyperglycemia increases the ratio of proinflammatory M1 to proregenerative M2 macrophages.9-14 ON101 (supplied by Oneness Biotech Co, Ltd; previously given the research code WH-1) exerts its therapeutic effect through regulation of the balance between M1 and M2 macrophages. ON101 is composed of 2 active pharmaceutical ingredients: PA-F4 from an extract of Plectranthus amboinicus and S1 from an extract of Centella asiatica, 2 medicinal plants reported to have significant pharmacological activities in wound healing.10-12 With 48 in vitro and in vivo studies performed, these 2 ingredients, which contribute to a synergistic effect on regulation of the M1:M2 macrophage ratio, have been defined and formulated in a cream base using a proprietary formula. One of these ingredients, PA-F4, significantly attenuates M1 macrophages by suppressing the NLRP3-mediated inflammasome pathway and the production of downstream inflammatory cytokines such as interleukin 1β and interleukin 6,13 which arrest the inflammation phase. On the other hand, the extract of C asiatica has been reported to activate M2 macrophages by increasing collagen synthesis and by stimulating fibroblast proliferation and the migration of keratinocytes.14,15 ON101 has been further demonstrated to accelerate wound healing efficiently in a db/db mouse model of diabetes, obesity, and dyslipidemia by decreasing inflammatory M1 macrophage activity and enriching M2 macrophage populations through granulocyte colony-stimulating factor–mediated M2 polarization, which changed the ulcer status from the inflammatory phase to the proliferation and remodeling stages (eFigure 1 in Supplement 2).

    A clinical pharmacokinetic study on 12 patients with DFUs showed that topical administration of ON101 twice daily in single and multiple doses yielded very limited systemic exposure (Kai-Min Chu, MD, PhD, oral communication, September 4, 2017). Thus, the maximum body concentrations from days 1 and 14 were similar, demonstrating that topical ON101 has no obvious accumulation in the body. No treatment-related adverse events were observed. In a clinical research trial conducted in 24 patients with chronic DFUs classified as grade 3 according to the Wagner system,10 treatment with ON101 for 2 weeks resulted in an approximately 20% reduction in wound size, and no serious adverse events were reported. Of the 21 patients with evaluable data, the mean wound size at baseline was 359 (range, 20-2352) mm2, decreasing to 293 mm2 after 2 weeks of ON101 treatment.10 Another clinical trial was performed with 30 patients with Wagner grade 1 chronic DFUs treated with ON101 for as long as 12 weeks (Yu-Yao Huang, MD, PhD, oral communication, August 22, 2011). The final incidence of healing was 50%. The mean wound area at baseline was 577 (range, 303-1225) mm2, decreasing to 163 mm2 after 12 weeks of ON101 treatment.

    The topical use of ON101 is supported with a safety profile from the manufacturer and has clear therapeutic potential in promoting wound healing based on previous studies.10 This multicenter, phase 3 randomized clinical trial was designed to evaluate whether ON101 could treat chronic DFUs by comparing it with a standard primary wound care absorbent dressing.

    Methods

    We followed adequate and well-controlled studies as categorized by the US Food and Drug Administration16 to design a randomized, controlled, evaluator-blinded phase 3 trial to evaluate the efficacy of ON101 applied topically twice daily for treating chronic DFUs (the trial protocol is available in Supplement 1). This treatment was compared with an absorbent dressing (Hydrofiber; ConvaTec Ltd) as a comparator in the control group for treating chronic DFUs. This multicenter study was performed with institutional review board approval from 21 medical/clinical centers (eTable 1 in Supplement 2) with wound care specialty across the US, China, and Taiwan, where these investigational new drug programs were initiated; all patients provided written informed consent at enrollment. The study followed the International Council on Harmonization guideline17 and the Consolidated Standards of Reporting Trials (CONSORT) reporting guideline.

    From November 23, 2012, to May 11, 2020, we enrolled outpatients with type 1 or 2 diabetes (as defined by World Health Organization criteria) aged 20 to 80 years, with a baseline HbA1c level of less than 12% measured during screening or within 3 months before randomization (to convert to proportion of total hemoglobin, multiply by 0.01). The target ulcer classified as grade 1 or 2 based on the Wagner system on the foot (below the ankle) needed to measure from 1 to 25 cm2 after debridement, without active infection, and present for at least 4 weeks despite receiving standard of care (according to the International Working Group on the Diabetic Foot guidelines18) before randomization. To avoid possible premature discontinuation of the patient treatments during the trial, we excluded patients with an ankle-brachial before randomization; those with necrosis, purulence, or sinus tracts in the target ulcer not removable by debridement during the screening visit; or those with acute Charcot neuroarthropathy as defined by the American Diabetes Association and the American Podiatric Medical Association, which indicates perturbations of bone metabolism.19 In addition, revascularization procedures aimed at increasing blood flow in the target limb must have been performed at least 4 weeks before randomization.

    Eligible participants judged by the principal investigators (Y.-Y.H., N.-C.C., H.-H.C., K.-F.H., K.-Y.T., H.-L.H., P.-Y.L., C.-K.P., B.S., C.L., Y.M., Y.C., Y.L., Y.X., Q.L., G.N., and S.-C.C.) on completion of the screening period (≤7 days) were assigned to receive ON101 or absorbent dressing for as long as 16 weeks in a 1:1 allocation by a computer-generated block randomization scheme (eMethods 1 in Supplement 2).20 Individual investigators and research staff were blinded to the size of the block and remained blinded to the treatment assignment before randomization, eliminating the possibility of predetermining the prospective participant’s treatment assignment. The investigator was informed of the randomized treatment assignment in a sealed envelope containing the individual treatment code at the baseline visit.

    The end-of-treatment visit (visit 10) was the visit in the 16th week after randomization or the visit in which complete wound closure was confirmed, whichever happened first. The independent evaluator assessed the degree of wound closure. The independent evaluator and the study statistician were blinded to the participants’ treatment throughout the study until the clinical database had been locked. To ensure masking throughout the trial, a standardized procedure was established including camera settings, photographing and image-encoding, image delivery to the independent evaluator, and outcome assessment based on the digitally encoded images to delink the patients’ identification, treatment groups, visits, or site information. The detailed blinding procedure is described in eMethods 2 in Supplement 2.

    Interventions

    Demographic data, medical history, disease status, radiography, and eligibility were evaluated during the screening period (before randomization). Participants were scheduled for return visits every 2 weeks to receive wound cleansing and debridement with an assessment of wound status, wound size measurement, physical examination results, and concomitant medication records throughout the 16 weeks of the study period once the interventions were administered. The principal investigators and nurses were trained to use standardized study materials, ON101 or absorbent dressings, camera setting, and off-loading recommendations. The instruction for use of off-loading devices was given to the patients with plantar ulcers as assessed by the clinical investigators. All adverse events were recorded at every visit once the intervention was applied. Blood samples for laboratory tests (including hematologic and biochemical analysis) were collected at the screening visit, then every 4 weeks during the treatment period and at the last visit of the follow-up period to detect the levels of factors such as alanine aminotransferase and aspartate aminotransferase to measure liver status, creatinine and blood urea nitrogen to measure kidney status, and albumin to measure nutritional status. Levels of HbA1c and blood glucose were measured to monitor diabetes-related safety concerns.

    ON101, a topical cream composed of PA-F4 and S1, was supplied by Oneness Biotech Co, Ltd, and manufactured in Taiwan in a facility in compliance with Good Manufacturing Practice certified by the Pharmaceutical Inspection Cooperation Scheme. Participants in the ON101 treatment group were shown how to self-administer the cream twice daily in an amount to cover the target ulcer fully without exceeding 2 mm in thickness at each visit. The absorbent dressing containing sodium carboxymethylcellulose (Aquacel; ConvaTec Ltd) needed to be changed daily or 2 to 3 times weekly subject to exudate level following the product’s instructions or the investigators’ discretion. The only secondary dressing allowed was sterile gauze for both groups. The amount of ON101 used or the frequency of absorbent dressing changes for each patient was recorded at every visit during the treatment period. No systemic prescriptions were contraindicated during the treatment period, whereas topical antimicrobials and antiseptic agents were not allowed.

    In cases where the target ulcer worsened (defined as Wagner grade 3), the investigators could determine whether to terminate treatment. If the ulcer was judged by the blinded evaluator as having undergone complete epithelialization for 2 consecutive visits during the treatment period (at or before visit 10), the intervention (ON101 or absorbent dressing) was stopped, and a visit 10 was scheduled after this judgment. If the patients were confirmed to have an unhealed target ulcer at visit 10, continual standard of care with the absorbent dressings was provided to them regardless of the allocated group during the 12-week follow-up period.

    Data Collection and Outcome Measures

    The primary efficacy outcome was to compare the incidence of complete healing between the 2 groups at the end of the 16-week treatment period. Complete healing, defined as complete epithelialization maintained without drainage or requirement of dressings for at least 2 consecutive visits, was determined by an independent evaluator blinded to the patient’s information and treatment allocation. Secondary ulcer-related outcomes included time to complete ulcer healing (from baseline visit to first 100% re-epithelialization visit), percentage of change in ulcer surface area from baseline (to the latest treatment visit or complete wound closure), percentage of patients with a 50% reduction in ulcer surface area, and incidence of infection of the target ulcer. The exploratory, ulcer-related outcome data included any incidence of ulcer recurrence during the 12-week follow-up period. Target wound size was measured by an investigator using digital planimetry at every visit after any necessary debridement. In addition, efficacy variables were further assessed for subgroups for the incidence of complete healing, characterized according to the prior duration of ulcers recorded at the baseline visit (6 months as a cutoff),21 ulcer size (5 cm2 as a cutoff),22 and HbA1c level (9% as a cutoff regarded as poor glycemic control according to the definition of the American Diabetes Association). Safety outcomes were used to assess adverse events and clinical laboratory values.

    Statistical Analysis

    The sample size was calculated based on the results of ON101 in the previous trial by hypothesizing a 20% superiority in the incidence of wound closure compared with the efficacy of the absorbent dressing (Yu-Yao Huang, MD, PhD, oral communication, August 22, 2011). With a 1:1 randomization ratio in the 2 groups, 236 participants were required to be enrolled to ensure that at least 212 had evaluable data for achieving 80% power with a 2-sided α value of 5% nominal significance. All analyses were performed using SAS software, version 9.4 (SAS Institute Inc). The intention-to-treat (ITT) principle was applied to the full-analysis set (FAS), which included all randomized patients irrespective of the actual receipt of study intervention and adherence to the protocol or the occurrence of adverse events. The FAS was used to analyze all efficacy and safety data. A modified ITT (mITT) protocol was applied to exclude patients in the FAS with ineligible target ulcers at baseline. The mITT was used for supportive analysis of efficacy data as appropriate.

    For the primary end point, we used a χ2 test and a logistic regression model with intervention as a fixed factor, with the baseline ulcer size and Wagner grade adjusted as covariates. The results of the logistic regression model are presented in terms of the odds ratio (OR), with P values and associated 95% CIs. Some outcomes are expressed as the hazard ratio (HR). Exploratory post hoc analyses of pertinent variables, such as ulcer duration, ulcer size, and patients’ HbA1c levels, were also performed. The time to complete ulcer healing was calculated using the Kaplan-Meier method with a log-rank test. The HRs and 95% CIs were estimated using a Cox proportional hazards regression model. The percentile changes in ulcer surface area and ulcer surface area change from baseline were subjected to regression analysis adjusted by baseline ulcer area and Wagner grades. The incidences of infection of target ulcers and of recurrence were evaluated using the Fisher exact test.

    The adverse events were regarded as treatment emergent if they occurred after the intervention started. Adverse events, treatment-emergent adverse events, and serious adverse events were summarized by frequency and proportion of total patients by system organ class and by preferred terms. All adverse event–related comparisons between the 2 groups were performed using the Fisher exact test. The clinical laboratory test data were used to tabulate the change in values from baseline and were compared between groups using analysis of covariance. All tests were 2 tailed, and P < .05 was considered statistically significant.

    For possible early study termination, an independent data monitoring committee was established to monitor data when the patient numbers reached approximately 50% and 90% of the planned enrollment. The futility or superiority of ON101 cream was assessed by the independent data monitoring committee using the Lan-DeMets alpha-spending approach, in which the boundaries were determined by the type of O’Brien-Fleming spending function.23 The superiority of ON101 was confirmed by the independent data monitoring committee (P < .001, much less than the boundary of 0.03476) on achieving 90% of the planned enrollment (212 participants with evaluable data) so the interim analysis could proceed. The trial was not terminated despite ON101 achieving superiority in the interim analysis because the 236th patient with evaluable data was already enrolled before this point.

    Results

    A total of 236 patients were included in the FAS (175 men [74.2%]; 61 women [25.8%]; mean [SD] age, 57.0 [10.9] years). The mean (SD) HbA1c level was 8.1% (1.6%) at baseline and did not change significantly at the end of treatment (mean [SD] HbA1c of 8.0% [1.8%] in the ON101 group vs 7.9% [1.6%] in the comparator group), and 144 patients (61.0%) were diagnosed as having had diabetes for more than 10 years. Patients in the FAS were randomly allocated to treatment: 114 (48.3%) to the comparator group and 122 (51.7%) to the ON101 group. Sixteen patients (13.1%) in the ON101 group vs 21 (18.4%) in the comparator group had an early termination (total of 37) (Figure 1). The instructions for using off-loading devices were given to the patients who were assessed by the clinical investigators. Some patients did not follow the suggestion because of the humidity in Taiwan (Table 1). Among the 236 patients in the FAS, 184 (78.0%) were classified as having Wagner grade 2 ulcers, 117 (49.6%) had ulcers in the plantar region, and 64 (27.1%) had a baseline HbA1c level of at least 9%. The mean (SD) ulcer size was 4.8 (4.4) cm2, and the mean (SD) prior duration of the target ulcer was 7.2 (13.4) months at entry (Table 1).

    Primary Outcome

    Seventy-four patients (60.7%) in the ON101 group vs 40 (35.1%) in the comparator group achieved ulcer closure within 16 weeks (OR, 2.84; 95% CI, 1.66-4.84; P < .001) (Table 2). Similar results were also noted in the mITT population, where 73 of 118 patients (61.9%) in the ON101 group and 38 of 112 (33.9%) in the comparator group had ulcer closure (OR, 3.15; 95% CI, 1.82-5.43; P < .001) (Table 2 and eTable 2 in Supplement 2). The independent evaluator assessed the degree of wound closure.

    Ulcer duration, ulcer size, and HbA1c levels are known to be associated with poor prognosis of DFUs.9,24,25 Therefore, a subgroup analysis was conducted on baseline ulcer duration (6 months as a cutoff), baseline ulcer area (5 cm2 as a cutoff size), and baseline HbA1c level (9% as a cutoff). The subgroup analysis displayed a significant OR in favor of the ON101 group compared with the comparator group (OR, 3.14 [95% CI, 1.04-9.50; P = .04] for HbA1c level ≥9%; OR, 3.99 [95% CI, 1.09-14.63; P = .04] for ulcer duration ≥6 months; OR, 4.09 [95% CI, 1.42-11.80; P = .009] for ulcer size >5 cm2) (Table 2). In addition, we subgrouped patients with an ulcer reduction of less than 10% during the screening period and analyzed the primary efficacy variable. The result also favored the ON101 treatment (32 of 64 [50.0%] vs 18 of 66 [27.3%]; P = .02) (eTable 6 in Supplement 2).

    Secondary Outcome

    Patients in the ON101 group had a better healing rate than those in the comparator group (HR, 1.80 [95% CI, 1.23-2.65; P = .002]) (Figure 2) in the FAS as well as the mITT population (HR, 1.91 [95% CI, 1.29-2.83; P = .001]) (eFigure 2 in Supplement 2). The cumulative incidence of complete healing at each week also reflected the continual higher probability in the ON101 group for reaching complete wound closure from week 4 onward. The time to reach median population healing was 98 days in the ON101 group, whereas it was undeterminable in the comparator group because ulcers of only 40 patients (35.1%) healed in this group during the treatment period (Figure 2). The mean reduction in ulcer surface area (from the last treatment visit to baseline) was 78.0% in both groups (SDs, 42.6% for the ON101 group and 34.9% for the comparator group; P = .89), and the incidence of a 50% reduction in ulcer surface area was not significantly different between both groups (101 of 122 [82.8%] vs 98 of 114 [86.0%]) (Table 2). Only a few episodes of target ulcer infection occurred in both groups during the treatment period (6 in the ON101 group and 7 in the comparator group; P = .78) (Table 2). The incidence of recurrence in completely healed wounds during the follow-up phase was 15 of 74 (20.3%) in the ON101 group and 7 of 40 (17.5%) in the comparator group without statistical significance (P = .81) (Table 2).

    Adverse Events

    In terms of safety, there were no clinically significant changes or differences between the 2 treatment groups in hematology, biochemistry (including HbA1c and fasting glucose levels), or vital signs (Table 2 and eTable 3 in Supplement 2). Treatment-emergent adverse events were reported in 76 patients in the ON101 group and 77 in the comparator group, of whom 7 of 122 (5.7%) in the ON101 group and 5 of 114 (4.4%) from the comparator group were considered related to the treatments (Table 2 and eTable 4 in Supplement 2). None of the serious adverse events was related to ON101 treatment, whereas there was 1 case of osteomyelitis reported to be linked to the comparator group in which 1 patient (0.8%) assigned to ON101 died of septic shock, acute kidney injury, and acute respiratory failure, which were not considered to be related to treatment or to ulcer progression (eTable 5 in Supplement 2).

    Discussion

    To our knowledge, this study is the first international phase 3 randomized clinical trial of an investigational drug able to regulate M1/M2 macrophage activities in patients with DFUs. ON101 exhibited better efficacy in facilitating the complete healing of DFUs. Hyperglycemia is an underlying cause of chronic DFUs in which the M1-to-M2 macrophage transition is delayed and the inflammatory stage is prolonged.26,27 ON101 can restore the balance of M1/M2 macrophages caused by hyperglycemia. The robust efficacy in patients with high-risk factors suggests that ON101 might provide multiple and proactive ways to improve wound healing by promoting the M1-to-M2 transition and thereby accelerating wound healing for ulcers not only in terms of early formation but also with high-risk factors including ulcer duration of at least 6 months, ulcer size greater than 5 cm2, and an HbA1c level of at least 9%.

    The design of this study followed US Food and Drug Administration guidelines.16 The complete healing rate of the comparator group at 16 weeks (35.1%) was consistent with the 28.2% shown by ITT analysis at week 12 disclosed in a previous trial by Jeffcoate et al.28 This finding verifies the suitability of the design and implementation of this study in conforming to other randomized clinical trials. The application of ON101 after debridement—which can be self-administered at home—indicated the same level of convenience of use as for the absorbent dressing.

    Despite the statistically significant wound closure and healing rates provided by ON101, the ulcer reduction outcomes, including changes in ulcer area from baseline and rate of 50% reduction in the wound area, were not statistically significant between the 2 groups during the treatment period. This discrepancy possibly arose from the use of 2-dimensional measurements on the wound area without considering the wound depth. In this study, 78.0% of the ulcers were Wagner grade 2, meaning that they extended into tendon, bone, or capsule. Thus, the measurement of wound area instead of volume might not reflect the actual volumetric change. Similar outcomes were also noted in the pivotal study (study 92-22120-K) of becaplermin (Regranex; Smith & Nephew plc). The use of 3-dimensional measurement tools should be considered in future studies.

    Limitations

    This study has some limitations. The first was the open-label design, which did not allow us to mask the interventions to patients or clinical investigators; therefore, blinded evaluation was implemented to minimize any possible bias. Second, the inclusion and exclusion criteria ruled out patients requiring dialysis, which, to a certain extent, reflects some types of patients with DFUs. Using the ankle-brachial index as the sole criterion in judging blood perfusion could not exclude patients with ischemia completely. Last, the lack of a 2-week run-in period was a potential flaw in the design, because possible rapid healers might not have been excluded in the study. To assess whether this factor affected the trial results, a separate analysis of the complete ulcer healing rate was performed by excluding those patients with an ulcer reduction of at least 10% during the screening period, the results of which favored ON101 treatment (32 of 64 [50.0%] vs 18 of 66 [27.3%]; P = .02) (eTable 6 in Supplement 2).

    Conclusions

    The results of this randomized clinical trial demonstrate a clinically and statistically superior therapeutic efficacy of ON101 in the treatment of DFUs in both FAS and mITT populations in terms of complete healing rate and time to complete healing compared with absorbent dressing. For chronic wounds in patients with high-risk factors, the therapeutic efficacy of ON101 could be sustained in ulcers that last for more than 6 months or measure greater than 5 cm2 or in patients with high HbA1c levels. The findings of this study suggest that ON101, a macrophage regulator that behaves differently from moisture-retaining dressings, represents an active-healing alternative for home and primary care of patients with chronic DFUs.

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    Article Information

    Accepted for Publication: June 15, 2021.

    Published: September 3, 2021. doi:10.1001/jamanetworkopen.2021.22607

    Open Access: This is an open access article distributed under the terms of the CC-BY-NC-ND License. © 2021 Huang YY et al. JAMA Network Open.

    Corresponding Author: Shun-Cheng Chang, MD, Division of Plastic Surgery, Department of Surgery, Integrated Burn and Wound Care Center, Taipei Medical University-Shuang Ho Hospital, Ministry of Health and Welfare, No. 291 Zhongzheng Rd, Zhonghe Dist, New Taipei City 235, Taiwan (csc901515@gmail.com).

    Author Contributions: Drs Y.-Y. Huang and Chang had full access to all the data in the study and take responsibility for the integrity of the data and the accuracy of the data analysis.

    Concept and design: Chen, H.-L. Huang, P.-Y. Lin, Chang.

    Acquisition, analysis, or interpretation of data: Y.-Y. Huang, C.-W. Lin, Cheng, Cazzell, K.-F. Huang, Tung, H.-L. Huang, Perng, Shi, Liu, Ma, Cao, Y. Li, Xue, Yan, Q. Li, Ning.

    Drafting of the manuscript: C.-W. Lin, Chen, P.-Y. Lin, Perng, Liu.

    Critical revision of the manuscript for important intellectual content: Y.-Y. Huang, Cheng, Cazzell, K.-F. Huang, Tung, H.-L. Huang, Shi, Ma, Cao, Y. Li, Xue, Yan, Q. Li, Ning, Chang.

    Statistical analysis: C.-W. Lin.

    Administrative, technical, or material support: Y.-Y. Huang, Cheng, Chen, K.-F. Huang, Tung, P.-Y. Lin, Shi, Liu, Ma, Cao, Y. Li, Xue, Yan, Q. Li, Ning, Chang.

    Supervision: Chang.

    Conflict of Interest Disclosures: Dr C.-W. Lin reported receiving personal fees from Oneness Biotech Co, Ltd, during the conduct of the study. Dr Chang reported receiving a speakers fee from Oneness Biotech Co, Ltd. No other disclosures were reported.

    Funding/Support: This study was supported by Oneness Biotech Co, Ltd, Microbio Co, Ltd, and Shanghai Haihe Pharmaceutical Co, Ltd.

    Role of the Funder/Sponsor: The sponsors were responsible for designing the study and preparing the manuscript. The sponsors had no role in the conduct of the study; collection, management, analysis, and interpretation of the data; review or approval of the manuscript; and decision to submit the manuscript for publication.

    Data Sharing Statement: See Supplement 3.

    Additional Contributions: We thank all the staff and participants at the sites involved in the study. Pan-Chyr Yang, MD, PhD (Genomic Research Center, Academia Sinica), Tsen-Wen Chen, MD, PhD (Department of Internal Medicine, Wei Gong Memorial Hospital), and Wayne Huey-Herng Sheu, MD, PhD (Division of Endocrinology and Metabolism, Department of Internal Medicine, Taichung Veterans General Hospital), acted as medical consultants, for which they were not compensated. Ya-Yuan Lu, MS (Bestat Pharmaservices Corporation), was responsible for statistical analysis and data interpretation. The sponsors paid Bestat Pharmaservices Corporation for their services. OnLine English editors assisted with revision, for which they were compensated.

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