Abbreviations:AD (atopic dermatitis), Th (T helper type)
Atopic dermatitis (AD) is a common, chronic inflammatory skin condition characterized by intense pruritis and recurrent eczematous lesions (
Weidinger and Novak, 2016). An unmet medical need exists for a well-tolerated and effective topical treatment (
Diaz and Guttman-Yassky, 2019).
AD pathogenesis involves a complex interplay between genetic predisposition to skin barrier dysfunction and immune dysregulation. Staphylococcus aureus colonization, occurring in >80% of AD lesions, can further alter the skin barrier and stimulate immune responses (
Li et al., 2019,
Mempel et al., 2003,
Weidinger and Novak, 2016).
SB414, an investigational topical product, is a cream containing berdazimer sodium (CAS Registry Number 1846565-00-1), a macromolecule composed of a polysiloxane backbone with covalently bound N-diazeniumdiolate nitric oxide donors (
Stasko et al., 2018).
Here, we report the effect of SB414 on S. aureus colonization in an FLG-deficient AD murine model (FLGft/ft) and the subsequent cutaneous immune response. A clinical study assessed the mechanism of action of SB414 by analyzing the changes in gene expression of biomarkers indicative of AD-related immune pathways and also evaluated systemic exposure, short-term efficacy, and safety.
The University of California San Diego Institutional Animal Care and Use Committee reviewed and approved the animal protocol (approval number: S09074). In the AD skin model of ovalbumin-sensitized and tape-stripped FLGft/ft Balb/c mice (
Nakatsuji et al., 2016), S. aureus levels in the skin of SB414 6%–treated mice were decreased to 5.3 × 105 colony forming units within 24 hours, representing >90% reduction in S. aureus versus untreated (P = 0.005) and vehicle-treated (P = 0.002) mice (Figure 1a, Supplementary Figure S1). In addition, skin biopsies of S. aureus-colonized lesions revealed that SB414 treatment led to substantial reductions in T helper type (Th) 2–related cytokines, including a 76% reduction in IL-13 gene expression (P = 0.002) and an 87% reduction in IL-4 expression (P = 0.008) versus vehicle treatment (Figure 1b). SB414 treatment also reduced TSLP, IFNγ, and CXCL2, but not IL-17A (data not shown). After confirming modification of the biomarkers in the mouse model, SB414 was advanced to a human clinical trial to prove the mechanism of action.
An institutional review board (Quorum Review Institutional Review Board, Seattle, WA) reviewed and approved all materials for the clinical study, and patients provided written, informed consent before screening procedures were initiated. A total of 48 adults with mild to moderate AD were randomized to SB414 2% (n = 17), SB414 6% (n = 17), and vehicle (n = 14) applied twice daily. Of five patients who discontinued (SB414 2%, n = 1; SB414 6%, n = 2; and vehicle, n = 2), three were due to adverse events (SB414 6%, n = 2; vehicle, n = 1; all three experienced moderate application-site reactions such as pruritus, pain, or swelling). Mean Eczema Area and Severity Index scores (SD) were 8.2 (5.2) for vehicle, 4.7 (2.0) for SB414 2%, and 7.2 (3.3) for SB414 6%. Mean (SD) Eczema Area and Severity Index scores decreased for all treatment groups from baseline to week 2, with the greatest percent reduction of –23.22% (29.14) for SB414 2%. There were no statistical comparisons of efficacy measures.
To understand the mechanism of action of SB414, RNA sequencing and RT-PCR were conducted from patient skin biopsy samples. RNA sequencing showed 612 differentially expressed genes (fold change > 2, false discovery rate < 0.05) between lesional and nonlesional skin at baseline (363 upregulated and 249 downregulated). Following 2 weeks of treatment, the study transcriptome showed 65.5% improvement to more closely resemble nonlesional skin for SB414 2% as compared with a worsening of –20.9% for SB414 6% and a minor improvement of 4.5% for vehicle (P < 0.05) (Figure 2a–c).
Only SB414 2% significantly suppressed the immune signature of AD lesions, whereas exacerbation of the signature for lesions treated with either SB414 6% or vehicle was apparent. Gene set variation analysis, an unsupervised sample-wise enrichment method that produces a score of activity for a gene subset or pathway for each sample (
Hänzelmann et al., 2013), also indicated a statistically significant decrease in a subset of immune genes (
Bissonnette et al., 2019,
Ewald et al., 2015,
Guttman-Yassky et al., 2019,
Pavel et al., 2019), including genes related to innate immunity (IL-6), Th2 pathway (IL-4R and CCL11), Th17/Th22 (IL-36G; S100A7, 8, 9,12; IL-22), Th9 (IL-9), and Th1 pathway (CXCL10 and CXCL11) with SB414 2% at week 2 (data not shown).
Based on gene set variation analysis, SB414 2% was associated with statistically significantly greater suppression of Th17-related gene expression versus both vehicle and baseline (P < 0.01 for both comparisons). Other AD-related pathways showed similar changes using gene set variation analysis, including the meta-analysis–derived AD transcriptome and Th1-, Th2-, and Th22-specific pathways (data not shown).
Changes in expression of key AD-related biomarkers identified by RNA sequencing data were validated using quantitative RT-PCR analysis of 45 immune response genes. Overall, changes in AD-related biomarkers were observed with the SB414 2% treatment after 2 weeks (Supplementary Figure S2). Although all immune axes showed significant modulation with treatment, the most robust changes were observed in Th17/Th22 genes such as S100A8, 9, and 12; STAT3; IL-6; IL-6R; CXCL2 (P < 0.01); and IL-22 (P < 0.05).
Only descriptive pharmacokinetic analyses were performed owing to limited systemic exposure to SB414. No SB414 2% patient had quantifiable concentrations of hydrolyzed N-methylaminopropyl-trimethoxysilane, a backbone of the polymeric drug substance, postdose on day 1 or 15. Only five SB414 6% patients had quantifiable hydrolyzed N-methylaminopropyl-trimethoxysilane concentrations at 1 hour postdose on day 1 or 15 (highest concentration, 34.46 ng/ml; lower limit of quantitation, 5 ng/ml); levels then declined, becoming undetectable in all but two samples (14.3%) by 12 hours postdose.
Adverse events were reported by one SB414 2% patient (headache), two SB414 6% patients (application-site reactions), and three vehicle patients (application-site reactions, accidental product exposure, and AD), with no correlation between adverse events and systemic hydrolyzed N-methylaminopropyl-trimethoxysilane concentrations.
The exacerbation in gene expression of AD-related biomarkers observed following SB414 6% treatment may be due to skin irritation. In patients who reported irritation (SB414 2%, n = 2; SB414 6%, n = 4; vehicle, n = 2), a post hoc analysis revealed that SB414 6% was associated with greater expression of Th1 and innate cytokines, including IL-1β, IFNγ, CXCL9, CXCL10, and CCL3 in lesional skin, compatible with a possible irritant contact dermatitis profile (
Nosbaum et al., 2009). TSLP did not change significantly following treatment with either 2% or 6% SB414 twice daily. In a toxicology study of minipigs, SB414 10% increased skin pH and irritation over time, whereas SB414 0.5% and 2% did not (data not shown), supporting the tolerability concern with SB414 6%.
Although this study is preliminary, evaluating short-term immunomodulatory effects of SB414 on AD lesions, topical nitric oxide–releasing SB414 2% showed significant reductions in Th2-, Th22-, Th1-, and Th17-related biomarker expression and may help treat the various clinical and molecular endotypes of AD (
Czarnowicki et al., 2019;
Renert-Yuval and Guttman-Yassky, 2019).
Data availability statement
Datasets related to this article can be found at https://www.ncbi.nlm.nih.gov/geo/query/acc.cgi?acc=GSE141571, hosted at the National Center for Biotechnology Information’s Gene Expression Omnibus (accession number: GSE141571; secure reviewer's token: qpwzewmadpuvdcx).
Emma Guttman-Yassky: https://orcid.org/0000-0002-9363-324X
Richard L. Gallo: https://orcid.org/0000-0002-1401-7861
Ana B. Pavel: https://orcid.org/0000-0002-0497-8351
Teruaki Nakatsuji: https://orcid.org/0000-0002-6187-2991
Randall Li: https://orcid.org/0000-0002-7504-0723
Ning Zhang: https://orcid.org/0000-0002-9754-1561
Elizabeth Messersmith: https://orcid.org/0000-0003-4689-4006
Tomoko Maeda-Chubachi: https://orcid.org/0000-0002-3614-1505
Conflict of Interest
EGY is an employee of Mount Sinai and has received research funds (grants paid to the institution) from Abbvie, Almirall, Amgen, AnaptysBio, Asana Biosciences, Boerhinger-Ingelhiem, Celgene, Dermavant, DS Biopharma, Eli Lilly, Glenmark, Galderma, Innovaderm, Janssen, Kiniska, Kyowa Kirin, Leo Pharma, Novan, Pfizer, Ralexar, Regeneron, Sienna Biopharma, UCB, and Union Therapeutics. EGY is also a consultant for Abbvie, Almirall, Amgen, Asana Biosciences, Boerhinger-Ingelhiem, Cara Therapeutics, Celgene, Concert, DBV, Dermira, DS Biopharma, Eli Lilly, EMD Serono, Escalier, Galderma, Glenmark, Kyowa Kirin, Leo Pharma, Mitsubishi Tanabe, Pfizer, RAPT Therapeutics, Regeneron, Sanofi, Sienna Biopharma, and Union Therapeutics.
RLG has received research funding from Novan. RLG is a cofounder of and has equity interest in MatriSys.
EM and TMC are employees of and stockholders in Novan, Inc.
Research was sponsored by Novan, Inc, ClinicalTrials.gov Identifier: NCT03431610. The authors are grateful to the patients who gave the gift of clinical research participation. In addition, the authors thank all the principal investigators for their contributions to the study: David M. Pariser, MD, Norfolk, VA; Michael A. Hassman, DO, Berlin, NJ; Mae Gutierrez, MD, Coral Gables, FL; Diane O. McConnehey, DO, CPI, Boise, ID; Phoebe Rich, MD, Portland, OR; Scott A. Fretzin, MD, Indianapolis, IN; Edward L. Lain, MD, MBA, Pflugerville, TX; Ana M. Elosegui, MD, Sweetwater, FL; and Stephen M. Schleicher, MD, Hazleton, PA. The authors also thank Carolyn Enloe of Novan, Inc for operational support of the phase 1b study. Medical writing assistance was provided by Dana L. Randall, MS, PharmD, Intuitive Graphite, Inc and funded by Novan , Inc.
Data included in this manuscript were presented in part as a poster at the International Investigative Dermatology 2018 Meeting (Hollenbach S, Nakatsuji T, Gallo R, Stasko N. Effects of SB414 cream on S. aureus and tissue cytokines in an atopic dermatitis mouse model. May 16–19, 2018; Orlando, FL) and at the 3rd Inflammatory Skin Disease Summit in December 2018 in Vienna, Austria (Maeda-Chubachi T, Durham T, Schleicher S, Rich P, Guttman-Yassky E. Abstract 30. A topical nitric oxide-releasing cream SB414: results of a phases 1b double-blind, randomized, vehicle-controlled study in patients with mild-to-moderate atopic dermatitis. Exp Dermatol. 2018;27(Suppl 2):14).
Novan, Inc was involved in the study design; collection, analysis, and interpretation of data; writing of the report; and the decision to submit the article for publication.
Conceptualization: EGY, RG, EM, TMC; Data Curation: ABP, TN; Formal Analysis: ABP, TN; Funding Acquisition: TMC; Investigation: RL, NZ, TN; Methodology: EGY, RG, TN, TMC; Project Administration: EGY, RG, TMC; Resources: EGY, ABP, TN; Software: ABP, TN; Supervision: EGY, RG, TMC, EM; Validation: ABP, TN; Visualization: ABP, TN; Writing - Original Draft Preparation: ABP; Writing - Review and Editing: EGY, RG, ABP, EM, TMC
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Published online: May 15, 2020
© 2020 The Authors. Published by Elsevier, Inc. on behalf of the Society for Investigative Dermatology.
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