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Development of a human skin commensal microbe for bacteriotherapy of atopic dermatitis and use in a phase 1 randomized clinical trial

Abstract

Staphylococcus aureus colonizes patients with atopic dermatitis (AD) and exacerbates disease by promoting inflammation. The present study investigated the safety and mechanisms of action of Staphylococcus hominis A9 (ShA9), a bacterium isolated from healthy human skin, as a topical therapy for AD. ShA9 killed S. aureus on the skin of mice and inhibited expression of a toxin from S. aureus (psmα) that promotes inflammation. A first-in-human, phase 1, double-blinded, randomized 1-week trial of topical ShA9 or vehicle on the forearm skin of 54 adults with S. aureus-positive AD (NCT03151148) met its primary endpoint of safety, and participants receiving ShA9 had fewer adverse events associated with AD. Eczema severity was not significantly different when evaluated in all participants treated with ShA9 but a significant decrease in S. aureus and increased ShA9 DNA were seen and met secondary endpoints. Some S. aureus strains on participants were not directly killed by ShA9, but expression of mRNA for psmα was inhibited in all strains. Improvement in local eczema severity was suggested by post-hoc analysis of participants with S. aureus directly killed by ShA9. These observations demonstrate the safety and potential benefits of bacteriotherapy for AD.

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Fig. 1: Preclinical validation of the activity of ShA9 on mice.
Fig. 2: Anti-inflammatory action of ShA9 is mediated by mechanisms independent of antimicrobial activity against S. aureus.
Fig. 3: Efficacy of ShA9 correlates with sensitivity to lantibiotics.
Fig. 4: Survival of ShA9 on AD skin and correlations to autoinducing peptide expression.
Fig. 5: Bacterial species on the skin of participants treated with ShA9.

Data availability

All data were independently validated by Rho Federal Systems Division, Inc. in a blinded manner. The detailed data analysis plan is available in DAIT/Rho Statistical analysis plan v.2.0, dated 15 May 2019, uploaded to Clinicaltrials.gov (https://clinicaltrials.gov/ProvidedDocs/48/NCT03151148/Prot_000.pdf). The 16S rRNA gene sequence data for the present study have been published in open-source web application server, MG-RAST (www.mg-rast.org) with project ID mgp92321. Any materials that can be shared will be released via a material transfer agreement. Source data are provided with this paper.

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Acknowledgements

The present study was funded by the Atopic Dermatitis Research Network (ADRN) (grant nos. U19AI117673 and 1UM1AI151958 to R.L.G. and D.Y.M.L., and UM2AI117870 to Rho Federal Systems Division), and National Institutes of Health grants (nos. R01AR076082, R01AR064781 and R01AI116576 to R.L.G. and T.N., R37AI052453 and R01AI153185 to R.L.G.) and NIH/NCATS Colorado CTSA grant no. UL1 TR002535 to D.Y.M.L.

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Authors

Contributions

R.L.G. conceived the idea for the present study. R.L.G., T.N. and T.R.H. designed the study and wrote the manuscript. A.K.R.S. and D.Y.M.L. reviewed the manuscript. K.J., T.R.H., Y.L.T., J.Y.C., F.S., M.R.G., P.T., D.Y.M.L. and R.L.G. coordinated the clinical trial. G.D. and A.K.R.S. developed the protocol. T.N., A.M.B., S.S.S. and S.B .collected the data from mouse experiments and clinical samples. T.N., T.R.H., A.B. and R.L.G. evaluated and analyzed the data. A.C. and B.J. conducted the statistical analysis. T.R.H., D.Y.M.L. and R.L.G. directed this project as the principal investigators in the ADRN.

Corresponding author

Correspondence to Richard L. Gallo.

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Competing interests

T.N. and R.L.G. are co-inventors of UCSD technology related to the bacterial antimicrobial peptides discussed herein. R.L.G. is co-founder and has equity interest in MatriSys Bioscience and Sente Inc. A.K.R.S.’s co-authorship of this publication does not necessarily constitute endorsement by the NIAID, the NIH or any other agency of the US government. All other authors declare no conflicts of interest.

Additional information

Peer review information Nature Medicine thanks Alice Prince, Katrina Abuabara and the other, anonymous, reviewers for their contribution to the peer review of this work. Alison Farrell was the primary editor on this article and managed its editorial process and peer review in collaboration with the rest of the editorial team.

Publisher’s note Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Extended data

Extended Data Fig. 1

Low levels of bacteria are detected after topical application of bacteria on mice with inflamed skin. Colony forming units (CFU) of S. aureus and CoNS were measured from the spleen of OVA-sensitized FLGft/ft Balb/c mice that were colonized by S. aureus for 4 days and treated by ShA9 for 3 days. Spleens (50–100 mg) were surgically excised and homogenized in 1 mL PBS. Live S. aureus and CoNS CFU were counted on mannitol salt agar with egg yolk and CFU counts adjusted based on wet weight of the excised spleens. Data represent mean ± SEM of biological replicates in individual mouse (n = 8). No statistical difference was detected by two-tailed unpaired parametric t-test.

Source data

Extended Data Fig. 2

Gating strategy for flow cytometry to identify CD4 + , IL4 + or CD4 + , IL17A + cells associated with experiments in Fig. 2g-i.

Extended Data Fig. 3

a-b, Flow diagram of participants (a) and illustration of measurements conducted for the clinical trial (b). § One individual withdrew due to inability to keep the time commitments of the study and was excluded from data analysis according to our study protocol. This individual reported <75% of treatment being applied and was excluded from data analysis according to our study protocol. This individual was lost to follow up due to personal reasons but completed the study. He/she was included to data analysis according to our study protocol.

Extended Data Fig. 4 Change in abundance of S. aureus mprF mRNA on the lesional skin of subjects treated by ShA9.

Each dot represent data from individual subject. Data are shown as mean±95% confidence interval (n = 32 independent subjects). Data at each time point was compared by two-tailed paired parametric t-test and no change was detected.

Source data

Extended Data Fig. 5

a, Minimal inhibitory concentration (MIC) of ShA9 supernatant to inhibit growth of S. aureus isolated from each subject is correlated with change in local EASI at the indicated time points after treatment with ShA9. S. aureus resistant to ShA9 was defined as its MIC > 100% of conditioned media. Statistical analysis for correlation was carried out by two-tailed t distribution (n = 32 independent subjects). Dotted line represents detection limit of the assay. b, Correlation between relative change in live S. aureus abundance and change in local EASI scores from baseline at indicated time points for subjects treated with ShA9 (top row, blue) or vehicle (bottom row, red). Statistical analysis for correlation was carried out by two-tailed t distribution (ShA9: n = 35; Vehicle: n = 17 independent subjects).

Source data

Extended Data Fig. 6

a-c, Abundance of DNA for ShA9 lantibiotic-a (a), S. hominis-specific gap gene (b) and universal 16S rRNA (c) on the lesional and nonlesional skin of patients treated by ShA9 or vehicle at indicated time points. Data represent Mean ± 95% confidence interval (ShA9: n = 35; Vehicle: n = 17 independent subjects) (a-c). A linear mixed-model approach was used to take into account the repeated aspect of the trial (a-c).

Source data

Extended Data Fig. 7

a, Abundance of ShA9 lantibiotic-α mRNA recovered by skin swabs from lesional skin at indicated time points for subjects treated with ShA9 or vehicle. Data represent Mean ± 95% confidence interval (ShA9: n = 35; Vehicle: n = 17 independent subjects). A linear mixed-model approach was used to take into account the repeated aspect of the trial (a). b-c, Correlation between abundance of ShA9 lantibiotic-α mRNA (b) or ShA9 AIP mRNA (c) and S. hominis DNA on the lesional skin of subjects with ShA9 at indicated time points. Statistical analysis for correlation was carried out by t distribution. Dotted line represents detection limit of the assay. Statistical analysis for correlation was carried out by two-tailed t distribution (n = 35 independent subjects) (b,c).

Source data

Extended Data Fig. 8 AIP activity from ShA9 against S. aureus psmα expression is independent from antimicrobial activity of ShA9.

MIC of ShA9 conditioned media (CM) against each patient’s S. aureus was correlated with relative change in S. aureus psmα expression from base line (BL) at indicated time point. CM was precipitated from ShA9 culture supernatant by 70% ammonium sulfate and % calculated from the original volume of medium. Statistical analysis for correlation was carried out by t distribution. Dotted line represents detection limit of the assay. Statistical analysis for correlation was carried out by two-tailed t distribution (n = 35 independent subjects).

Source data

Extended Data Fig. 9

Change in S. aureus psmα expression after ShA9 treatment does not correlate with a decrease in S. aureus survival on AD lesional skin. Statistical analysis for correlation was carried out by two-tailed t distribution (n = 35 independent subjects).

Source data

Extended Data Fig. 10 AIP activity of ShA9 on S. aureus isolates that resistant to ShA9 lantibiotics.

All 11 isolates of S. aureus identified by analyses in Fig. 3d,f,g were cultured in TSB with ShA9 conditioned media (25%) or TSB (Vehicle) at 30 °C for 18 hrs. Abundance in mRNA for Psmα was measured as described in Fig. 2j. Data represent mean of 2 technical replicates in independent bacterial culture.

Source data

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Nakatsuji, T., Hata, T.R., Tong, Y. et al. Development of a human skin commensal microbe for bacteriotherapy of atopic dermatitis and use in a phase 1 randomized clinical trial. Nat Med 27, 700–709 (2021). https://doi.org/10.1038/s41591-021-01256-2

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