Treating severe junctional epidermolysis bullosa with artesunate

1 Treating severe junctional epidermolysis bullosa with artesunate Sophie Marie Sinz1, Ulrich Koller2, Bernadette Liemberger2, Stefan Hainzl2, Alfred Klausegger2, Joerg von Hagen3, Norbert Müller4, Thomas Mohr5, Christopher Gerner5, Gazmend Temaj6, Martin Laimer1, Hannelore Breitenbach-Koller7, Johann W. Bauer1 1 Department of Dermatology and Allergology, University Hospital of the Paracelsus Medical University Salzburg, Muellner Hauptstraße 48, 5020 Salzburg, Austria 2 EB House Austria, Research Program for Molecular Therapy of Genodermatoses, Department of Dermatology and Allergology, University Hospital of the Paracelsus Medical University Salzburg, Muellner Hauptstraße 48, 5020 Salzburg, Austria 3 ryon-Greentech Accelerator, Mainzer Straße 41, 64579 Gernsheim, Germany 4 DepartmentofChemistry,FacultyofScience,UniversityofSouthBohemia,ČeskýchBudějovicích,Branišovská1760,37005ČeskéBudějovice,CzechRepublic 5 Department of Analytical Chemistry, Faculty of Chemistry, University of Vienna, 1090 Vienna, Austria 6 Human Genetics, Faculty of Pharmacy, College UBT, 10000 Pristina, Kosovo 7 Department of Biosciences and Medical Biology, University of Salzburg, 5020 Salzburg, Austria Corresponding author: Johann W. Bauer ([email protected]) Letter to the editor Severe junctional epidermolysis bullosa (sJEB) is a rare genetic, often postpartum lethal epithelial disease that is predominantly caused by biallelic nonsense/premature termination codon (PTC) sequence variants in the LAMB3 gene [1]. LAMB3 gene encodes the β3 chain of the trimeric, hemidesmosomal skin anchor protein laminin 332. PTC mutations prevent the synthesis of full-length functional proteins, and truncated laminin ß3 (lamß3) prevents the formation of laminin 332, with devastating clinical manifestations. These include generalized blistering at birth, periorificial erosions around the mouth, eyes, and nares, significant loss of mucosal surfaces of inner organs, hoarse cry, cough and respiratory difficulties. Sepsis and general failure to thrive result in the demise of patients in early infancy [2]. The condition is currently incurable and would require long-term systemic correction. Artesunate is an FDA-approved, bioavailable small molecule drug for systemic treatment of malaria [3]. It is primarily effective in the blood compartment and can be administered intravenously or orally with minimal side effects. Artesunate is a derivative of artemisinin, a compound originally isolated from the plant Artemisia annua. Its mode of action primarily involves the generation of reactive oxygen species (ROS) within the malaria parasite Plasmodium. It is targeting the parasite’s haemoglobin digestion process inside red blood cells. The drug is activated by the iron present in the heme (from broken-down hemoglobin) and forms free radicals which damage the parasite’s proteins and lipids, leading to its death [4]. Targeted ribosome editing in yeast and human cells identified artesunate as an efficient enhancer of lamß3 protein translation [5–8]. Here, application of artesunate to both sJEB-LAMB3 HaCat model cells and to immortalized sJEBLAMB3 patient keratinocytes revealed a significant increase of lamß3 protein after 4 days of treatment with 1 µM artesunate/0.01% DMSO in CnT Prime medium (Fig. 1). Thus, we describe the use of artesunate in a patient with sJEB, homozygous for a LAMB3 PTC mutation. A girl of 3 months of age was Subject editor: Georg Stingl Received: 23 August 2025 Accepted: 3 October 2025 Published: 2 December 2025 Citation: Sinz SM, Koller U, Liemberger B, Hainzl S, Klausegger A, Hagen Jvon, Müller N, Mohr T, Gerner C, Temaj G, Laimer M, Breitenbach-Koller H, Bauer JW (2025) Treating severe junctional epidermolysis bullosa with artesunate. SKINdeep 1: e169584. https://doi. org/10.1553/skindeep.2025.169584 SKINdeep 1: e169584 (2025) DOI: 10.1553/skindeep.2025.169584 Copyright: © Sophie Marie Sinz et al. This is an open access article distributed under terms of the Creative Commons Attribution License (Attribution 4.0 International – CC BY-NC 4.0). 2 SKINdeep 1: e169584 (2025), DOI: 10.1553/skindeep.2025.169584 Sophie Marie Sinz et al.: Treating severe junctional epidermolysis bullosa with artesunate presented to the clinic with the phenotype of sJEB. In addition to widespread chronic wounds, she had growth retardation, anemia, and repeating episodes of infections (i.e., respiratory infections, bacterial wound infections, candida infection in oral mucous membranes). Since birth, her wounds were treated with various wound dressings, that were adjusted to the local condition. In general, for this indication, no approved treatment is available. Therefore, the parents agreed to start the off-label treatment with artesunate. Artesunate was administered initially in calendar week (cw) 39 2023 intravenously with a total amount of 3 mg q.d., then switched to an oral application with 10 mg daily (1 ml of 10 mg/ml oral suspension) and increased to 20 mg after 3 months and to 40 mg after 6 months (Table 1). Artesunate was well tolerated; side effects resulting from anemic episodes most likely due to iron Figure 1. Immunofluorescence (IF) analysis of keratinocytes stained with α-laminin ß3 antibody. HaCat model cells comprise samples PARENTAL CONTROL (original HaCat keratinocytes), HETERO CONTROL and HOMO CONTROL, where homozygous and heterozygous deficiencies in LAMB3 expression were generated using CRISPR/Cas9 [7]. HOMO ART, a homozygote-deficient LAMB3 R635X cell culture was treated with artesunate 1 uM for 4 days. Samples of E6/E7-immortalized keratinocytes were derived from cells isolated from an unaffected proband, HKC CONTROL, from a LAMB3 R635X heterozygous carrier, HKC HET CONTROL, and from the homozygous patient, JEB-CONTROL, and from cells isolated from the patient treated with artesunate 1uM for 4 days, JEB-ART (p-value JEB-control vs. JEB-ART: 0,053). Scale bar: 100 um. 3 SKINdeep 1: e169584 (2025), DOI: 10.1553/skindeep.2025.169584 Sophie Marie Sinz et al.: Treating severe junctional epidermolysis bullosa with artesunate deficiency and chronic disease were met with discontinuing treatment. Clinical response showed a mixed picture: During the first 5 months of treatment, no effect on wound healing (nates and flanks) was observed according to the treating physician’s assessment. An immunofluorescence (IF) investigation of patient skin was performed after these 5 months of treatment and showed only a marginal increase in expression of lamß3 protein (Fig. 2). However, respiratory complications improved with the start of oral administration (cw 41) and returned when oral treatment was paused during infectious episodes. From December 2023 to January 2024, artesunate was also administered topically (10 mg/ml) q.d. to the affected areas on the flanks with no signs of re-epithelialization (not shown). After 6 months, oral dosing was increased to 40 mg daily. After one week of treatment at this dosage, the patient’s wounds on the nates began to show gradual healing (Fig. 3). However, the patient´s general condition worsened, and she died in cw 19 2024. Table 1. Timing and dose of artesunate at 10 mg/ml preparation (cw: calendar week). calendar week dosage artesunate complications cw 39 2023 300ul iv cw 41 2023 1 ml orally cw 51 2023 2 ml orally, 4 ml topically cw 3 2024 break, restart afterwards with 2ml orally episode of oral Candida cw 8 2024 break, restart with 2ml orally fever, anaemia cw 12 2024 4 ml orally, amnion-transplant Figure 2. Immunofluorescence investigation of patient skin (biopsies taken from the right upper thigh): Residual expression of laminin ß3 after a total of 5 months of treatment, of which the last 2 months were dosed with 20 mg orally daily (left); compared to laminin a3 expression of the patient skin (right), see white arrows. Scale bar: 100 um. IF images were split into green and blue channels. In the blue channel, DAPI-stained nuclei were counted using an Otsu binarization algorithm. The green channel was background corrected for non-specific lamß3 staining using a rolling ball algorithm, followed by integration of the pixel intensities. Values were normalized by dividing the integrated intensity by the number of nuclei counted (normalized integrated intensity). Normalized integrated fluorescence intensities were divided by the mean normalized fluorescence intensities of the overall controls (parental control for the three left columns, HKC-control for the three right columns). Representative green and blue channel images are shown for each group. 4 SKINdeep 1: e169584 (2025), DOI: 10.1553/skindeep.2025.169584 Sophie Marie Sinz et al.: Treating severe junctional epidermolysis bullosa with artesunate Discussion Biochemical and in vitro evidence suggests that the specific binding of artesunate to ribosomal LP35 leads to stop codon readthrough in the premature termination stop codons of the LAMB3 gene. Here, a patient with s-JEB-LAMB3 was treated with intravenous, oral and topical artesunate to determine safety in neonates. Safety was acceptable, and in areas where artesunate had local access (oral cavity, perianal), positive clinical effects were reported by treating physicians and visually documented (Fig. 3). Other systemic treatments for s-JEB include aminoglycosides, such as gentamicin [9], and non-aminoglycosides, such as ataluren [10], to promote general ribosomal readthrough. Gentamicin is not available orally and treatment options are limited to pulsed, short-term intravenous applications due to nephrotoxic and ototoxic effects. Ataluren is available orally, but there are no safety studies on long-term use in EB of this compound. Both compounds are compromised by their lack of specificity, i.e. they are general PTC readthrough drugs that are unable to distinguish between PTC signals resulting from mutations and endogenous PTC signals that regulate the production of protein isoforms. Possible reasons why the clinical outcome in this patient could not be changed include: 1) Bioavailability of artesunate in the skin as opposed to the blood system. 2) The initiation and dosage of treatment, which probably preclude the timely accumulation of laminin ß3 protein in epithelial tissues. 3) Effects of prenatal loss of laminin ß3 [11] leading to immune dysregulation. Future studies that treat patients as early as possible to reduce the formation of large wounds and promote the formation of intact oral and intestinal mucosa, as well as strategies to develop artesunate derivatives with improved bioavailability, are warranted. Acknowledgements We thank the diagnostic laboratory of the Department of Dermatology and Allergology for performing the IF. We thank Dr.s Diem, Ude-Schoder, Welponer for performing clinical care of the patient. The support of the patient´s family is gratefully acknowledged. Figure 3. Wound healing during treatment with 40 mg artesunate orally daily for 1 week starting in cw 13 2024 (right) compared to cw 9 2024 at 20 mg orally daily (left). 5 SKINdeep 1: e169584 (2025), DOI: 10.1553/skindeep.2025.169584 Sophie Marie Sinz et al.: Treating severe junctional epidermolysis bullosa with artesunate Additional information Conflict of interest Dr. Bauer and Dr. Breitenbach-Koller disclose funding from Land Salzburg WISS 2025 Research Initiative Grant HIRIBO (PFL181001_03) and thank Maestro Mandozzi Locarno/ Salzburg for continuous support of our work to develop ribosome editing for sJEB. They and Dr. von Hagen are shareholders of KBHB GmbH, a company developing ribosome editing technology to provide targeted therapy for rare and prevalent diseases. DEBRA Austria supported U.K., B.L., A.K., S.H. No other disclosures are reported. Ethical statements The authors declared that no clinical trials were used in the present study. The authors declared that experiments on humans or human tissues were performed for the present study. For the generation of cell lines, tissue samples from healthy donors and the JEB patient were obtained upon written informed consent at the Department of Dermatology and Allergology, University Hospital Salzburg. Ethical approval was granted by the ethics committee of the county of Salzburg (vote number: 415-E/2118/45-2024). All procedures described in this study were in full accordance with Austrian legislation and with the Declaration of Helsinki. The authors declared that no experiments on animals were performed for the present study. Use of commercially available immortalised human and animal cell lines: HaCaT, Cell Lines Service GmbH, 300493 https://www.selectscience.net/product/hacat. Use of AI No use of AI was reported. Funding No funding was reported. Author contributions All authors have contributed equally. We thank the patient’s parents for granting permission to publish this information. Author ORCIDs Sophie Marie Sinz https://orcid.org/0009-0001-8381-9147 Ulrich Koller https://orcid.org/0000-0002-6285-1789 Bernadette Liemberger https://orcid.org/0000-0002-1056-362X Alfred Klausegger https://orcid.org/0000-0001-6160-9934 Joerg von Hagen https://orcid.org/0000-0001-9810-3590 Norbert Müller https://orcid.org/0000-0002-7621-3980 Thomas Mohr https://orcid.org/0000-0002-1933-847X Christopher Gerner https://orcid.org/0000-0003-4964-0642 Gazmend Temaj https://orcid.org/0000-0003-4807-2938 Hannelore Breitenbach-Koller https://orcid.org/0000-0002-8387-6408 Johann W. Bauer https://orcid.org/0000-0002-6085-9170 6 SKINdeep 1: e169584 (2025), DOI: 10.1553/skindeep.2025.169584 Sophie Marie Sinz et al.: Treating severe junctional epidermolysis bullosa with artesunate Data availability All of the data that support the findings of this study are available in the main text. References 1. Has C, Bauer JW, Bodemer C, Bolling MC, Bruckner-Tuderman L, Diem A et al. Consensus reclassification of inherited epidermolysis bullosa and other disorders with skin fragility. Br J Dermatol. 2020;183:614–27. https://doi.org/10.1111/bjd.18921 2. Laimer M, Lanschuetzer CM, Diem A, Bauer JW. Herlitz junctional epidermolysis bullosa. Dermatol Clin. 2010;28:55–60. https://doi.org/10.1016/j.det.2009.10.006 3. U.S. centers for disease control and prevention. Treatment of severe malaria [Internet]. 07.03.2024 [last access 12.12.2024] at https://www.cdc.gov/malaria/ hcp/clinical-guidance/treatment-of-severe-malaria.html 4. Ruwizhi N, Maseko RB, Aderibigbe BA. Recent advances in the therapeutic efficacy of artesunate. Pharmaceutics. 2022 Feb 25;14(3):504. https://doi.org/10.3390/ pharmaceutics14030504 5. Bauer JW, Brandl C, Haubenreisser O, Wimmer B, Weber M, Karl T et al. Specialized yeast ribosomes: a customized tool for selective mRNA translation. PLoS One. 2013 Jul 8;8(7):e67609. https://doi.org/10.1371/journal.pone.0067609 [Epub 08.07.2013] 6. Rathner A, Rathner P, Friedrich A, Wießner M, Kitzler CM, Schernthaner J et al. Drug development for target ribosomal protein rpL35/uL29 for repair of LAMB3R635X. rare skin disease epidermolysis bullosa. Skin Pharmacol Physiol. 2021;34:167– 82. https://doi.org/10.1159/000513260 7. Moßhammer C. Ribosomal protein rpL35/uL29 as target for systemic repair of LAMB3R635XPTC mutation in epidermolysis bullosa: Studies in yeast and human PTC/PTC model system [Ph.D. Thesis]. Salzburg: Department of Biosciences and Medical Biology, University of Salzburg. 2021 8. Wimmer B, Friedrich A, Poeltner K, Edobor G, Mosshammer C, Temaj G, et al. En route to targeted ribosome editing to replenish skin anchor protein LAMB3 in junctional epidermolysis bullosa. JID Innov. 2023 Nov 10;4(1):100240. https://doi. org/10.1016/j.xjidi.2023.100240 [Epub January 2024] 9. Woodley T, Hao M, Kwon A, Levian B, Cogan J, Hou Y, et al. Intravenous gentamicin therapy induces functional type VII collagen in patients with recessive dystrophic epidermolysis bullosa: an open-label clinical trial. Br J Dermatol. 2024;191:267– 74. https://doi.org/10.1093/bjd/ljae063 10. Orlowski G, Amano S, Flanagan. K, Rieger K, Marinkovich M, Wiss. K. Treatment with ataluren for wound healing and health complications in a patient with junctional epidermolysis bullosa. JAMA Dermatol 2023;159:1145–7. https://doi. org/10.1001/jamadermatol.2023.2077 11. Nyström A, Bornert O, Kühl T, Gretzmeier C, Thriene K, Dengjel J et al. Impaired lymphoid extracellular matrix impedes antibacterial immunity in epidermolysis bullosa. Proc Natl Acad Sci U S A. 2018;115:E705–E714. https://doi.org/10.1073/ pnas.1709111115 [Epub 05.01.2018]