b85a2a84cf1d5b66e6f71afb952e25cc ijms-26-02641.pdf d536d32ee141ce2d6716926874c4cbd1c584e18c ijms-26-02641.pdf 6bf71c5fd4539398736c5b86240ea87b484d677fb680453814094e93865c4ece ijms-26-02641.pdf Title: Topical miRNA Delivery via Elastic Liposomal Formulation: A Promising Genetic Therapy for Cutaneous Lupus Erythematosus (CLE) Subject: Cutaneous lupus erythematosus (CLE) is a chronic autoimmune skin disorder with limited therapeutic options, particularly for refractory discoid lupus (DLE), which often results in scarring and atrophy. Recent studies have identified miR-31, miR-485-3p, and miR-885-5p as key regulators of inflammation, apoptosis, and fibrosis in CLE skin lesions. This research investigates a novel topical miRNA therapy using DDC642 elastic liposomes to target these pathways in CLE. DDC642 liposomes were complexed with miRNAs (anti-miR-31, anti-miR-485-3p, pre-miR-885-5p) and characterized through dynamic light scattering and Cryo-TEM. Cytotoxicity, cellular penetration, and therapeutic efficacy were evaluated in primary keratinocytes, PBMCs, and immune 3D-skin organoids. miRNA lipoplexes were successfully synthesized with optimized particle size, surface charge, and encapsulation efficiency. These lipoplexes exhibited effective cellular penetration and low cytotoxicity. Anti-miR-31 lipoplexes reduced miR-31 and NF-B levels while increasing STK40 and PPP6C expression. Pre-miR-885-5p lipoplexes elevated miR-885-5p levels and downregulated PSMB5 and NF-B in keratinocytes. While anti-miR-485-3p lipoplexes reduced T-cell activation markers. Anti-miR-31 and pre-miR-885-5p lipoplexes successfully modulated inflammatory pathways in 3D-skin CLE models. miRNA lipoplexes represent promising candidates for pioneering topical genetic therapies for CLE. Further studies, including animal models, are necessary to validate and optimize these findings. Keywords: lupus cutaneous; lipid-based nanoparticles; miRNA; topical drug delivery; gene therapy Author: Blanca Joseph-Mullol, Maria Royo, Veronique Preat, Teresa Moliné, Berta Ferrer, Gloria Aparicio, Josefina Cortés-Hernández and Cristina Solé Creator: LaTeX with hyperref Producer: pdfTeX-1.40.25 CreationDate: Mon Mar 17 08:23:03 2025 CET ModDate: Mon Mar 17 09:31:27 2025 CET Custom Metadata: no Metadata Stream: no Tagged: no UserProperties: no Suspects: no Form: none JavaScript: no Pages: 17 Encrypted: no Page size: 595.276 x 841.89 pts (A4) Page rot: 0 File size: 3107311 bytes Optimized: no PDF version: 1.7 name type encoding emb sub uni object ID ------------------------------------ ----------------- ---------------- --- --- --- --------- JDWIZJ+EURM10 Type 1 Builtin yes yes yes 10 0 MIKRSI+VnURWPalladioL Type 1 Custom yes yes yes 15 0 GDXAQS+URWPalladioL-Roma Type 1 Custom yes yes yes 21 0 CRYXFH+URWPalladioL-Bold Type 1 Custom yes yes yes 27 0 QQBTYL+URWPalladioL-Ital Type 1 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yes 219 0 AIIELO+PalatinoLinotype-Roman CID TrueType Identity-H yes yes yes 227 0 AIIENO+PalatinoLinotype-Italic CID TrueType Identity-H yes yes yes 233 0 AIIENP+PalatinoLinotype-Italic TrueType WinAnsi yes yes no 239 0 AIIEPA+PalatinoLinotype-Bold TrueType WinAnsi yes yes no 242 0 AIIEPB+PalatinoLinotype-Roman TrueType WinAnsi yes yes no 245 0 AIIHLC+PalatinoLinotype-Roman TrueType MacRoman yes yes no 248 0 AIIEKN+PalatinoLinotype-Bold CID TrueType Identity-H yes yes yes 261 0 AIIELO+PalatinoLinotype-Roman CID TrueType Identity-H yes yes yes 267 0 AIIENO+PalatinoLinotype-Italic CID TrueType Identity-H yes yes yes 273 0 AIIENP+PalatinoLinotype-Italic TrueType WinAnsi yes yes no 279 0 AIIEPA+PalatinoLinotype-Bold TrueType WinAnsi yes yes no 282 0 AIIEPB+PalatinoLinotype-Roman TrueType WinAnsi yes yes no 285 0 AIIHLC+PalatinoLinotype-Roman TrueType MacRoman yes yes no 288 0 MPUDFL+PazoMath Type 1 Builtin yes yes yes 312 0 Jhove (Rel. 1.28.0, 2023-05-18) Date: 2026-05-30 04:31:58 CEST RepresentationInformation: ijms-26-02641.pdf ReportingModule: PDF-hul, Rel. 1.12.4 (2023-03-16) LastModified: 2026-05-29 16:03:01 CEST Size: 3107311 Format: PDF Version: 1.7 Status: Well-Formed and valid SignatureMatches: PDF-hul MIMEtype: application/pdf PDFMetadata: Objects: 506 FreeObjects: 1 IncrementalUpdates: 0 DocumentCatalog: PageLayout: SinglePage PageMode: UseNone Outlines: Item: Title: Introduction Destination: section.1 Item: Title: Results Destination: section.2 Children: Item: Title: Characterization of DDC642 Liposomes and Their Complexes Destination: subsection.2.1 Item: Title: Cytotoxic Effect and Cell Penetration Destination: subsection.2.2 Item: Title: Transfection Efficiency and Modulation of miRNA-Targeted Pathway Regulation Destination: subsection.2.3 Item: Title: Lipoplex Penetration Capacity and Therapeutic Efficacy in Immune 3D-Skin Organoids Destination: subsection.2.4 Item: Title: Discussion Destination: section.3 Item: Title: Materials and Methods Destination: section.4 Children: Item: Title: Materials and Reagents Destination: subsection.4.1 Item: Title: Particle Preparation and Complex Formation Destination: subsection.4.2 Item: Title: Stability Studies of Lipoplexes Destination: subsection.4.3 Item: Title: Isolation of Peripheral Blood Mononuclear Cells (PBMCs) from CLE Patients Destination: subsection.4.4 Item: Title: Cell Culture and In Vitro Experiments Destination: subsection.4.5 Item: Title: Confocal Live-Cell Microscopy: Interaction Between Liposomes and HEKa or PBMCs Destination: subsection.4.6 Item: Title: Immune 3D-Skin Organoid for Skin Penetration and Therapeutic Efficacy Destination: subsection.4.7 Item: Title: MTT Assay Destination: subsection.4.8 Item: Title: miRNA and RNA Quantification by RT-qPCR Destination: subsection.4.9 Item: Title: Flow Cytometry Destination: subsection.4.10 Item: Title: Immune 3D-Skin Organoid Immunofluorescence Destination: subsection.4.11 Item: Title: Immunofluorescence in Cultured Cells Destination: subsection.4.12 Item: Title: Western Blot Analysis Destination: subsection.4.13 Item: Title: Statistical Analysis Destination: subsection.4.14 Item: Title: References Destination: appendix.A. Info: Title: Topical miRNA Delivery via Elastic Liposomal Formulation: A Promising Genetic Therapy for Cutaneous Lupus Erythematosus (CLE) Author: Blanca Joseph-Mullol, Maria Royo, Veronique Preat, Teresa Moliné, Berta Ferrer, Gloria Aparicio, Josefina Cortés-Hernández and Cristina Solé Subject: Cutaneous lupus erythematosus (CLE) is a chronic autoimmune skin disorder with limited therapeutic options, particularly for refractory discoid lupus (DLE), which often results in scarring and atrophy. Recent studies have identified miR-31, miR-485-3p, and miR-885-5p as key regulators of inflammation, apoptosis, and fibrosis in CLE skin lesions. This research investigates a novel topical miRNA therapy using DDC642 elastic liposomes to target these pathways in CLE. DDC642 liposomes were complexed with miRNAs (anti-miR-31, anti-miR-485-3p, pre-miR-885-5p) and characterized through dynamic light scattering and Cryo-TEM. Cytotoxicity, cellular penetration, and therapeutic efficacy were evaluated in primary keratinocytes, PBMCs, and immune 3D-skin organoids. miRNA lipoplexes were successfully synthesized with optimized particle size, surface charge, and encapsulation efficiency. These lipoplexes exhibited effective cellular penetration and low cytotoxicity. Anti-miR-31 lipoplexes reduced miR-31 and NF-B levels while increasing STK40 and PPP6C expression. Pre-miR-885-5p lipoplexes elevated miR-885-5p levels and downregulated PSMB5 and NF-B in keratinocytes. While anti-miR-485-3p lipoplexes reduced T-cell activation markers. Anti-miR-31 and pre-miR-885-5p lipoplexes successfully modulated inflammatory pathways in 3D-skin CLE models. miRNA lipoplexes represent promising candidates for pioneering topical genetic therapies for CLE. Further studies, including animal models, are necessary to validate and optimize these findings. 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