Background/Purpose: Long Interspersed Element-1 (LINE-1) are retrotransposable DNA elements that make up ~17% of the human genome, and their role in health and disease is still being evaluated. Published studies have suggested that LINE-1 may contribute to the development and progression of autoimmune diseases such as lupus by triggering the type I interferon (IFN) production via activation of the innate immune system through nucleic acid sensing pathways. Additionally, studies have shown that lupus patients have higher level of LINE-1 in kidneys and blood compared to healthy controls, but expression of LINE-1 in SLE skin, where type I IFN signatures are dominant, has not been studied. We thus examined LINE-1 RNA and protein expression in lupus patient skin biopsies, and the impact on inflammatory signals upon a LINE-1 reverse transcriptase (RT) inhibitor in cellular and murine models.

Methods: All human subjects gave informed, written consent for the study. Formalin-fixed, paraffin embedded skin sections from non-lesional, non-sun exposed and UVB-treated skin were examined for changes in interferon-stimulated gene (ISG) expression via RNA-seq, LINE-1 ORF1p and phospho-STING via immunohistochemistry, and for LINE-1 ORF1 and ORF2 transcripts by RNA-scope. A novel, potent LINE-1 RT inhibitor, RPT-A, was characterized using LINE-1 RT enzyme assay, cellular LINE-1 retrotransposition assay, a cellular model of Aicardi-Goutières syndrome, and a UV-irradiated keratinocyte model. To investigate its impact on type I interferon pathway in lupus settings, we assessed its activity in regulating ISGs in UV-irradiated keratinocytes from lupus patients and healthy controls by RNA-seq. We also studied the impact of RPT-A on disease in a TREX1 knockout interferonopathy mouse model.

Results: Non-lesional biopsies from SLE skin exhibit increased ORF1p protein and increased ORF2 transcript staining and a concurrent increase in phospho-STING staining, suggestive of activation of nucleic acid sensing pathways. UV stimulation increased expression of LINE-1 transcript and protein in both healthy control and SLE skin. Tri-phosphorylated form of RPT-A inhibited enzymatic activity of LINE-1 RT with an IC₅₀ of 0.03 µM. RPT-A inhibited cellular LINE-1 retrotransposition, decitabine-induced interferon response in TREX1 deficient THP-1 cells, and UV-induced phospho-TBK1 in human HaCaT keratinocytes. LINE-1 knockdown with shRNA and siRNA in the THP1 and HaCaT cells mimicked the inhibitory effect of RPT-A. Keratinocyte cell lines derived from lupus patients pre-treated with RPT-A exhibit reduced ISGs upon UV irradiation in a dose-dependent fashion. Further, 5-6 week old TREX1 knockout mice dosed with RPT-A orally once a day for 28 days exhibited reduced serum anti-dsDNA antibodies, reduced myocardial ISGs and reduced immune infiltrates in the heart and kidney.

Conclusion: LINE-1 RNA and protein are increased in SLE skin and is induced by UV exposure. Inhibition of LINE-1 RT activity results in decreased ISG expression and improved disease activity in a murine interferonopathy model. Inhibition of LINE-1 reverse transcriptase holds promise as a novel therapy for diseases in which type I IFNs drive disease.

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ACR Meeting Abstracts - https://acrabstracts.org/abstract/elevated-line-1-expression-in-sle-keratinocytes-leads-to-line-1-reverse-transcriptase-dependent-type-i-interferon-responses/