Chuang-Wei Wang1, Wen-Hung Chung2, Yi-Fang Cheng3, Nien-Wen Ying3, Konan Peck3, Yuan-Tsong Chen3, Shuen-Iu Hung4. 1. Department and Institute of Pharmacology, School of Medicine, Infection and Immunity Research Center, National Yang-Ming University, Taipei, Taiwan; Molecular Medicine Program, National Yang-Ming University and Academia Sinica, Taipei, Taiwan. 2. Department of Dermatology, Drug Hypersensitivity Clinical and Research Center, Chang Gung Memorial Hospitals, Keelung, Linko, Chang Gung University College of Medicine, Taoyuan, Taiwan. 3. Institute of Biomedical Science, Academia Sinica, Taipei, Taiwan. 4. Department and Institute of Pharmacology, School of Medicine, Infection and Immunity Research Center, National Yang-Ming University, Taipei, Taiwan; Molecular Medicine Program, National Yang-Ming University and Academia Sinica, Taipei, Taiwan. Electronic address: sihung@ym.edu.tw.
Abstract
BACKGROUND: Stevens-Johnson syndrome (SJS), toxic epidermal necrolysis (TEN), and graft-versus-host disease (GVHD) are distinct immune reactions elicited by drugs or allogeneic antigens; however, they share a pathomechanism with the activation of cytotoxic T lymphocytes (CTLs). CTLs produce cytotoxic proteins, cytokines, chemokines, or immune alarmins, such as granulysin (GNLY), leading to the extensive tissue damage and systemic inflammation seen in patients with SJS/TEN or GVHD. Currently, there is no effective therapeutic agent specific for CTL-mediated immune disorders. OBJECTIVES: By targeting GNLY(+) CTLs, we aimed to develop a nucleic acid-based agent consisting of an anti-CD8 aptamer with GNLY small interfering RNA (siRNA). METHODS: We performed systematic evolution of ligands using exponential enrichment to select and identify effective anti-CD8 aptamers. We developed an aptamer-siRNA chimera using a "sticky bridge" method by conjugating the aptamer with siRNA. We analyzed the inhibitory effects of the aptamer-siRNA chimera on CTL responses in patients with SJS/TEN or GVHD. RESULTS: We identified a novel DNA aptamer (CD8AP17s) targeting CTLs. This aptamer could be specifically internalized into human CTLs. We generated the CD8AP17s aptamer-GNLY siRNA chimera, which showed a greater than 79% inhibitory effect on the production of GNLY by drug/alloantigen-activated T cells. The CD8AP17s aptamer-GNLY siRNA chimera decreased cytotoxicity in in vitro models of both SJS/TEN (elicited by drug-specific antigen) and GVHD (elicited by allogeneic antigens). CONCLUSIONS: Our results identified a new nucleic acid-based agent (CD8 aptamer-GNLY siRNA chimera) that can significantly inhibit CTL-mediated drug hypersensitivity, such as that seen in patients with SJS/TEN, as well as the alloreactivity seen in patients with GVHD. This study provides a novel therapeutic strategy for CTL-mediated immune disorders.
BACKGROUND:Stevens-Johnson syndrome (SJS), toxic epidermal necrolysis (TEN), and graft-versus-host disease (GVHD) are distinct immune reactions elicited by drugs or allogeneic antigens; however, they share a pathomechanism with the activation of cytotoxic T lymphocytes (CTLs). CTLs produce cytotoxic proteins, cytokines, chemokines, or immune alarmins, such as granulysin (GNLY), leading to the extensive tissue damage and systemic inflammation seen in patients with SJS/TEN or GVHD. Currently, there is no effective therapeutic agent specific for CTL-mediated immune disorders. OBJECTIVES: By targeting GNLY(+) CTLs, we aimed to develop a nucleic acid-based agent consisting of an anti-CD8 aptamer with GNLY small interfering RNA (siRNA). METHODS: We performed systematic evolution of ligands using exponential enrichment to select and identify effective anti-CD8 aptamers. We developed an aptamer-siRNA chimera using a "sticky bridge" method by conjugating the aptamer with siRNA. We analyzed the inhibitory effects of the aptamer-siRNA chimera on CTL responses in patients with SJS/TEN or GVHD. RESULTS: We identified a novel DNA aptamer (CD8AP17s) targeting CTLs. This aptamer could be specifically internalized into human CTLs. We generated the CD8AP17s aptamer-GNLY siRNA chimera, which showed a greater than 79% inhibitory effect on the production of GNLY by drug/alloantigen-activated T cells. The CD8AP17s aptamer-GNLY siRNA chimera decreased cytotoxicity in in vitro models of both SJS/TEN (elicited by drug-specific antigen) and GVHD (elicited by allogeneic antigens). CONCLUSIONS: Our results identified a new nucleic acid-based agent (CD8 aptamer-GNLY siRNA chimera) that can significantly inhibit CTL-mediated drug hypersensitivity, such as that seen in patients with SJS/TEN, as well as the alloreactivity seen in patients with GVHD. This study provides a novel therapeutic strategy for CTL-mediated immune disorders.
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