title: A new nucleic acid-based agent inhibits cytotoxic T lymphocyte-mediated immune disorders.
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.
  
  
  OBJECTIVE
  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.
authors.affiliation: 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.
authors.forename: Chuang-Wei
authors.initials: CW
authors.lastname: Wang
authors.affiliation: Department of Dermatology, Drug Hypersensitivity Clinical and Research Center, Chang Gung Memorial Hospitals, Keelung, Linko, Chang Gung University College of Medicine, Taoyuan, Taiwan.
authors.forename: Wen-Hung
authors.initials: WH
authors.lastname: Chung
authors.affiliation: Institute of Biomedical Science, Academia Sinica, Taipei, Taiwan.
authors.forename: Yi-Fang
authors.initials: YF
authors.lastname: Cheng
authors.affiliation: Institute of Biomedical Science, Academia Sinica, Taipei, Taiwan.
authors.forename: Nien-Wen
authors.initials: NW
authors.lastname: Ying
authors.affiliation: Institute of Biomedical Science, Academia Sinica, Taipei, Taiwan.
authors.forename: Konan
authors.initials: K
authors.lastname: Peck
authors.affiliation: Institute of Biomedical Science, Academia Sinica, Taipei, Taiwan.
authors.forename: Yuan-Tsong
authors.initials: YT
authors.lastname: Chen
authors.affiliation: 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.
authors.forename: Shuen-Iu
authors.initials: SI
authors.lastname: Hung
mesh_terms: D000945:Antigens, Differentiation, T-Lymphocyte / Q000235:genetics*; D052157:Aptamers, Nucleotide / Q000008:administration & dosage*; D016827:CD8 Antigens / Q000235:genetics*; D006086:Graft vs Host Disease / Q000276:immunology; D006801:Humans; D034741:RNA, Small Interfering / Q000008:administration & dosage*; D013262:Stevens-Johnson Syndrome / Q000276:immunology; D013602:T-Lymphocytes, Cytotoxic / Q000276:immunology*
chemical_list: D000945:Antigens, Differentiation, T-Lymphocyte; D052157:Aptamers, Nucleotide; D016827:CD8 Antigens; C075005:GNLY protein, human; D034741:RNA, Small Interfering
keywords: Alloreactivity; CD8; CON; CTL; Cytotoxic T lymphocyte; Dissociation constant; GNLY; GVHD; Graft-versus-host disease; Granulysin; K(d); NF-κB; NK; Natural killer; Nonsilencing control; Nuclear factor κB; RNA interference; RNAi; SEAP; SELEX; SJS; Secreted embryonic alkaline phosphatase; Single-stranded DNA; Small interfering RNA; Stevens-Johnson syndrome; Systematic evolution of ligands by exponential enrichment; T-cell receptor; TCR; TEN; Toxic epidermal necrolysis; aptamer; drug hypersensitivity; graft-versus-host disease; granulysin; nucleic acid–based therapeutics; siRNA; ssDNA; systematic evolution of ligands by exponential enrichment; toxic epidermal necrolysis