Novel anti-tumor mechanism of galanin receptor type 2 in head and neck squamous cell carcinoma cells

Authors

  • Takayuki Uehara,

    1. Department of Otorhinolaryngology, Head and Neck Surgery, Graduate School of Medicine, University of the Ryukyus, Nishihara, Japan
    2. Division of Genetic Therapeutics, Center for Molecular Medicine, Jichi Medical University, Shimotsuke, Japan
    3. Department of Otolaryngology, Head and Neck Surgery, Jichi Medical University School of Medicine, Shimotsuke, Japan
    Search for more papers by this author
  • Takeharu Kanazawa,

    Corresponding author
    1. Department of Otolaryngology, Head and Neck Surgery, Jichi Medical University School of Medicine, Shimotsuke, Japan
    • Takeharu Kanazawa, Department of Otolaryngology, Head and Neck Surgery, Jichi Medical University School of Medicine, 3311-1 Yakushiji, Shimotsuke 329-0498, Japan.

      Tel: 0285-58-7381; Fax: 0285-44-5547;

      E-mail: kanatake@omiya.jichi.ac.jp

    Search for more papers by this author
    • These three authors made an equal contribution towards the conduct and reporting of this study.
  • Hiroaki Mizukami,

    1. Division of Genetic Therapeutics, Center for Molecular Medicine, Jichi Medical University, Shimotsuke, Japan
    Search for more papers by this author
    • These three authors made an equal contribution towards the conduct and reporting of this study.
  • Ryosuke Uchibori,

    1. Division of Genetic Therapeutics, Center for Molecular Medicine, Jichi Medical University, Shimotsuke, Japan
    Search for more papers by this author
  • Tomonori Tsukahara,

    1. Division of Genetic Therapeutics, Center for Molecular Medicine, Jichi Medical University, Shimotsuke, Japan
    Search for more papers by this author
  • Masashi Urabe,

    1. Division of Genetic Therapeutics, Center for Molecular Medicine, Jichi Medical University, Shimotsuke, Japan
    Search for more papers by this author
  • Akihiro Kume,

    1. Division of Genetic Therapeutics, Center for Molecular Medicine, Jichi Medical University, Shimotsuke, Japan
    Search for more papers by this author
  • Kiyoshi Misawa,

    1. Department of Otolaryngology, Head and Neck Surgery, Hamamatsu University School of Medicine, Hamamatsu, Japan
    Search for more papers by this author
  • Thomas E. Carey,

    1. Laboratory of Head and Neck Center Biology, Department of Otolaryngology, Head and Neck Surgery, The University of Michigan, Ann Arbor, Michigan, USA
    Search for more papers by this author
  • Mikio Suzuki,

    1. Department of Otorhinolaryngology, Head and Neck Surgery, Graduate School of Medicine, University of the Ryukyus, Nishihara, Japan
    Search for more papers by this author
  • Keiichi Ichimura,

    1. Department of Otolaryngology, Head and Neck Surgery, Jichi Medical University School of Medicine, Shimotsuke, Japan
    Search for more papers by this author
  • Keiya Ozawa

    1. Division of Genetic Therapeutics, Center for Molecular Medicine, Jichi Medical University, Shimotsuke, Japan
    Search for more papers by this author
    • These three authors made an equal contribution towards the conduct and reporting of this study.

Abstract

Galanin and its receptors, GALR1 and GALR2, are known tumor suppressors and potential therapeutic targets in head and neck squamous cell carcinoma (HNSCC). Previously, we demonstrated that, in GALR1-expressing HNSCC cells, the addition of galanin suppressed tumor proliferation via upregulation of ERK1/2 and cyclin-dependent kinase inhibitors, whereas, in GALR2-expressing cells, the addition of galanin not only suppressed proliferation, but also induced apoptosis. In this study, we first transduced HEp-2 and KB cell lines using a recombinant adeno-associated virus (rAAV)-green fluorescent protein (GFP) vector and confirmed a high GFP expression rate (>90%) in both cell lines at the standard vector dose. Next, we demonstrated that GALR2 expression in the presence of galanin suppressed cell viability to 40–60% after 72 h in both cell lines. Additionally, the annexin V-positive rate and sub-G0/G1 phase population were significantly elevated in HEp-2 cells (mock vs GALR2: 12.3 vs 25.0% (P < 0.01) and 9.1 vs 32.0% (P < 0.05), respectively) after 48 h. These changes were also observed in KB cells, although to a lesser extent. Furthermore, in HEp-2 cells, GALR2-mediated apoptosis was caspase-independent, involving downregulation of ERK1/2, followed by induction of the pro-apoptotic Bcl-2 protein, Bim. These results illustrate that transient GALR2 expression in the presence of galanin induces apoptosis via diverse pathways and serves as a platform for suicide gene therapy against HNSCC.

Ancillary