title: Molecular targeted photoimmunotherapy for HER2-positive human gastric cancer in combination with chemotherapy results in improved treatment outcomes through different cytotoxic mechanisms.
abstract: BACKGROUND
  Photoimmunotherapy (PIT) is a novel type of molecular optical imaging-guided cancer phototherapy based on a monoclonal antibody conjugated to a photosensitizer, IR700, in combination with near-infrared (NIR) light. PIT rapidly causes target-specific cell death by inducing cell membrane damages and appears to be highly effective; however, we have previously demonstrated that tumor recurrences were eventually seen in PIT-treated mice, likely owing to inhomogeneous mAb-IR700 conjugate distribution in the tumor, thus limiting the effectiveness of PIT as a monotherapy. Here, we examined the effects of human epidermal growth factor-2 (HER2)-targeted PIT in combination with 5-fluorouracil (5-FU) compared to PIT alone for HER2-expressing human gastric cancer cells.
  
  
  METHODS
  NCI-N87 cells, HER2-positive human gastric cancer cells, were used for the experiments. Trastuzumab, a monoclonal antibody directed against HER2, was conjugated to IR700. To assess the short-term cytotoxicity and examine the apoptotic effects upon addition of 5-FU in vitro, we performed LIVE/DEAD and caspase-3 activity assays. Additionally, to explore the effects on long-term growth inhibition, trypan blue dye exclusion assay was performed. NCI-N87 tumor xenograft models were prepared for in vivo treatment studies and the tumor-bearing mice were randomized into various treatment groups.
  
  
  RESULTS
  Compared to PIT alone, the combination of HER2-targeted PIT and 5-FU rapidly induced significant cytotoxicity in both the short-term and long-term cytotoxicity assays. While both 5-FU and/or trastuzumab-IR700 conjugate treatment induced an increase in caspase-3 activity, there was no additional increase in caspase-3 activity upon NIR light irradiation after incubation with 5-FU and/or trastuzumab-IR700. The combination of HER2-targeted PIT and 5-FU resulted in greater and longer tumor growth inhibition than PIT monotherapy in vivo. This combined effect of PIT and 5-FU is likely owing to their different mechanisms of inducing tumor cell death, namely necrotic membrane damage by PIT and apoptotic cell death by 5-FU and trastuzumab.
  
  
  CONCLUSIONS
  PIT in combination with 5-FU resulted in enhanced antitumor effects compared to PIT alone for HER2-expressing human gastric cancer in vitro and in vivo. This combination photoimmunochemotherapy represents a practical method for treating human gastric cancer and should be investigated further in the clinical setting.
authors.affiliation: Division of Gastroenterology and Hepatology, Department of Internal Medicine, The Jikei University School of Medicine, 3-25-8 Nishishinbashi, Minato, Tokyo, 105-8461, Japan. kimihiroitou88@gmail.com.
authors.forename: Kimihiro
authors.initials: K
authors.lastname: Ito
authors.affiliation: Division of Gastroenterology and Hepatology, Department of Internal Medicine, The Jikei University School of Medicine, 3-25-8 Nishishinbashi, Minato, Tokyo, 105-8461, Japan. mit@jikei.ac.jp.
authors.forename: Makoto
authors.initials: M
authors.lastname: Mitsunaga
authors.affiliation: Division of Gastroenterology and Hepatology, Department of Internal Medicine, The Jikei University School of Medicine, 3-25-8 Nishishinbashi, Minato, Tokyo, 105-8461, Japan. sarihiro@gmail.com.
authors.forename: Seiji
authors.initials: S
authors.lastname: Arihiro
authors.affiliation: Division of Gastroenterology and Hepatology, Department of Internal Medicine, The Jikei University School of Medicine, 3-25-8 Nishishinbashi, Minato, Tokyo, 105-8461, Japan. m.saruta@jikei.ac.jp.
authors.forename: Masayuki
authors.initials: M
authors.lastname: Saruta
authors.affiliation: Division of Gastroenterology and Hepatology, Department of Internal Medicine, The Jikei University School of Medicine, 3-25-8 Nishishinbashi, Minato, Tokyo, 105-8461, Japan. mat@jikei.ac.jp.
authors.forename: Mika
authors.initials: M
authors.lastname: Matsuoka
authors.affiliation: Molecular Imaging Program, Center for Cancer Research, National Cancer Institute, NIH, Building 10, RoomB3B69, MSC1088, Bethesda, MD, 20892-1088, USA. kobayash@mail.nih.gov.
authors.forename: Hisataka
authors.initials: H
authors.lastname: Kobayashi
authors.affiliation: Division of Gastroenterology and Hepatology, Department of Internal Medicine, The Jikei University School of Medicine, 3-25-8 Nishishinbashi, Minato, Tokyo, 105-8461, Japan. tajiri@jikei.ac.jp.
authors.forename: Hisao
authors.initials: H
authors.lastname: Tajiri
mesh_terms: D000818:Animals; D017209:Apoptosis / Q000187:drug effects; D045744:Cell Line, Tumor; D003131:Combined Modality Therapy; D005472:Fluorouracil / Q000008:administration & dosage; D006801:Humans; D007167:Immunotherapy / Q000379:methods*; D051379:Mice; D058990:Molecular Targeted Therapy; D010789:Phototherapy / Q000379:methods*; D018719:Erb-b2 Receptor Tyrosine Kinases / Q000235:genetics* / Q000276:immunology; D013274:Stomach Neoplasms / Q000235:genetics / Q000276:immunology* / Q000473:pathology / Q000628:therapy*; D000068878:Trastuzumab / Q000008:administration & dosage; D016896:Treatment Outcome; D023041:Xenograft Model Antitumor Assays
chemical_list: D005472:Fluorouracil; D018719:Erb-b2 Receptor Tyrosine Kinases; D000068878:Trastuzumab; C508053:ERBB2 protein, human
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