Molecular targeting of HER2-overexpressing biliary tract cancer cells with trastuzumab emtansine, an antibody–cytotoxic drug conjugate

  • Yoriko Yamashita-Kashima
  • Yasushi Yoshimura
  • Takaaki Fujimura
  • Sei Shu
  • Mieko Yanagisawa
  • Keigo Yorozu
  • Koh Furugaki
  • Ryota Higuchi
  • Junichi Shoda
  • Naoki HaradaEmail author
Original Article



Trastuzumab emtansine (T-DM1) provides clinical benefit in breast cancers overexpressing human epidermal growth factor receptor 2 (HER2). However, its efficacy against biliary tract cancers (BTC) has not been evaluated. In this study, the effectiveness of T-DM1 in various BTC cell lines and xenograft models with different levels of HER2 expression was investigated.


HER2 expression status in xenografts and patient tissue microarrays was assessed by immunohistochemistry (IHC) or fluorescence in situ hybridization (FISH). Cell-surface HER2 expression levels and cell growth inhibition in response to T-DM1 were examined in 17 BTC cell lines. The antitumor activity of T-DM1 was evaluated in four xenograft mouse models with different levels of HER2 expression. The effects of T-DM1 on HER2 signaling, antibody-dependent cell-mediated cytotoxicity (ADCC), cell cycle, and apoptosis were assessed in vitro.


Cell-surface expression of HER2 was observed in both gallbladder carcinoma and cholangiocarcinoma tissues. The anti-proliferative activity of T-DM1 was higher in BTC cell lines and breast cancer cell lines with higher levels of HER2 expression. The HER2 status (IHC score|HER2-to-CEP17 ratio by FISH testing) of each BTC xenograft was 3 +|8.3 for KMCH-1, 2 +|4.7 for Mz-ChA-1, 1 +/0|1.4 for OCUG-1, and 0|1.1 for KKU-100, and T-DM1 showed antitumor activity in proportion to the HER2 status. T-DM1 inhibited HER2 signaling and induced ADCC, mitotic arrest, and apoptosis in KMCH-1 cells.


T-DM1 exhibited preclinical activity in HER2-overexpressing BTC. Further evaluation in clinical studies is warranted.


T-DM1 Trastuzumab emtansine HER2 Biliary tract cancer 



We thank Marie Mochizuki and Mirei Kouno (Product Research Department, Chugai Pharmaceutical Co., Ltd.) for technical assistance; Dr. Kaori Fujimoto-Ouchi (Product Research Department, Chugai Pharmaceutical Co., Ltd.) for helpful suggestions and comments about this study; and Dr. Banchob Spira (Khon Kaen University) for kindly providing KKU-100, KKU-055 and KKU-0213 cell lines.

Compliance with ethical standards

Conflict of interest

Junichi Shoda received research fee and grants from Chugai Pharmaceutical Co., Ltd. and other authors have no conflict of interest.

Ethical standards

All procedures performed in studies involving human participants were in accordance with the ethical standards of the institutional research committee at Chugai Pharmaceuticals, Co., Ltd. and with the 1964 Helsinki declaration and its later amendments or comparable ethical standards. The human tissues were processed anonymously inhibiting the revelation of the donor’s identity.

Supplementary material

280_2019_3768_MOESM1_ESM.docx (21 kb)
Supplementary material 1 (DOCX 21 KB)


  1. 1.
    Chan E, Berlin J (2015) Biliary tract cancers: understudied and poorly understood. J Clin Oncol 33(16):1845–1848. CrossRefGoogle Scholar
  2. 2.
    Recio-Boiles A, Babiker HM (2017) Cancer, gallbladder. In: StatPearls. StatPearls, Treasure IslandGoogle Scholar
  3. 3.
    Furuse J, Okusaka T (2011) Targeted therapy for biliary tract cancer. Cancers (Basel) 3(2):2243CrossRefGoogle Scholar
  4. 4.
    Yarden Y, Sliwkowski MX (2001) Untangling the ErbB signalling network. Nat Rev Mol Cell Biol 2:127. CrossRefGoogle Scholar
  5. 5.
    Wang Z (2017) ErbB receptors and cancer. In: Wang Z (ed) ErbB receptor signaling: methods and protocols. Springer, New York, pp 3–35. CrossRefGoogle Scholar
  6. 6.
    Chua TC, Merrett ND (2012) Clinicopathologic factors associated with HER2-positive gastric cancer and its impact on survival outcomes—a systematic review. Int J Cancer 130(12):2845–2856. CrossRefGoogle Scholar
  7. 7.
    Wong H, Yau T (2012) Targeted therapy in the management of advanced gastric cancer: are we making progress in the era of personalized medicine? Oncologist 17(3):346–358. CrossRefGoogle Scholar
  8. 8.
    Terashima M, Kitada K, Ochiai A, Ichikawa W, Kurahashi I, Sakuramoto S, Katai H, Sano T, Imamura H, Sasako M, Group A-G (2012) Impact of expression of human epidermal growth factor receptors EGFR and ERBB2 on survival in stage II/III gastric cancer. Clin Cancer Res 18(21):5992–6000. CrossRefGoogle Scholar
  9. 9.
    Galdy S, Lamarca A, McNamara MG, Hubner RA, Cella CA, Fazio N, Valle JW (2017) HER2/HER3 pathway in biliary tract malignancies; systematic review and meta-analysis: a potential therapeutic target? Cancer Metastasis Rev 36(1):141–157. CrossRefGoogle Scholar
  10. 10.
    Nam AR, Kim JW, Cha Y, Ha H, Park JE, Bang JH, Jin MH, Lee KH, Kim TY, Han SW, Im SA, Kim TY, Oh DY, Bang YJ (2016) Therapeutic implication of HER2 in advanced biliary tract cancer. Oncotarget 7(36):58007–58021. CrossRefGoogle Scholar
  11. 11.
    Kawamoto T, Ishige K, Thomas M, Yamashita-Kashima Y, Shu S, Ishikura N, Ariizumi S, Yamamoto M, Kurosaki K, Shoda J (2015) Overexpression and gene amplification of EGFR, HER2, and HER3 in biliary tract carcinomas, and the possibility for therapy with the HER2-targeting antibody pertuzumab. J Gastroenterol 50(4):467–479. CrossRefGoogle Scholar
  12. 12.
    Erickson HK, Park PU, Widdison WC, Kovtun YV, Garrett LM, Hoffman K, Lutz RJ, Goldmacher VS, Blattler WA (2006) Antibody-maytansinoid conjugates are activated in targeted cancer cells by lysosomal degradation and linker-dependent intracellular processing. Cancer Res 66(8):4426–4433. CrossRefGoogle Scholar
  13. 13.
    Barok M, Tanner M, Koninki K, Isola J (2011) Trastuzumab-DM1 causes tumour growth inhibition by mitotic catastrophe in trastuzumab-resistant breast cancer cells in vivo. Breast Cancer Res 13(2):R46. CrossRefGoogle Scholar
  14. 14.
    Oroudjev E, Lopus M, Wilson L, Audette C, Provenzano C, Erickson H, Kovtun Y, Chari R, Jordan MA (2010) Maytansinoid-antibody conjugates induce mitotic arrest by suppressing microtubule dynamic instability. Mol Cancer Ther 9(10):2700–2713. CrossRefGoogle Scholar
  15. 15.
    Junttila TT, Li G, Parsons K, Phillips GL, Sliwkowski MX (2011) Trastuzumab-DM1 (T-DM1) retains all the mechanisms of action of trastuzumab and efficiently inhibits growth of lapatinib insensitive breast cancer. Breast Cancer Res Treat 128(2):347–356. CrossRefGoogle Scholar
  16. 16.
    Yamashita-Kashima Y, Shu S, Yorozu K, Moriya Y, Harada N (2017) Mode of action of pertuzumab in combination with trastuzumab plus docetaxel therapy in a HER2-positive breast cancer xenograft model. Oncol Lett 14(4):4197–4205. CrossRefGoogle Scholar
  17. 17.
    Yamashita-Kashima Y, Iijima S, Yorozu K, Furugaki K, Kurasawa M, Ohta M, Fujimoto-Ouchi K (2011) Pertuzumab in combination with trastuzumab shows significantly enhanced antitumor activity in HER2-positive human gastric cancer xenograft models. Clin Cancer Res 17(15):5060–5070. CrossRefGoogle Scholar
  18. 18.
    Yamashita-Kashima Y, Shu S, Yorozu K, Hashizume K, Moriya Y, Fujimoto-Ouchi K, Harada N (2014) Importance of formalin fixing conditions for HER2 testing in gastric cancer: immunohistochemical staining and fluorescence in situ hybridization. Gastric Cancer 17(4):638–647. CrossRefGoogle Scholar
  19. 19.
    Wolff AC, Hammond MEH, Hicks DG, Dowsett M, McShane LM, Allison KH, Allred DC, Bartlett JMS, Bilous M, Fitzgibbons P, Hanna W, Jenkins RB, Mangu PB, Paik S, Perez EA, Press MF, Spears PA, Vance GH, Viale G, Hayes DF (2013) Recommendations for Human Epidermal Growth Factor Receptor 2 testing in breast cancer: American Society of Clinical Oncology/College of American Pathologists Clinical Practice Guideline Update. J Clin Oncol 31(31):3997–4013. CrossRefGoogle Scholar
  20. 20.
    Stephen L, Bastiaan DBW, Soraya F, Michael P, Priyanthi KM (2011) Human epidermal growth factor receptor 2 testing in gastric carcinoma: issues related to heterogeneity in biopsies and resections*. Histopathology 59(5):832–840. CrossRefGoogle Scholar
  21. 21.
    Tanner M, Hollmén M, Junttila TT, Kapanen AI, Tommola S, Soini Y, Helin H, Salo J, Joensuu H, Sihvo E, Elenius K, Isola J (2005) Amplification of HER-2 in gastric carcinoma: association with Topoisomerase IIα gene amplification, intestinal type, poor prognosis and sensitivity to trastuzumab. Ann Oncol 16(2):273–278. CrossRefGoogle Scholar
  22. 22.
    Fujimoto-Ouchi K, Sekiguchi F, Yasuno H, Moriya Y, Mori K, Tanaka Y (2007) Antitumor activity of trastuzumab in combination with chemotherapy in human gastric cancer xenograft models. Cancer Chemother Pharmacol 59(6):795–805. CrossRefGoogle Scholar
  23. 23.
    Li Y, Zhang R, Han Y, Lu T, Ding J, Zhang K, Lin G, Xie J, Li J (2016) Comparison of the types of candidate reference samples for quality control of human epidermal growth factor receptor 2 status detection. Diagn Pathol 11(1):85. CrossRefGoogle Scholar
  24. 24.
    Kim SY, Kim HP, Kim YJ, Oh DY, Im SA, Lee D, Jong HS, Kim TY, Bang YJ (2008) Trastuzumab inhibits the growth of human gastric cancer cell lines with HER2 amplification synergistically with cisplatin. Int J Oncol 32(1):89–95. Google Scholar
  25. 25.
    Beeram M, Krop IE, Burris HA, Girish SR, Yu W, Lu MW, Holden SN, Modi S (2012) A phase 1 study of weekly dosing of trastuzumab emtansine (T-DM1) in patients with advanced human epidermal growth factor 2-positive breast cancer. Cancer 118(23):5733–5740. CrossRefGoogle Scholar
  26. 26.
    Lewis Phillips GD, Li G, Dugger DL, Crocker LM, Parsons KL, Mai E, Blattler WA, Lambert JM, Chari RV, Lutz RJ, Wong WL, Jacobson FS, Koeppen H, Schwall RH, Kenkare-Mitra SR, Spencer SD, Sliwkowski MX (2008) Targeting HER2-positive breast cancer with trastuzumab-DM1, an antibody-cytotoxic drug conjugate. Cancer Res 68(22):9280–9290. CrossRefGoogle Scholar
  27. 27.
    Erickson HK, Park PU, Widdison WC, Kovtun YV, Garrett LM, Hoffman K, Lutz RJ, Goldmacher VS, Blättler WA (2006) Antibody-maytansinoid conjugates are activated in targeted cancer cells by lysosomal degradation and linker-dependent intracellular processing. Cancer Res 66(8):4426–4433. CrossRefGoogle Scholar
  28. 28.
    Baselga J, Swain SM (2009) Novel anticancer targets: revisiting ERBB2 and discovering ERBB3. Nat Rev Cancer 9(7):463–475. CrossRefGoogle Scholar
  29. 29.
    Burris HA 3rd, Rugo HS, Vukelja SJ, Vogel CL, Borson RA, Limentani S, Tan-Chiu E, Krop IE, Michaelson RA, Girish S, Amler L, Zheng M, Chu YW, Klencke B, O’Shaughnessy JA (2011) Phase II study of the antibody drug conjugate trastuzumab-DM1 for the treatment of human epidermal growth factor receptor 2 (HER2)-positive breast cancer after prior HER2-directed therapy. J Clin Oncol 29(4):398–405. CrossRefGoogle Scholar
  30. 30.
    Barok M, Tanner M, Koninki K, Isola J (2011) Trastuzumab-DM1 is highly effective in preclinical models of HER2-positive gastric cancer. Cancer Lett 306(2):171–179. CrossRefGoogle Scholar
  31. 31.
    Yamashita-Kashima Y, Shu S, Harada N, Fujimoto-Ouchi K (2013) Enhanced antitumor activity of trastuzumab emtansine (T-DM1) in combination with pertuzumab in a HER2-positive gastric cancer model. Oncol Rep 30(3):1087–1093. CrossRefGoogle Scholar
  32. 32.
    Cretella D, Saccani F, Quaini F, Frati C, Lagrasta C, Bonelli M, Caffarra C, Cavazzoni A, Fumarola C, Galetti M, La Monica S, Ampollini L, Tiseo M, Ardizzoni A, Petronini PG, Alfieri RR (2014) Trastuzumab emtansine is active on HER-2 overexpressing NSCLC cell lines and overcomes gefitinib resistance. Mol Cancer 13:143. CrossRefGoogle Scholar
  33. 33.
    Hayashi T, Seiler R, Oo HZ, Jager W, Moskalev I, Awrey S, Dejima T, Todenhofer T, Li N, Fazli L, Matsubara A, Black PC (2015) Targeting HER2 with T-DM1, an antibody cytotoxic drug conjugate, is Effective in HER2 over expressing bladder cancer. J Urol 194(4):1120–1131. CrossRefGoogle Scholar
  34. 34.
    Hofmann M, Stoss O, Shi D, Buttner R, van de Vijver M, Kim W, Ochiai A, Ruschoff J, Henkel T (2008) Assessment of a HER2 scoring system for gastric cancer: results from a validation study. Histopathology 52(7):797–805. CrossRefGoogle Scholar
  35. 35.
    Bang YJ, Van Cutsem E, Feyereislova A, Chung HC, Shen L, Sawaki A, Lordick F, Ohtsu A, Omuro Y, Satoh T, Aprile G, Kulikov E, Hill J, Lehle M, Ruschoff J, Kang YK (2010) Trastuzumab in combination with chemotherapy versus chemotherapy alone for treatment of HER2-positive advanced gastric or gastro-oesophageal junction cancer (ToGA): a phase 3, open-label, randomised controlled trial. Lancet 376(9742):687–697. CrossRefGoogle Scholar
  36. 36.
    Van Cutsem E, Bang Y-J, Feng-yi F, Xu JM, Lee K-W, Jiao S-C, Chong JL, López-Sanchez RI, Price T, Gladkov O, Stoss O, Hill J, Ng V, Lehle M, Thomas M, Kiermaier A, Rüschoff J (2015) HER2 screening data from ToGA: targeting HER2 in gastric and gastroesophageal junction cancer. Gastric Cancer 18(3):476–484. CrossRefGoogle Scholar
  37. 37.
    Lee HE, Park KU, Yoo SB, Nam SK, Park DJ, Kim HH, Lee HS (2013) Clinical significance of intratumoral HER2 heterogeneity in gastric cancer. Eur J Cancer 49(6):1448–1457. CrossRefGoogle Scholar

Copyright information

© Springer-Verlag GmbH Germany, part of Springer Nature 2019

Authors and Affiliations

  • Yoriko Yamashita-Kashima
    • 1
  • Yasushi Yoshimura
    • 1
  • Takaaki Fujimura
    • 1
  • Sei Shu
    • 1
  • Mieko Yanagisawa
    • 1
  • Keigo Yorozu
    • 1
  • Koh Furugaki
    • 1
  • Ryota Higuchi
    • 2
  • Junichi Shoda
    • 3
  • Naoki Harada
    • 1
    Email author
  1. 1.Product Research DepartmentChugai Pharmaceutical Co., Ltd.KamakuraJapan
  2. 2.Department of Gastroenterological SurgeryTokyo Women’s Medical UniversityTokyoJapan
  3. 3.Department of Medical Sciences, Faculty of MedicineUniversity of TsukubaTsukubaJapan

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