Cancer Therapy: Preclinical A Novel Toll-Like Receptor 9 Agonist Cooperates with Trastuzumab in Trastuzumab-Resistant Breast Tumors through Multiple Mechanisms of Action

Purpose: Resistance to anti-HER2 monoclonal antibody trastuzumab is a relevant issue in breast cancer patients. Among the mechanisms implicated in trastuzumab resistance, increasing evidence supports a role of tumor microenvironment. We previously found that a novel toll-like receptor 9 agonist, referred to as immune modulatory oligonucleotide (IMO) and currently under clinical investigation, acts through epidermal growth factor receptor (EGFR) and showsdirect antiangiogenic effects by cooperatingwith anti-EGFR or anti-VEGF drugs, thus interfering with cancer cells and microenvironment. Experimental Design: In this study, we used KPL-4 and JIMT-1 trastuzumab-resistant breast cancer cells to evaluate the combination IMO plus trastuzumab as a therapeutic option for trastuzumab-resistant breast cancers. Results: IMO inhibits KPL-4 and JIMT-1 xenografts growth and potentiates trastuzumab antitumor effect, with complete suppression of tumor growth, potent enhancement of trastuzumab-mediated antibody-dependent cell-mediated cytotoxicity, and strong inhibition of EGFR/HER2-related signaling. In KPL-4 xenografts, IMO alone interferes with HER signal transduction, whereas trastuzumab is ineffective. IMO induces an HERdependent signal inhibition also in vitro by modulating a functional interaction between toll-like receptor 9 and HER receptors occurring at membrane level. Finally, IMO plus trastuzumab produces a cooperative antiangiogenic effect related to suppression of endothelial HER-related signaling. Conclusions: We showed a cooperative effect of IMO plus trastuzumab in trastuzumabresistant breast cancers due to IMO direct antitumor and antiangiogenic activity and antibody-dependent cell-mediated cytotoxicity enhancement. Moreover, we provided first evidence of a toll-like receptor 9/HER interaction at membrane level as novel mechanism of action. Altogether, we propose IMO plus trastuzumab as an effective strategy in trastuzumab-resistant breast cancers. (Clin Cancer Res 2009;15(22):6921–30) The aberrant activity of HER2 signaling is associated with breast cancer development and progression, and ∼20% to 25% of invasive breast cancers exhibit overexpression of HER2. Because elevated HER2 levels are associated with reduced disease-free and overall survival in metastatic breast cancer, therapeutic strategies are being developed to target this oncoprotein (1). Trastuzumab, a recombinant humanized monoclonal antibody (mAb) directed against an extracellular region of HER2, was the first HER2-targeted therapy approved by the U.S. Food and Drug Administration for the treatment of HER2-overexpressing metastatic breast cancer. In addition, trastuzumab with adjuvant chemotherapy (either in sequence or in combination) significantly improved disease-free and overall survival rates in patients with early stage HER2overexpressing breast cancer (2). However, despite the great Authors' Affiliations: Departments of Endocrinologia ed Oncologia Molecolare e Clinica, and Biologia e Patologia Cellulare e Molecolare “L. Califano,” Università di Napoli Federico II, Napoli, Italy; and Idera Pharmaceuticals, Cambridge, Massachusetts Received 6/22/09; revised 7/27/09; accepted 8/14/09; published OnlineFirst 11/10/09. Grant support: TheMinistry of Health, TheMinistry of Education, University and Research (PRIN), and the Regione Campania, a fellowship from the Associazione Italiana per la Ricerca sul Cancro (V. Damiano). The costs of publication of this article were defrayed in part by the payment of page charges. This articlemust therefore be herebymarked advertisement in accordance with 18 U.S.C. Section 1734 solely to indicate this fact. Note: V. Damiano and S. Garofalo contributed equally to this work. Requests for reprints: Giampaolo Tortora, Cattedra di Oncologia Medica, Dipartimento Endocrinologia e Oncologia Molecolare e Clinica, Università di Napoli Federico II, Via S. Pansini 5, 80131 Napoli, Italy. Phone: 39-0817462061; Fax: 39-081-7462061; E-mail: [email protected]. F 2009 American Association for Cancer Research. doi:10.1158/1078-0432.CCR-09-1599 6921 Clin Cancer Res 2009;15(22) November 15, 2009 www.aacrjournals.org Research. on June 6, 2017. © 2009 American Association for Cancer clincancerres.aacrjournals.org Downloaded from Published OnlineFirst November 10, 2009; DOI: 10.1158/1078-0432.CCR-09-1599