Abstract 3826: Phenethyl isothiocyanate hampers growth and progression of HER2-positive breast cancers

Abstract

Abstract Background: Almost 90% of all breast cancers (BCs) express a splice variant of HER2 lacking exon 16 (d16HER2), which promotes the generation of stable and active d16HER2 homodimers on the tumor cell surface. Comparison of the tumorigenic potential of human d16HER2 and the full-length HER2 (WTHER2) in transgenic mice revealed a significantly shorter latency period and a higher incidence of tumors in the d16HER2 line, suggesting enrichment of this variant in HER2+ BC stem cells. Based on reports that cruciferous vegetable-derived compounds such as isothiocyanates (ITCs) mediate strong anti-tumorigenic effects on numerous oncotypes and target BC stem cells, we tested the effect of the phenethyl ITC (PEITC), alone or together with Trastuzumab (T), which reportedly targets HER2+ BC stem cells, on d16HER2-positive tumor growth and progression in transgenic mice. Methods: The percentage of tumor cells expressing high levels of CD29 and CD24 and low levels of SCA-1 (CD29high/CD24+/SCA-1low) murine stem cell markers was evaluated by multiparametric flow cytometry in cell models derived from spontaneous d16HER2 and WTHER2 transgenic lesions. Mammosphere forming efficiency (MFE%) was calculated in d16HER2- and WTHER2-positive tumor cells. The therapeutic activity of PEITC, T and their combination was assessed in vitro based on their ability to decrease the MFE% and the expression of stem cell markers in d16HER2- vs. WTHER2-positive tumor cells, and in vivo in d16HER2 transgenic mice. Analyses by western blot, histology/immunohistochemistry and magnetic resonance spectroscopy (MRS) are ongoing to assess apoptosis, necrosis and metabolomic profiles of treated tumors vs. controls. Results: d16HER2 tumor cells showed a significantly higher MFE% and higher expression of stem cell markers than did WTHER2 tumor cells. PEITC and T significantly reduced MFE% as compared to controls (p = 0.0342 and p = 0.0083, respectively), and their combination exerted significantly stronger inhibition than T (p = 0.001) or PEITC (p = 0.0003) alone only in d16HER2 cells. The percentage of tumor cells expressing CD29high/CD24+/SCA-1low in d16HER2 tumor cells after treatment with T, PEITC or both was also reduced. Whereas PEITC monotherapy did not significantly reduce mammary tumor incidence compared to controls, while T effectively suppressed d16HER2-driven tumor growth (p = 0.0002), the combination of PEITC and T significantly heightened the impairment of mammary lesion development compared to T alone (p = 0.0046). Ex vivo metabolomic analyses by MRS of d16HER2-positive murine and human BC cell lines showed an increase in glycolytic pathway as compared to the respective WTHER2 cells. Conclusions: Our results provide in vivo evidence that the combination of T and PEITC mediates a significantly greater anti-tumor effect than either alone, most likely targeting BC stem cells. Supported by the Italian Ministry of Health and AIRC. Citation Format: Ada Koschorke, Lorenzo Castagnoli, Gaia C. Ghedini, Tiziana Triulzi, Claudia Chiodoni, Rossella Canese, Egidio Iorio, Manuela Iezzi, Patrizia Nanni, Elda Tagliabue, Serenella M. Pupa. Phenethyl isothiocyanate hampers growth and progression of HER2-positive breast cancers. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 3826.