Abstract 4237: Decreased hypoxia in a HER2⁺ breast cancer model following trastuzumab therapy

Introduction: Hypoxic tumors demonstrate treatment resistance with an increased risk of metastasis. Therefore, alleviating hypoxia has potential to improve efficiency of combination therapies. The primary goal of this study is to quantify alterations in hypoxia in response to trastuzumab in a murine model of HER2+ breast cancer through imaging and histology.

Experimental Design: Mice were implanted subcutaneously with BT474 breast cancer cells (10⁷) and randomly assigned into treated (10 mg/kg trastuzumab) or control (saline) groups. After tumors reached ∼250 mm³, animals (n = 32) were utilized to identify longitudinal changes in functional vascular through an intravenous injection of Hoechst 33342 nuclear stain (immediately prior to sacrifice) and hypoxia through pimonidazole intravenous injection (one hour prior to sacrifice). Tumors were extracted for immunofluorescence between days 0 through 7. Tumor sections were flash frozen and stained with anti-pimonidazole and propidium iodide (nuclear counterstain). Additionally, a set of tumors (n = 36) were sacrificed for immunohistochemistry (days 0 through 7), formalin fixed and stained for CA-IX. All stained sections were scanned in high resolution (20×) and quantitatively analyzed with Leica SCN400 software. Another cohort of animals (n = 10) were imaged with ¹⁸F-FMISO (fluoromisonidazole) PET between days 0 through 7 and quantified via mean tracer concentration (%ID/g).

Results: Immunohistochemistry revealed significantly increased hypoxia in the control group compared to treated on days 3 (p = 0.03) and 7 (p = 0.002), as measured through CA-IX staining. Additionally, on day 4, functional vascular delivery was increased while hypoxia (pimonidazole) decreased in treated tumors compared to control. ¹⁸F-FMISO PET imaging corroborated histology findings with significantly decreased hypoxia in treated tumors compared to control tumors on day 7 (-47%; p<0.0001).

Conclusion: Trastuzumab has been shown to decrease hypoxia, as measured through gold-standard immunofluorescence. Additionally, this trastuzumab-induced improvement in tumor hypoxia can be measured via clinically relevant, noninvasive imaging (¹⁸F-FMISO PET). Temporarily improving the tumor's functional vasculature and decreasing hypoxia during trastuzumab treatment has potential to enhance the effectiveness of combination therapies. Identifying windows of improved vascular and cellular normalization with noninvasive medical imaging provide opportunity to decrease resistance to standard-of-care treatments and optimize therapeutic timing and regimens.

Citation Format: Anna G. Sorace, C. Chad Quarles, Violeta Sanchez, Thomas E. Yankeelov. Decreased hypoxia in a HER2⁺ breast cancer model following trastuzumab therapy. [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 4237.