Abstract PS10-21: Identifying efficacy of targeted HER2 antibodies in sensitization of HER2 positive breast cancer to fractionated radiation
Authors: Song Patrick N. , Lu Yun , Napier Tiara , Samuel Sharon , Heinzman Katherine , Lapi Suzanne E. , Sorace Anna G. 
Published: February 2021

Introduction: Tumor hypoxia contributes to intra-tumoral heterogeneity and decreases efficacy of cytotoxic therapy. Fractionated radiation therapy has emerged as an alternative to single dose radiation, and is a component of standard-of-care therapy for patients with unresectable human epidermal growth factor receptor 2 (HER2+) breast cancer. Because radiation therapy is dependent on tissue oxygenation, therapies that increase oxygenation could radio-sensitize tumors. The goal of this study is to investigate if molecular imaging with [18F]-fluoromisonidazole (FMISO)-PET can quantify anti-HER2 therapy-induced changes in tumor oxygenation and utilize imaging metrics to enhance the effectiveness of fractionated radiation. Improving treatment synergy in HER2+ breast cancer has potential to increase therapeutic effectiveness without increasing cytotoxic therapy.

Methods: For in vitro studies, HER2+ breast cancer cells (BT474, SKBR3, MDA-MB-361 and MDA-MB-453) were treated with various sequencing of combination trastuzumab (1 µg/mL) and fractionated radiation (6/3 Gy) and assessed for cell death. HER2+ cancer cells (BT474 and MDA-MB-361) were also treated with combination trastuzumab and fractionated radiation and analyzed for DNA double strand breaks (DSB) through flow cytometry. For in vivo studies, HER2+ cell line (BT474 and MDA-MB-361) and HER2+ patient derived xenograft (BCM 3472) tumors were engrafted into mice and treated with trastuzumab (4 mg/kg) on days 0 and 3 and fractionated radiation (6/3 Gy) on days 1, 2 and 3 (or single agent control). [18F]-FMISO-PET imaging was conducted on day 0, 3 and 7. At the imaging endpoint, tumors were either extracted for biological validation or continued to measure tumor size changes for longitudinal assessment of response. Bliss test of independence and a non-parametric T-test was used to assess for treatment synergy and significance, respectively.

Results: In vitro cell death assay revealed single agent trastuzumab or fractionated radiation treated groups exhibited 15.2% ± 6.5% or 53.6% ± 9% cell death respectively, while trastuzumab prior to fractionated radiation groups exhibited 72.5% ± 2.5% cell death (p = 0.01) on day 7 in MDA-MB-361 cells. Flow cytometry analysis showed MDA-MB-361 cells treated with trastuzumab prior to fractionated radiation exhibited 62.4% ± 8.7% DSB, which significantly increased from single agent trastuzumab (0.74% ± 0.39%) or fractionated radiation groups (37.3% ± 7.3%) (p = 0.01). In vivo, MDA-MB-361 tumors treated with trastuzumab and fractionated radiation had a [18F]-FMISO SUVmean of 0.20 ± 0.09, while tumors treated with fractionated radiation had a [18F]-FMISO SUVmean of 0.31 ± 0.06 on day 7 (p = 0.05). MDA-MB-361 tumors treated with trastuzumab and fractionated radiation experienced a 26.1% ± 16.8% decrease in tumor volume, while tumors treated with single agent fractionated radiation experienced a 10.4% ± 2.8% decrease in tumor volume from day 0 to day 7 (p = 0.11). 30 days after start of therapy, MDA-MB-361 tumors treated with single agent fractionated radiation had a tumor volume of 471.8 ± 120 mm3, whereas tumors treated with combination trastuzumab and fractionated radiation had a tumor volume of 116 ± 38 mm3 (p &lt; 0.01). Bliss test of independence confirmed in vivo treatment synergy of trastuzumab and fractionated radiation starting 14 days after start of therapy.

Conclusion: HER2+ breast cancer treated with trastuzumab prior to fractionated radiation synergistically increases efficacy of radiotherapy in vitro and in vivo. [18F]-FMISO-PET imaging has potential to identify in vivo response to combination therapies and better understand changes in the tumor microenvironment to guide combination therapy.

Acknowledgements: We thank the American Cancer Society for RSG-18-006-01-CCE and NIH NCI R01 CA240589.

Citation Format: Patrick N. Song, Yun Lu, Tiara Napier, Sharon Samuel, Katherine Heinzman, Suzanne E. Lapi, Anna G. Sorace. Identifying efficacy of targeted HER2 antibodies in sensitization of HER2 positive breast cancer to fractionated radiation [abstract]. In: Proceedings of the 2020 San Antonio Breast Cancer Virtual Symposium; 2020 Dec 8-11; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2021;81(4 Suppl):Abstract nr PS10-21.