Abstract 1395: GRP78 is induced and can be targeted after systemic therapy in breast cancer.
Authors: Thaiparambil Jose T. , Wen Jing , Patton Samuel K. , Diaz Roberto 
Published: August 2014

Purpose/Objectives: Currently, there is no effective treatment or drug delivery system to specifically target cancer metastasis. Our approach is to use a systemic agent to induce neoantigens in breast tumors and target these neoantigens using small peptides. The 78-kDa glucose regulated protein (GRP78) is a major endoplasmic reticulum chaperone that suppresses stress-induced apoptosis. Recent studies from our group show that GRP78 is an important neoantigen that is upregulated in response to ionizing radiation. We tested the hypothesis that GRP78 is induced by systemic agents and use our GIRLRG targeting peptide to recognize this receptor on the surface of breast tumors after treatment with systemic agents. Methods: We used ex-vivo immunohistochemistry (IHC) analyses to determine the levels of GRP78 in breast tumors. We used an in vivo imaging system to determine peptide binding on breast tumors in mice. Nude mice were injected with MDA-MB-231 tumor cells in both hind limbs. Mice were treated with vehicle control, sunitinib, sorafenib, pazopanib or 5-FU once daily for 5 consecutive days. The tumors on the left hind limb remained unirradiated. The tumors on the right hind limb received 3 Gy of radiation daily for 3 days. For the IHC analyses, the mice were sacrificed at 24 hours after the last treatment, and the tumors were processed using an anti-GRP78 antibody. For the imaging studies, the mice were analyzed immediately after the last therapy was applied. This novel 3-step imaging protocol involves injecting the peptide conjugated to biotin, a 12 hour delay, an intraperitoneal injection of the avidin chase, a 2 hour delay, and a tail-vein injection of streptavidin conjugated to Alexa Fluor 680. Results: IHC analyses revealed that GRP78 is induced in these breast tumors in response to sunitinib, sorafenib, pazopanib and 5-FU ± radiation, and not to untreated controls. We found that the GIRLRG peptide discriminately binds to breast tumors treated with sunitinib, sorafenib, pazopanib, and 5-FU ± radiation. These findings were statistically significant (p &lt; 0.005). We discovered a strong binding of GIRLRG to treated breast tumors at 24 hours and not to untreated tumors, which was consistent with the results obtained with IHC analyses. Conclusions: We found that GRP78 is induced in these breast tumors in response to systemic treatment, alone or when combined with radiation. The GIRLRG peptide can discriminately bind to breast tumors that have been treated with 5-FU chemotherapy, tyrosine kinase inhibitors, or radiation. Since 5-FU is used clinically as capecitabine to treat metastatic breast cancer patients, our ultimate goal would be to conjugate this targeting peptide to a drug delivery system in order to increase chemotherapy efficacy to metastatic tumors while reducing systemic toxicities.

Citation Format: Jose T. Thaiparambil, Jing Wen, Samuel K. Patton, Roberto Diaz. GRP78 is induced and can be targeted after systemic therapy in breast cancer. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 1395. doi:10.1158/1538-7445.AM2013-1395