Abstract 4433: Nanoconjugate mediated inhibition of EGFR expression of triple negative breast cancer
Authors: Inoue Satoshi , Patil Rameshwar , Portilla Jose , Ding Hui , Konda Bindu , Espinoza Andres , Phillips H. Westley , Elramsisy Adam , Holler Eggehard , Black Keith L. , Ljubimova Julia 
Published: June 2012

Abstract
Objective: We synthesized novel nanoconjugate based on a polymalic acid (PMLA) nanoplatform for triple negative breast cancer treatment. Aberrant expression of EGFR correlates with poor prognosis of breast cancer and anti-EGFR therapy has been recognized to play an important role for triple negative breast cancer patients. However, conventional small molecule drugs lack tumor specificity and cause adverse effect on normal cells. To improve anti-cancer and tumor specific targeting effects, we synthesized a new nanoconjugate carrying the monoclonal antibody (mAb) 2C5. This antibody has binding specificity for tumor cells, while PMLA harbors a mouse anti-transferrin receptor antibody (TfR) and a antisense oligonucleotide to EGFR (AON) allowed us to inhibit breast cancer growth.
Materials and Methods: The nanoconjugates are: (1) PMLA-AON with 2C5 and TfR for tumor endothelial and cancer cell targeting, (2) PMLA-AON with 2C5, and (3) only 2C5 without AON. (4) PBS and (5) PMLA were used as a control. Anti-mouse TfR antibody mediated polymer transcytosis through tumor vascular endothelial cells, and 2C5 targeted tumor cell specifically. EGFR overexpressing (MDA-MB-468, SKBR-3) breast cancer cells were used to evaluate drug effect after treatments in vitro. MDA-MB-468 triple negative breast cancer cells were inoculated subcutaneously in nude mice. Drugs were injected intravenously and tumor sizes were measured. Intravenous injections with a nanoconjugate labeled with Alexa Fluor 680 were done for imaging analysis. Drug accumulation was analyzed by Xenogen IVIS 200 and subsequent confocal microscopy. EGFR, phosphorylated Akt and total Akt expressions were detected by Western blotting using in vivo tumor samples undergoing different drug treatments as described above.
Results: In vitro Western blot showed that PMLA-AON with targeting antibody inhibited EGFR synthesis more than naked AON. In vivo imaging and confocal microscopy revealed increased drug accumulation with targeting mAbs. Nanoconjugates carrying both mAb and AON significantly inhibited breast tumor growth in vivo as compared to controls (P&lt;0.03). The highest inhibition of tumor growth was observed in mice treated with the leading nanoconjugate variant (1) PMLA-AON with 2C5 and TfR, (P=0.03 vs. controls; P=0.03 vs. nanoconjugate variant (2)). Leading nanoconjugate (1) also showed stronger inhibition of EGFR expression and Akt phosphorylation than variant (2) and controls on protein level expression.
Summary: The newly designed nanoconjugate with covalently attached monoclonal antibody (2C5 and TfR) delivered EGFR inhibitors specifically into the cancer cells by passing host endothelial system and tumor cell membrane. Systemic treatment of triple negative human breast tumor-bearing nude mice with a nanoconjugate showed significant anti-tumor effects in vivo.
Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 4433. doi:10.1158/1538-7445.AM2011-4433