Abstract B04: Systemic delivery of silver nanoparticles and targeting of the folate receptor alpha for the treatment of triple-negative breast cancer

Triple-negative breast cancer (TNBC) lacks expression of the estrogen and progesterone receptors and does not overexpress the HER-2 receptor. Thus, current targeted therapies are rendered ineffective against this subtype of breast cancer, leaving a gap in treatment options for these patients. The use of nanotechnology has the potential to dramatically enhance the way cancer is diagnosed and treated. However, concerns persist about the toxicity of nanomaterials, which may not degrade, or are only slowly degraded and excreted after long body residence time. For clinical translation to become a reality, the potential risk–benefit balance for these materials must be resolved. We recently identified that silver nanoparticles (AgNPs) have the capacity to act as a single, self-therapeutic agent with a desirable combination of selective cytotoxicity and radiation dose enhancement effects in TNBC cells in vitro and in vivo at doses that that are non-toxic to non-cancerous breast and other cells. This is the first time any nanomaterial has been demonstrated to possess a TNBC specific toxicity profile and provides evidence that a therapeutic window exists for the safe use of AgNPs. Our current studies focus on determining the properties of the nanomaterial that are important to retain or enhance this TNBC selective response. Nanoparticle size and aggregation can have a dramatic effect on the cytotoxic and tumor targeting properties of nanomaterials. We used AgNP preparations with varying diameters (5 nm, 25 nm, 50 nm, 75 nm) to determine the influence of AgNP size on the increased sensitivity of TNBC cell lines to AgNPs. Nanoparticles were characterized by transmission electron microscopy, dynamic light scattering, and nanoparticle tracking analysis. Using this combinatorial analysis, we identify spherical AgNPs preparations that resisted aggregation or dissolution in physiologic solutions for up to one month. We find that the increased sensitivity of TNBC cells as compared to non-cancerous cells was independent of nanoparticle size, and TNBC cell lines (MDA-MB-231, BT-549, SUM-159) are more sensitive to AgNP exposure than luminal A (MCF-7) or non-cancerous breast (MCF-10A, 184B5). We then performed tumor treatment studies in nude mice bearing MDA-MB-231 TNBC tumors in the 4th mammary fat pad. Mice were intravenously injected three times per week with AgNPs (6 mg/kg) or phosphate buffered saline (PBS). Remarkably, AgNP treatment significantly slows tumor TNBC tumor growth in vivo with no apparent systemic toxicity. Targeting has the potential to increase selectivity and uptake of AgNPs leading to increased therapeutic efficacy. Therefore, we functionalized our AgNPs with folic acid (FA) to exploit the folate receptor α (FRA), which is overexpressed in approximately 80% of TNBC. AgNPs were functionalized with a SH-PEG-FA targeting moiety to form FRA targeted AgNPs (FA-AgNPs). Using a modified dot blot probed with an anti-FA antibody, we demonstrate that FA-AgNPs were free of unbound FA and that AgNP bound- FA was available for binding with the FRA. In comparison to non-targeted AgNPs, FA-AgNPs significantly increase the cytotoxic effects of our AgNPs against TNBC in vitro. Furthermore, excess FA decreased the cytotoxicity of FA-AgNPs but had no effect on the cytotoxicity of the non-targeted AgNPs, indicating that the increased cytotoxicity of FA-AgNPs was FRA mediated. In summary, we identified that AgNPs exert significant anti-cancer activity toward TNBC cells in vitro and in vivo, and point to potential vulnerabilities in TNBC cells that could be exploited for the development of new therapeutic agents. Additionally, these AgNPs are suitable for systemic delivery and can be modified with FA to increase the specificity which further decreases the potential for off-target toxicity. Citation Format: Jessica L. Swanner, Cale D. Fahrenholtz, Ravi N. Singh. Systemic delivery of silver nanoparticles and targeting of the folate receptor alpha for the treatment of triple-negative breast cancer. [abstract]. In: Proceedings of the AACR Special Conference on Advances in Breast Cancer Research; Oct 17-20, 2015; Bellevue, WA. Philadelphia (PA): AACR; Mol Cancer Res 2016;14(2_Suppl):Abstract nr B04.