Abstract 1952: Targeting the CCL2/CCR2 chemokine pathway in breast tumors through intratumoral delivery of calcium cross linked TAT peptide: siRNA complexes

Abstract

Abstract Breast tumors exhibit inflammatory responses through recruitment of stromal cells in the primary tumor microenvironment, including fibroblasts and macrophages. Accumulation of these stromal cells correlates with development of invasive breast cancer and poor patient prognosis. These stromal cells express high levels of the inflammatory chemokine CCL2, corresponding to overexpression of its receptor CCR2 in the tumor epithelium. Our laboratory and others have shown that CCL2/CCR2 signaling mediates stromal: tumor interactions to regulate breast cancer cell survival, invasion and metastasis. While the CCL2/CCR2 signaling represents a novel and potentially effective therapeutic target in invasive breast cancer, it is necessary to develop new tools to specifically inhibit CCL2/CCR2 signaling in vivo. siRNA knockdown could specifically target CCL2 or CCR2, but is unstable and toxic when delivered in vivo. The HIV-1 derived TAT peptide penetrates cell plasma membranes at high efficiency and contains nuclear translocation signals to enhance siRNA delivery to the nucleus. Recent studies from our research team have shown that CaCl2 condenses TAT/nucleic acids to nanoparticles facilitating cellular uptake. The goals of these studies are to design TAT nanoparticle formulas to target CCL2 and CCR2 protein expression in breast tumors. Using an in vitro luciferase reporter system, we identified specific N/P ratios and CaCl2 concentrations that would transfect stromal cells and PyVmT mammary tumor cells. TAT/CaCl2 nanocomplexes carrying GFP plasmid or Cy3 labeled siRNAs were then injected into PyVmT mammary tumors. We observed visible GFP expression and Cy3 labeled siRNA uptake in over 50% of the PyVmT mammary tumor at low and high N/P ratios. Normal mammary tissues did not show significant uptake of TAT nanoparticles. Flow cytometry analysis showed that high N/P ratios resulted in efficient TAT/CaCl2/siRNA uptake in breast cancer associated fibroblasts and macrophages. TAT/CaCl2/siRNA nanocomplexes formed at low N/P ratios resulted in efficient siRNA uptake in tumor epithelial cells. Preliminary studies of PyVmT mammary tumors showed a reduction in CCL2 and CCR2 expression upon delivery of TAT/CaCl2 nanoparticles carrying CCL2 or CCR2 siRNAs. These data indicate this nanoparticle strategy represents a novel, feasible approach to target the CCL2/CCR2 signaling in breast cancer, with important implications on therapeutic targeting. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 1952. doi:1538-7445.AM2012-1952