Abstract 3630: Immunoengineering of tumor associated macrophages using targeted, siRNA delivering nanoparticles
Authors: Ortega Ryan , Barham Whitney , Tikhomirov Oleg , Sharman Kavya , Yull Fiona , Giorgio Todd 
Published: May 2015

Abstract
Tumor associated macrophages (TAMs) can modify the tumor microenvironment to create a pro-tumor niche. Dysregulation of NF-κB signaling is implicated in creating a pro-tumor phenotype in TAMs. NF-κB signaling consists of a classical pathway and a less understood alternative pathway. Suppression of NF-κB activation in TAMs is predicted to decrease local smoldering inflammation and inhibit the pro-tumor TAM phenotype. However, recent studies have reported anti-tumor macrophage behavior induced by activation of the classical NF-κB pathway. We have successfully used mannosylated polymer nanoparticles (Mn-NP) to deliver siRNA for RNAi of NF-κB proteins in bone marrow derived macrophages (BMDMs) and mouse TAMs in vivo. Mannose serves as a macrophage targeting ligand via the mannose receptor (CD206). This work aims to characterize these nanomaterials for targeted delivery of functional nucleic acid sequences into macrophages in vitro and in vivo. These studies will also use the Mn-NP to investigate the effects of specific manipulation of NF-κB activation on macrophage phenotype.
Mn-NPs are micelles formed by self-assembly of triblock polymers with a poly(BMA-co-PAA-co-DMAEMA) core, a DMAEMA siRNA condensing block, and an azide-containing outer block for further functionalization. Alkyne functionalized mannose is then “clicked” on the end of the polymer and presented on the micellar surface. In vitro cell studies carried out with BMDMs from mice that express luciferase as a reporter of total NF-κB activity. Mn-NP delivery of siRNA (50 nM for 24 hrs) against each of several NF-κB pathway proteins confirms efficacious knockdown of total NF-κB activity, comparable to transfection with the commercial agent Lipofectamine. Mn-NPs are also highly biocompatible in vitro and exhibit no significant liver or kidney toxicity in mice at doses of 5 mg/kg. The mannosylated carrier effectively delivers cy3 fluorescently labeled nucleotides to TAMs in multiple model systems, including in primary mammary tumors arising spontaneously in polyoma mice and an implanted ovarian tumor model, . Currently, studies are focused on a potential therapeutic strategy that involves RNAi of an inhibitor of the classical NF-κB pathway, IκBα, in order to strategically activate the classical pathway in TAMs. Quantitative RT-PCR analysis of macrophages treated with Mn-NP formulated with siRNA against IκBα show a decrease in M2 markers, a decrease in smoldering inflammation associated cytokines, and an increase in immune cell recruiting chemokines.
Citation Format: Ryan Ortega, Whitney Barham, Oleg Tikhomirov, Kavya Sharman, Fiona Yull, Todd Giorgio. Immunoengineering of tumor associated macrophages using targeted, siRNA delivering nanoparticles. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 3630. doi:10.1158/1538-7445.AM2014-3630