Abstract 3439: Tissue tranglutaminase (TG2) targeting by multifunctional field responsive gold nanoparticles

Abstract

Abstract Background: Ovarian cancer (OC) spreads by direct exfoliation into the peritoneal cavity, bypassing vascular mechanisms. Our previous studies demonstrated that intraperitoneal (IP) metastasis is critically dependent on TG2, an enzyme that catalyzes protein post-translational modifications and crosslinking. In this study we assessed TG2 expression in OC, measured effects of TG2 overexpression on OC cell proliferation and tumorigenicity, and designed a novel method for cancer-specific TG2 targeting. As in-vivo delivery of siRNAs is inefficient and nonspecific, we engineered folate-coated gold nanorods (GNR) that deliver and release TG2 siRNAs into tumor cells by photothermal activation, and used small animal models to evaluate the efficacy of siRNA delivery. Materials & Methods: Immunohistochemistry (IHC) evaluated TG2 expression in 58 OC specimens. Correlation with histological subtype, grade, and stage was performed by Kruskal-Wallis test. Human full length TG2 and mutants lacking enzymatic activity (C277S), GTPase activity (R580A), or both (TG1-140) were stably expressed in OV90 cells. Cell proliferation was measured by MTT and clonogenic assays. Tumorigenicity and IP metastasis were assessed in orthotopic xenografts. GNRs were synthesized by seed mediated growth, cleansed by treatment with polystyrenesulfonate, then functionalized with a mixture of thiolated siRNA and folate-conjugated PEG for targeted TG2 knockdown in tumor cells overexpressing the folate receptor (FR). Localization of FR-targeted GNRs in orthotopic ovarian tumors and metastatic colonies in the peritoneum was assessed ex vivo by confocal microscopy, two-photon luminescence (TPL), and fluorescence imaging. Results: TG2 expression was noted in 84% of OC specimens, of which 71% had > 2+ expression. TG2 expression levels correlated with histological subtype (p=0.005), and with high grade (p=0.005), but not with surgical stage. Forced expression of TG2 or of one of the three mutants did not affect OV90 cell proliferation in vitro or tumor volume in vivo. Mice injected with TG2-OV90 cells developed milliary peritoneal metastases (8/8) compared to mice implanted with control cells (2/9, p=0.002). TG2 knockdown via siRNA yielded >70% reduction in TG2 expression level in vitro. The uptake of folate-labeled GNRs into SKOV3 cells was confirmed in vitro by confocal microscopy and TPL. FR-targeted GNR delivery in vivo was most notable in peritoneal metastases, primary ovarian tumor, and liver. Little or no uptake was noted in other normal tissues, suggesting that this modality achieves cancer-specific delivery. Conclusions: TG2 is overexpressed in OC and its overexpression increases peritoneal metastasis. GNR uptake and delivery to ovarian tumors is facilitated by FR targeting. Ongoing studies are assessing the effects of GNRs carrying TG2 siRNA on OC metastasis in an orthotopic xenograft model. 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 3439. doi:1538-7445.AM2012-3439