Abstract 1407A: Imaging the coordinated expression of cell surface receptors in gliomas using a novel multi-functional enzyme reporter complementation complex

Abstract

Abstract Diagnosis of any disease is crucial in the prevention, treatment and reduction of the mortality rate. Tailoring clinical diagnostics and therapeutics to specific forms of cancer and even to the precise aberrant biological events underlying cancer development in a particular patient has focused significant research on molecular signatures. For instance, cell surface receptors important in tumorigenesis are expressed at different levels during the multi-stage process and are routinely used to guide treatment regimes. Mutational events that drive a normal cell to become a cancer cell require the coordinated overexpression of not just one receptor at a time, but rather multiple biomarkers. Many cancers are characterized by an abnormal increase in the activity of epidermal growth factor receptors (EGFR) and transferrin receptors (TfR). Our data of representative human cancer cell lines demonstrate unique, observable expression patterns for the two receptors. Current diagnostic techniques such as medical imaging, tissue biopsy, and bio-analytical assay of body fluids are relatively insensitive and lack the specificity to detect these physiologic changes simultaneously. To develop imaging tools that take advantage of the molecular signature, new technologies must employ a contrast agent or signal-amplifying material conjugated to a targeting agent. By linking a reporter enzyme, such as β-gal, to a targeting moiety, signal-amplification at the molecular level is achieved. We divided β-gal into unique, independent polypeptides that reassemble and complement enzymatic activity in bacteria and in mammalian cells. We created two sets of complementing pairs that individually have no enzymatic activity. However, when brought into close proximity, complementing pairs associate, resulting in detectable enzymatic activity. We then constructed a targeting complex composed of reporter fragment, linker, and targeting moiety. Our studies demonstrate a time course- and dose-dependent uptake in vitro. Further, we were able to simultaneously visualize the two cell surface receptors implicated in gliomas, EGFR and TfR, using complementing pairs of the targeted-reporter fragment complex. In addition, we were able to image orthotopic brain tumor accumulation and localization of the targeted-complexes when a fluorescence reporter was added to the complex, as well as immunohistochemical staining of the β-gal reporter complex ex vivo. After fluorescence imaging localized the β-gal complexes to the brain tumor, we topically applied a bioluminescent β-gal substrate to serial sections of the brain to evaluate the delivery and integrity of enzyme. Robust bioluminescence was captured within the tumor after 3 min, indicating that β-gal maintained its activity. Current experiments are underway to image the enzyme complementation non-invasively. 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 1407A. doi:10.1158/1538-7445.AM2011-1407A