I. A. Naqvi1, R. Gunaratne1, J. McDade1, D. Rouse1, J. Lee1, B. Sullenger1, R. White2 1Duke University Medical Center,Surgery,Durham, NC, USA 2University Of California – San Diego,Surgery,San Diego, CA, USA
Introduction: Pancreatic cancer (PC) has the poorest prognosis of all major cancers with a 5-year survival of less than 5%. Cell-free DNA (cfDNA) has been well-validated as a biomarker that correlates with tumor burden. More recently, a role for cfDNA in tumor progression has also been identified. The latter occurs through activation of toll-like receptors (TLRs) in both tumor cells and the host environment, which can upregulate pro-metastatic signaling pathways. In addition, tumor-derived extracellular vesicles (EVs) such as microparticles and exosomes have also been implicated in promoting metastasis by activating pro-invasive pathways in tumor cells and pre-conditioning secondary sites for metastatic establishment. Our laboratory has previously shown that cationic polymers can neutralize cfDNA and abrogate inflammation in disease models of autoimmunity and infection. We investigated the ability of the cationic polymer, PAMAM-G3, to bind and inhibit cfDNA and EVs and thereby inhibit tumor invasion in vitro and metastasis in vivo.
Methods: Transwell-Matrigel invasion assays were performed using multiple PC cell lines. Cells were plated in the upper chamber with no serum +/- cfDNA, EVs, or PAMAM-G3. The bottom chamber was plated with complete media as a chemo-attractant. Cells treated with cfDNA, EVs, or PAMAM-G3 were also analyzed for activation of pro-inflammatory pathways (NFkB, MAPK, etc.). The in vivo model was performed in C57B6 mice that were injected with bioluminescent murine PC cells into their spleens and followed for liver metastases with bioluminescent imaging. Experimental mice were treated biweekly with intraperitoneal PAMAM-G3 (20 mg/kg) or saline starting 48 hours after pancreatic tumor cell implantation.
Results: PAMAM-G3 significantly inhibited in vitro invasion of pancreatic cancer cell lines in response to cancer patient serum-derived cfDNA and EVs. Moreover, PAMAM-G3 (20 mg/kg) treatment led to a significant reduction in liver metastasis without affecting primary tumor growth in vivo. Serum derived from saline treated mice induced higher levels of invasion than serum from polymer treated mice and exogenous polymer blocked this effect (Figure). (**** = p<0.0001, *** = p<0.001, ** = p<0.01, NS = not significant by t-test)
Conclusion: Cationic polymers such as PAMAM-G3 may represent a novel class of therapeutics to combat pancreatic cancer metastasis. These polymers can bind and neutralize cfDNA and tumor-derived EVs, thereby blocking activation of pro-inflammatory pathways within tumor cells, and reducing tumor invasion and metastasis. Cationic polymers will be tested in other pre-clinical models of cancer and in dosing studies for potential progression to clinical translation.
