Circulating DNA released from brain tumors in response to conditional cytotoxic/immune stimulatory gene therapy induces activation of TLR9 on pDCs, leading to antitumor immunity

CANDOLFI; YAGIZ; BALASUBRAMANIAN; PUNTEL; KING; MINEHARU; MUHAMMAD; FOULAD; BARNETT; KROEGER; LOWENSTEIN; CASTRO

Orlando, Florida, US

Congreso; American Association for Cancer Research; 2011

Complexes of DNA and proinflammatory proteins are found in serum during autoimmune responses and activate antigen presenting cells, contributing to autoimmune pathogenesis. Although circulating DNA has been extensively evaluated as a biomarker for cancer progression, its role in the onset of anti-tumor immunity remains elusive. Here we aimed to evaluate the role of circulating DNA in the induction of anti-tumor immunity induced by the combination of immune-stimulant Flt3L and conditionally cytotoxic thymidine kinase (TK), which leads to long-term survival when delivered into intracranial brain tumor models using adenoviral vectors (Ad). Treatment of CNS-1 glioblastoma cells with Ad.TK+GCV led to substantial release of DNA in vitro. Similar results were observed in vivo; intratumoral treatment of established intracranial CNS-1 glioblastomas in rats with Ad-TK/GCV+Ad-Flt3L led to increased levels of circulating DNA in serum when compared to saline-treated tumor-bearing rats. Since our previous results suggest that tumor immunity was dependent on the function of activated pDCs, we aimed to evaluate the hypothesis that circulating tumor DNA induces pDC activation. Using specific inhibitors, such as DNAse, micrococcal DNAse, CpG2088 (an inhibitor of TLR9) and cloroquine (an inhibitor of endocytosis, required for intracellular TLR9 activation), we found that circulating DNA from treated rats induced TLR9-dependent activation of pDCs. Taken together, our results suggest that after treatment of brain tumors with Ad.TK+Ad.Flt3L dying tumor cells release DNA to the general circulation, which mediate antitumor immunity by acting as danger-associated molecular patterns, leading to TLR9 signaling and pDC activation.