Citation

Therapeutic efficacy of bifunctional siRNA combining TGF-beta1 silencing with RIG-I activation in pancreatic cancer

 

  • Authors: Ellermeier, J., Wei, J., Duewell, P., Hoves, S., Stieg, M. R., Adunka, T., Noerenberg, D., Anders, H. J., Mayr, D., Poeck, H., Hartmann, G., Endres, S., Schnurr, M.
  • Year: 2013
  • Journal: Cancer Res 73 1709-20
  • Applications: in vivo / modified siRNA / in vivo-jetPEI

Method

50µg of ppp-siRNA were complexed with 6µl of in vivo-jetPEI (N/P=6) and injected intravenously through tail vein into mice with orthotopic tumors (Panc02 cell implantation into the pancreas. Injections started on day 10 after cell implantation and were performed twice weekly for 3 weeks.

Abstract

Deregulated TGF-beta signaling in pancreatic cancer promotes tumor growth, invasion, metastasis, and a potent immunosuppressive network. A strategy for disrupting this tumor-promoting pathway is silencing TGF-beta by siRNA. By introducing a triphosphate group at the 5' end of siRNA (ppp-siRNA), gene silencing can be combined with immune activation via the cytosolic helicase retinoic acid-inducible gene I (RIG-I), a ubiquitously expressed receptor recognizing viral RNA. We validated RIG-I as a therapeutic target by showing that activation of RIG-I in pancreatic carcinoma cells induced IRF-3 phosphorylation, production of type I IFN, the chemokine CXCL10, as well as caspase-9-mediated tumor cell apoptosis. Next, we generated a bifunctional ppp-siRNA that combines RIG-I activation with gene silencing of TGF-beta1 (ppp-TGF-beta) and studied its therapeutic efficacy in the orthotopic Panc02 mouse model of pancreatic cancer. Intravenous injection of ppp-TGF-beta reduced systemic and tumor-associated TGF-beta levels. In addition, it induced high levels of type I IFN and CXCL10 in serum and tumor tissue, systemic immune cell activation, and profound tumor cell apoptosis in vivo. Treatment of mice with established tumors with ppp-TGF-beta significantly prolonged survival as compared with ppp-RNA or TGF-beta siRNA alone. Furthermore, we observed the recruitment of activated CD8(+) T cells to the tumor and a reduced frequency of CD11b(+) Gr-1(+) myeloid cells. Therapeutic efficacy was dependent on CD8(+) T cells, whereas natural killer cells were dispensable. In conclusion, combing TGF-beta gene silencing with RIG-I signaling confers potent antitumor efficacy against pancreatic cancer by breaking tumor-induced CD8(+) T cell suppression.

Pubmed