Performance of Pyridylthiourea-Polyethylenimine Polyplex for siRNA-Mediated Liver Cancer Therapy in Cell Monolayer, Spheroid, and Tumor Xenograft Models.

Medical application of siRNAs relies on methods for delivering nucleic acids into the cytosol. Synthetic carriers, which assemble with nucleic acids into delivery systems, show promises for cancer therapy but efficiency remains to be improved. In here, the effectiveness of pyridylthiourea-polyethylenimine (πPEI), a siRNA carrier that favors both polyplex disassembly and endosome rupture upon sensing the acidic endosomal environment, in 3 experimental models of hepatocellular cancer is tested. The πPEI-assisted delivery of a siRNA targeting the polo-like kinase 1 into Huh-7 monolayer produces a 90% cell death via a demonstrated RNA interference mechanism. Incubation of polyplex with Huh-7 spheroids leads to siRNA delivery into the superficial first cell layer and a 60% reduction in spheroid growth compared to untreated controls. Administration of polyplexes into mice bearing subcutaneous implanted Huh-7Luc tumors results in a reduced tumor progression, similar to the one observed in the spheroid model. Altogether, these results support the in vivo use of synthetic and dedicated polymers for increasing siRNA-mediated gene knockdown, and their clinical promise in cancer therapeutics.

Sticky siRNAs targeting survivin and cyclin B1 exert an antitumoral effect on melanoma subcutaneous xenografts and lung metastases.

BACKGROUND: Melanoma represents one of the most aggressive and therapeutically challenging malignancies as it often gives rise to metastases and develops resistance to classical chemotherapeutic agents. Although diverse therapies have been generated, no major improvement of the patient prognosis has been noticed. One promising alternative to the conventional therapeutic approaches currently available is the inactivation of proteins essential for survival and/or progression of melanomas by means of RNA interference. Survivin and cyclin B1, both involved in cell survival and proliferation and frequently deregulated in human cancers, are good candidate target genes for siRNA mediated therapeutics. METHODS: We used our newly developed sticky siRNA-based technology delivered with linear polyethyleneimine (PEI) to inhibit the expression of survivin and cyclin B1 both in vitro and in vivo, and addressed the effect of this inhibition on B16-F10 murine melanoma tumor development. RESULTS: We confirm that survivin and cyclin B1 downregulation through a RNA interference mechanism induces a blockage of the cell cycle as well as impaired proliferation of B16-F10 cells in vitro. Most importantly, PEI-mediated systemic delivery of sticky siRNAs against survivin and cyclin B1 efficiently blocks growth of established subcutaneaous B16-F10 tumors as well as formation and dissemination of melanoma lung metastases. In addition, we highlight that inhibition of survivin expression increases the effect of doxorubicin on lung B16-F10 metastasis growth inhibition. CONCLUSION: PEI-mediated delivery of sticky siRNAs targeting genes involved in tumor progression such as survivin and cyclin B1, either alone or in combination with chemotherapeutic drugs, represents a promising strategy for melanoma treatment.

Systemic delivery of sticky siRNAs targeting the cell cycle for lung tumor metastasis inhibition.

RNA interference allows the design of new inhibitors that target deregulated pathways in cancer. However systemic delivery of siRNA for the treatment of solid tumors still remains an issue. In our study, in order to suppress the progression of lung cancer metastasis in mice, we developed sticky siRNA (ssiRNA) to inhibit survivin and cyclin B1, two candidates involved in cell survival and proliferation. We exploited the linear polyethylenimine (PEI) as potent non-viral carrier to efficiently deliver our inhibitors. As a proof of concept, we have chosen a very aggressive mammary adenocarcinoma model (TSA-Luc cells), which forms lung metastases upon systemic cell injection. We confirmed in vitro, that the ssiRNAs delivered with PEI are not only able to inhibit our target genes at the mRNA and protein levels, but are also able to block the cell cycle and cell proliferation through a mechanism of RNA interference. More importantly, we showed in vivo by luciferase dosage, bioimaging and tissue section, an inhibition of lung tumor metastases after systemic delivery of cyclin B1 and survivin ssiRNA complexed with PEI. Alternating treatment with cisplatin and ssiRNA/PEI showed an additive effect between the two anticancer drugs on lung tumor inhibition leading to a significant increase in animal survival. Moreover a promising window between activity (IC50) and toxicity (LD50), essential for therapeutic application, was observed. Our data show that systemic delivery of ssiRNA/PEI complexes targeting the cell cycle is a valuable strategy for the treatment of lung tumor metastasis and that it can be combined with chemotherapy.