Our results indicated that the gene silencing effects by CpG-siRNA in cultured cells requires high concentrations of the conjugates and are suboptimal relative to in vivo treatment. Work is underway to determine the possible cause(s) of this difference, which might include serum-associated degradation of CpG-siRNAs or reduction of the overall silencing effect in rapidly dividing cell populations. It is possible that in vivo repeated treatments allow accumulative gene silencing effects, and the crosstalk between various cells in the tumor microenvironment could lead to secondary inhibitory effects on Stat3 activity 23 . The halflife of the constructs at present is limited. Although the CpG ODN in the construct is phosphothioated, which should resist serum degradation, the siRNA in the chimeric construct is unmodified and negatively charged. Chemically modifying the siRNA to prolong serum stability and to neutralize the negative charge of the siRNA to facilitate endosomal release may improve the efficacy of TLR agonist-siRNA approach. Although more studies are necessary to further understand the mechanism(s), and to improve CpG-siRNA gene silencing effects, our results raise the possibility to use oligonucleotide TLR agonists for siRNA delivery into tumor-associated myeloid cells and B cells, and possibly certain tumor cells, to inhibit expression of tumor-promoting/immunosuppressive molecules while activating TLR(s) for immune activation.