Silencing of gene expression by small interfering RNAs (siRNAs) is rapidly becoming a powerful tool for genetic analysis and represents a potential strategy for therapeutic product development. However, there are no reports of systemic delivery for siRNAs toward treatment of bone-metastatic cancer. Accordingly, we report here that i.v. injection of GL3 luciferase siRNA complexed with atelocollagen showed effective reduction of luciferase expression from bone-metastatic prostate tumor cells developed in mouse thorax, jaws, and͞or legs. We also show that the siRNA͞ atelocollagen complex can be efficiently delivered to tumors 24 h after injection and can exist intact at least for 3 days. Furthermore, atelocollagen-mediated systemic administration of siRNAs such as enhancer of zeste homolog 2 and phosphoinositide 3-hydroxykinase p110-␣-subunit, which were selected as candidate targets for inhibition of bone metastasis, resulted in an efficient inhibition of metastatic tumor growth in bone tissues. In addition, upregulation of serum IL-12 and IFN-␣ levels was not associated with the in vivo administration of the siRNA͞atelocollagen complex. Thus, for treatment of bone metastasis of prostate cancer, an atelocollagen-mediated systemic delivery method could be a reliable and safe approach to the achievement of maximal function of siRNA.bone metastasis ͉ prostate cancer R NA interference (RNAi) induced by small interfering RNA (siRNA) has recently emerged as a powerful technique that is capable of suppressing expression of individual genes with a high degree of specificity (1). The technique has been used for studies of gene function in vivo, primarily in mice. The first demonstration of RNAi-mediated repression in an adult animal showed effective repression of a luciferase reporter gene after hydrodynamic transfection of siRNA expression plasmids into mouse liver (2, 3). Subsequent studies have delivered siRNA by various methods, including viral vector-mediated delivery (4, 5) and lipid-based delivery (6, 7). A more recent study showed that chemically modified siRNAs can silence an endogenous gene after i.v. injection in mice (8). These findings provide hope for using RNAi technology in disease control.Many studies have used siRNAs as an experimental tool to dissect the cellular pathways that lead to uncontrolled cell proliferation and cancer. To develop siRNAs for cancer therapy, several researchers have investigated them in animal models (9-13). However, reports of RNAi-delivery strategies for bonemetastatic cancer are very limited. For example, in advanced prostate cancer, the sites most frequently affected by metastasis are the bones and regional lymph nodes. Patients with these metastases suffer pain and low limb edema, making it extremely important to explore avenues of treating such bone metastases.We previously demonstrated the efficacy of atelocollagen for delivery of nucleotides, such as plasmid DNA and antisense oligonucleotides, in vitro and in vivo (14)(15)(16)(17)(18)(19). Recently, we also reported that a...

Treatment of patients with advanced sarcoma remains challenging due to lack of effective medicine, with the development of novel drugs being of keen interest. A pan-PI3K inhibitor, ZSTK474, has been evaluated in clinical trials against a range of advanced solid tumors, with clinical benefit shown in sarcoma patients. In the present study, we developed a panel of 14 human sarcoma cell lines and investigated the antitumor effect of 24 anticancer agents including ZSTK474, other PI3K inhibitors, and those clinically used for sarcoma treatment. ZSTK474 exhibited a similar antiproliferative profile to other PI3K inhibitors but was clearly different from the other drugs examined. Indeed, ZSTK474 inhibited PI3K-downstream pathways, in parallel to growth inhibition, in all cell lines examined, showing proof-of-concept of PI3K inhibition. In addition, ZSTK474 induced apoptosis selectively in Ewing's sarcoma (RD-ES and A673), alveolar rhabdomyosarcoma (SJCRH30) and synovial sarcoma (SYO-1, Aska-SS and Yamato-SS) cell lines, all of which harbor chromosomal translocation and resulting oncogenic fusion genes, EWSR1-FLI1, PAX3-FOXO1 and SS18-SSX, respectively. Finally, animal experiments confirmed the antitumor activity of ZSTK474 in vivo, with superior efficacy observed in translocation-positive cells. These results suggest that ZSTK474 could be a promising drug candidate for treating sarcomas, especially those harboring chromosomal translocation.

Drug resistance often critically limits the efficacy of molecular targeted drugs. Although pharmacological inhibition of phosphatidylinositol 3-kinase (PI3K) is an attractive therapeutic strategy for cancer therapy, molecular determinants for efficacy of PI3K inhibitors (PI3Kis) remain unclear. We previously identified that overexpression of insulin-like growth factor 1 receptor (IGF1R) contributed to the development of drug resistance after long-term exposure to PI3Kis. In this study, we examined the involvement of basal IGF1R expression in intrinsic resistance of drug-naïve cancer cells to PI3Kis and whether inhibition of IGF1R overcomes the resistance. We found that cancer cells highly expressing IGF1R showed resistance to dephosphorylation of Akt and subsequent antitumor effect by ZSTK474 treatment. Knockdown of IGF1R by siRNAs facilitated the dephosphorylation and enhanced the drug efficacy. These cells expressed tyrosine-phosphorylated insulin receptor substrate 1 at high levels, which was dependent on basal IGF1R expression. In these cells, the efficacy of ZSTK474 in vitro and in vivo was improved by its combination with the IGF1R inhibitor OSI-906. Finally, we found a significant correlation between the basal expression level of IGF1R and the inefficacy of ZSTK474 in an in vivo human cancer panel, as well as in vitro. These results suggest that basal IGF1R expression affects intrinsic resistance of cancer cells to ZSTK474, and IGF1R is a promising target to improve the therapeutic efficacy. The current results provide evidence of combination therapy of PI3Kis with IGF1R inhibitors for treating IGF1R-positive human cancers.