261. Gain-of-Function Effect Augments Therapeutic Efficacy of CpG-STAT3 Anti-Sense Oligonucleotide Against Castration-Resistant Prostate Cancers
Authors: Dayson Moreira, Xingli Zhao, Haejung Won, Priyanka Duttagupta, Summanta Pal, Marcin Kortylewski
Journal: Molecular Therapy (2016)

The treatment for castration-resistant prostate cancers (CRPC) remains a clinical challenge. STAT3 expression level correlates with poor prognosis in prostate cancer patients and resistance to hormone therapy. In addition to oncogenic role of STAT3 in cancer cells, it is also commonly activated in the prostate tumor microenvironment. Therefore, STAT3 is a promising target targeting STAT3 is expected to generate dual effects; anti-tumor activity and anti-immunosuppression in CRPC. Targeting transcriptions factors for cancer therapy has not been successful due to the lack of enzymatic activity. Recent advances in oligonucleotide-based therapy using anti-sense oligonucleotide (ASO) targeting STAT3 had shown promising results in phase I clinical trials in various human cancers, including liver metastasis. However, the limitation fo these reagents remained targeting We previously demonstrated that Toll-like Receptor 9 (TLR9) is up-regulated in prostate cancer stem-like cells in the late-stage prostate cancer. STAT3 signaling is activated in TLR9+ prostate cancer cells and tumor associated immune cells. Here, we describe a new molecule that enables specific delivery of STAT3 ASO to TLR9+ tumor and immune by linking STAT3 ASO to CpG ODN, TLR9 ligand. CpG-STAT3 ASO was quickly and efficiently internalized by human and mouse prostate cancer cell lines showing cytoplasmic localization after 15 minutes after incubation in vitro. Human and mouse TLR9+ immune cells; DCs, Macrophages and B cells were able to efficiently uptake CpG-STAT3 ASO in vitro. In a biodistribution experiment, CpG-STAT3 ASOCy3 was effectively internalized by TLR9+ myeloid cells in various organs including lymph node and bone marrow. Treatment with CpG-STAT3 ASO conjugates in vitro decreased the expression of STAT3 in TLR9+ human (DU145 and LNCaP-TLR9) and mouse (RM1 and RM9) prostate cancer cells more efficiently than unconjugated STAT3 ASO. Notably, CpG-STAT3 ASO showed significantly augmented cytotoxicity in prostate cancer cells compared to STAT3 ASO. In a preliminary anti-tumor efficacy study using syngeneic mouse model, repeated intra-tumoral injections of CpG-STAT3 ASO reduced the tumor growth of RM9. In contrast, the inhibitory effect of STAT3 ASO was transient leading to tumor relapse after the initial treatments. Noteworthy, CpG-STAT3 ASO reduced tumor growth and STAT3 expression in the untreated tumors in the contra-lateral site. Moreover, CpG-STAT3 ASO, but not the unconjugated STAT3 ASO, reduced activation of STAT3 in infiltrated myeloid-derived suppressor cells (MDSC) (CD11b+Gr1+) in both tumors and the percentage of T reg (CD4+FoxP3+) cells in the tumor draining lymph nodes. These results implicate that only when combined with concomitant TLR9 stimulation STAT3 inhibition can systemically overcome tumor immune tolerance. Overall, these results suggest that CpG-STAT3 ASO conjugate holds a great promise for an anti-cancer therapy of CRPC by simultaneously targeting STAT3 signaling in TLR9+ prostate cancer cells and tumor-associated immune cells.