Abstract 1135: Dextran based nanoparticles for immunotheranostics of prostate cancer

Abstract

Abstract Introduction: Targeted nanoparticles (NPs) containing imaging reporters provide exciting options for cancer-specific delivery of siRNA to downregulate specific pathways in theranostic strategies. We recently developed a biodegradable modified dextran NP to deliver effectively siRNA and downregulate the target in vivo (1). Here we have developed a prostate specific membrane antigen (PSMA)-targeted dextran NP to deliver siRNA to downregulate PD-L1 in PSMA expressing prostate cancer (PCa) cells for immunotheranostics. In proof-of-principle studies, the NP was decorated with an optical imaging reporter to allow noninvasive detection in vivo, and evaluated in PC3 human PCa cells genetically engineered to overexpress PSMA (PC3-PIP) and in non-PSMA expressing PC3-Flu cells. Since castration-resistant PCa cells express PSMA, the ability to down regulate PD-L1 specifically may prevent these cancer cells from evading immune surveillance, resulting in their destruction by the immune system. Methods: For mRNA and protein analysis, PSMA overexpressing PC3-PIP cells and low PSMA expressing PC3-Flu cells were treated with dextran NPs with PD-L1 siRNA or PBS. 6h later, IFNγ (10 ng/mL) or PBS was added to the medium. Cells were collected, and proteins and mRNA extracted, 24h after NP/PBS treatment. Total RNA was isolated, and cDNA prepared following standard protocols, with q-RT-PCR performed for PD-L1 expression. Western blots were probed with anti-PD-L1, and anti-PSMA antibodies. For in vivo studies, PC3-PIP and PC3-Flu tumors were inoculated bilaterally in the flanks of male SCID mice. Once tumors reached ~200 mm3, dextran NPs siRNA with PSMA binding peptide were injected intravenously. Dosage and frequency of IFNγ injection in vivo was established, and bio distribution studies performed. Tumor samples were harvested and processed for mRNA and protein to detect PD-L1 expression. Results and Conclusion: Specificity of the PSMA-targeted dextran NP delivery was confirmed by PSMA-dependent retention of the NPs following fluorescent imaging of PC3-PIP and Flu cells. We next assessed the ability of the dextran NPs carrying PD-L1 siRNA to down regulate PD-L1 expression in PC3-PIP and PC3-Flu cells. Studies were performed with or without pre-activation of the cells with IFNγ. q-RT-PCR and western blots analysis confirmed a clear decrease of PD-L1 after NP treatment in PC3-PIP and PC3-Flu cells. Effective NP delivery was confirmed in tumors with optical imaging, both in vivo and ex vivo in excised organs. Ongoing studies are characterizing PD-L1 downregulation in tumors in vivo following treatment with PSMA-targeted dextran NPs delivering PD-L1 siRNA.