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Sources of Funding: NIH grant 5P50CA058236-15 (to T.L. DeWeese and S.E._x000D_ Lupold), the David H. Koch Charitable Foundation (to S.E. Lupold)_x000D_

Introduction

Radiosensitizing agents can improve ionizing radiation (IR) potency. We have previously reported that an A10-3-DNAPK chimera can deliver radiation-sensitizing short interfering RNA (anti-DNA-PK siRNA) to prostate cancer cells and subcutaneous tumors through the PSMA-targeting RNA aptamer, A10-3. Here we describe the DNA-PK knock-down and radiation-sensitizing effect of the A10-3-DNAPK chimera on a human prostate cancer xenograft model in the mouse tibia, and its evaluation by bioluminescent imaging, immunohistochemistry and micro-CT.

Methods

Five to six-week-old male athymic nude mice were anesthetized by isoflurane, then luciferase expressing human prostate cancer cells (LNCaP-luc) were inoculated into tibias. Tumors were evaluated by IVIS bioluminescent imaging 6 weeks after implantation, then IVIS positive mice were separated into three groups and treated by PBS, A103-ctrl and A103-DNAPK via tail vein injection. For the confirmation of DNAPK knock-down, tibias were harvested 72hrs post injection and evaluated by HE staining and immunohistochemistry. In order to confirm radiation-sensitizing effect, half of the tumors received IR therapy (6 Gy) directly to the tumored leg. Four weeks after IR, the tibias were harvested and tumor burden assessed ex vivo by nanoScan PET/CT and VivoQuant software. The CT value (HU) from these images were applied to automatically assign regions of interest for bone (red, HU 5,000-10,000) and marrow (green, HU <5,000) (Fig 1B). The sum HU were then calculated for each treatment group to estimate tumor burden four weeks after treatment.

Results

Immunohistochemistry demonstrated reduction of DNAPK protein after A103-DNAPK treatment (Fig 1A: 50%, 38% less than PBS, A103-ctrl, respectively). The results of CT indicate significantly reduced tumor volume in A103-DNAPK pre-treated animals, with IR, when compared to animals pre-treated with PBS or A103-Ctrl (Fig 1C: p =0.045 and p=0.026, respectively).

Conclusions

Systemically injected aptamer siDNAPK can induce PSMA specific radiosensitization through DNAPK knock-down in human prostate tumor within mouse tibia. This system could be used to enhance radiation therapy of locally advanced PCa.

Funding

NIH grant 5P50CA058236-15 (to T.L. DeWeese and S.E._x000D_ Lupold), the David H. Koch Charitable Foundation (to S.E. Lupold)_x000D_