Novel Combination Oncolytic Adenoviral Gene Therapy Armed with Dm-dNK and CD40L for Breast Cancer

Background:Both Drosophila melanogaster deoxyribonucleoside kinase (Dm-dNK) suicide gene therapy and exogenous CD40 ligand (CD40L)-CD40 interaction in cancer via conditionally replicating adenovirus can selectively kill tumors without damaging normal tissues.Objective:To further improve the cancer killing effect, we investigated the therapeutic effect of combined cancer gene therapy based on a selective oncolytic adenovirus vector containing Dm-dNK suicide gene and exogenous CD40L on breast carcinoma cells in vitro and in vivo.Methods:A series of conditionally replicating adenoviruses using adenovirus vector P74 were generated: P74-dNK, P74-CD40L (expressing Dm-dNK or CD40L respectively), and P74-dNK-CD40L (expressing combined Dm-dNK and CD40L). Breast cancer cell lines (MDA-MB-231, MCF-7) and non-tumor cell line (MRC5) were treated with adenovirus and cytotoxicity determined by MTT assay, and apoptosis assessed by flow cytometry after 72h. We also assessed in vivo cell killing efficiency using a mouse xenograft model with MDA-MB-231 cells.Results and Discussion:Co-expression of Dm-dNK and CD40L reduced cell proliferation of MDAMB- 231 or MCF7 cancer cells, and induced more apoptosis in TERT and CD40 positive cancer cells, but not normal MRC5 cells. Significant reduction in tumor volume was also seen in combined treatment arms as compared to any single treatment.Conclusion:Our data suggest enhanced, selective tumor cell killing using combined gene therapy with conditionally replicating adenovirus containing Dm-dNK suicide gene and exogenous CD40 ligation (CD40L-CD40).

Cisplatin Synergistically Enhances Antitumor Potency of Conditionally Replicating Adenovirus via p53 Dependent or Independent Pathways in Human Lung Carcinoma

Cisplatin is ranked as one of the most powerful and commonly prescribed anti-tumor chemotherapeutic agents which improve survival in many solid tumors including non-small cell lung cancer. However, the treatment of advanced lung cancer is restricted due to chemotherapy resistance. Here, we developed and investigated survivin promoter regulating conditionally replicating adenovirus (CRAd) for its anti-tumor potential alone or in combination with cisplatin in two lung cancer cells, H23, H2126, and their resistant cells, H23/CPR, H2126/CPR. To measure the expression of genes which regulate resistance, adenoviral transduction, metastasis, and apoptosis in cancer cells, RT-PCR and Western blotting were performed. The anti-tumor efficacy of the treatments was evaluated through flow cytometry, MTT and transwell assays. This study demonstrated that co-treatment with cisplatin and CRAd exerts synergistic anti-tumor effects on chemotherapy sensitive lung cancer cells and monotherapy of CRAd could be a practical approach to deal with chemotherapy resistance. Combined treatment induced stronger apoptosis by suppressing the anti-apoptotic molecule Bcl-2, and reversed epithelial to mesenchymal transition. In conclusion, cisplatin synergistically increased the tumor-killing of CRAd by (1) increasing CRAd transduction via enhanced CAR expression and (2) increasing p53 dependent or independent apoptosis of lung cancer cell lines. Also, CRAd alone proved to be a very efficient anti-tumor agent in cancer cells resistant to cisplatin owing to upregulated CAR levels. In an exciting outcome, we have revealed novel therapeutic opportunities to exploit intrinsic and acquired resistance to enhance the therapeutic index of anti-tumor treatment in lung cancer.