ABSTRACT Despite modern advances, ovarian cancer remains one of the most common causes of cancer-related death of women in the US. Due to lack of an effective screening test, ovarian cancer typically presents at an advanced stage. Current treatment of ovarian cancer is primarily limited to surgery and chemotherapy, and the five-year survival rate is below 50%. New treatments are urgently needed to help patients suffering from this deadly disease. A unique feature of ovarian cancer is that more than 80% of patients express a high serum level of CA- 125. Bioinformatic analysis shows that CA-125 mRNA is also highly expressed in gynecological cancer cells, with the highest in ovarian cancer cells, but not in most other cancer cells or in normal cells. CA-125 has been regarded as an ideal and unique target for ovarian cancer treatment; however, targeting CA-125 protein for ovarian cancer treatment has never been successful. Specific transcriptional activation of MUC16 (the gene encoding CA-125) in ovarian cancer cells is poorly defined, and targeting specific MUC16 transactivation for ovarian cancer treatment has never been attempted. Nevertheless, our overall objective is to determine the feasibility of targeting the MUC16 gene specific transactivation by developing a conditionally replicative adenovirus (CRAd) that can only replicate in ovarian cancer cells expressing CA-125 and to gauge the ability of virus-infected cancer cells to induce a protective anti- cancer immune response that can be harnessed for ovarian cancer treatment. We hypothesize that using a CRAd with the MUC16 transactivation sequence to control an essential gene for virus replication is an innovative and practical way to target the specific MUC16 gene transactivation for ovarian cancer treatment. We have compelling preliminary data to support our hypothesis. We expect to accomplish our overall objective by pursuing the following Specific Aims: Aim 1: Develop potent CRAds that specifically replicate in ovarian cancer cells expressing CA-125. Aim 2: Evaluate the anti-cancer activity of CRAds in immunodeficient and immunocompetent mice. Aim 3: Select a potent CRAd and assess the oncolytic activity in primary ovarian cancer cells collected from patients. We anticipate that the strategy of constructing a CRAd dependent on MUC16 transactivation for replication will be feasible and effective for ovarian cancer treatment. Successful completion of the proposed studies will yield a potent targeted agent that not only specifically replicates in and lyses ovarian cancer cells, but also activates a protective anti-cancer immune response. Better definition of MUC16 transcriptional elements will also lay the groundwork for more detailed mechanistic investigations into CA-125 regulation in ovarian cancers, which may reveal new targets to explore for ovarian cancer treatment.