Abstract 3304: Oncolytic adenovirus with low-dose temozolomide induces autophagy and antitumor immune responses preclinically and in cancer patients.
Authors: Liikanen Ilkka , Ahtiainen Laura , Hirvinen Mari L.M , Barmante Simona , Cerullo Vincenzo , Nokisalmi Petri , Hemminki Otto , Diaconu Iulia , Pesonen Sari , Koski Anniina , Kangasniemi Lotta , Pesonen Saila K. , Oksanen Minna , Zhao Fang , Laasonen Leena , Partanen Kaarina , Joensuu Timo , Kanerva Anna , Hemminki Akseli 
Published: August 2014

Tumor cell autophagy appears useful for cancer therapeutics due to immunogenic cell death and possible induction of antitumor immunity. Oncolytic adenoviruses and alkylating chemotherapeutic temozolomide have been shown to induce autophagic cell death preclinically. We studied safety, efficacy and immunological effects of oncolytic adenovirus combined with low-dose pulse of temozolomide. Metronomic low-dose cyclophosphamide was added to treatments to selectively reduce regulatory T-cells.

We assessed the combination of oncolytic adenovirus with low-dose temozolomide and cyclophosphamide in breast and prostate cancer cells in vitro. Immunogenicity of cell death was studied by calreticulin exposure, adenosine triphosphate release, and nuclear protein high-mobility group box-1 (HMGB1) secretion assays. Efficacy was further tested in a prostate cancer xenograft mouse model, and tumors were assessed for autophagy by electron microscopy and LC3-immunohistochemistry. Furthermore, 41 combination treatments were given to 17 chemotherapy-refractory cancer patients in the context of an advanced therapy access program. The primary end-point was safety. In addition, autophagy assays, immunological analyses, and efficacy parameters were studied.

Combination therapy resulted in increased cytotoxicity and immunogenic cell death. Autophagy induction was associated with enhanced tumor-growth inhibition in combination-treated prostate tumors (P&lt;0.01). Patient treatments were well-tolerated with mostly grade 1-2 fever, nausea, lymphopenia and anemia. LC3-stained ascites tumor cells showed increased autophagy post-treatment together with upregulation of autophagic pathways in microarray. 8/15 evaluable cases showed induction of antitumor T-cells in peripheral blood. Interestingly, release of the immunogenic protein HMGB1 into serum increased in 60% of treatments, and seemed to correlate with simultaneous antitumor T-cell responses (P=0.0833). Objective antitumor efficacy was seen in 67% of evaluable treatments, with a trend for increased survival over matched controls treated with virus only (median overall survival 269 versus 170 days).

Low-dose pulse of temozolomide and cyclophosphamide in combination with oncolytic adenovirus increased autophagy, elicited antitumor T-cell activations, and showed promising safety and efficacy. Our results support the hypothesis that oncolysis and chemotherapy mediated autophagy might lead to antitumor immunity. Oncolytic adenovirus together with autophagy-inducing temozolomide is an attractive combination for future clinical studies.

Citation Format: Ilkka Liikanen, Laura Ahtiainen, Mari L.M, Hirvinen, Simona Barmante, Vincenzo Cerullo, Petri Nokisalmi, Otto Hemminki, Iulia Diaconu, Sari Pesonen, Anniina Koski, Lotta Kangasniemi, Saila K. Pesonen, Minna Oksanen, Fang Zhao, Leena Laasonen, Kaarina Partanen, Timo Joensuu, Anna Kanerva, Akseli Hemminki. Oncolytic adenovirus with low-dose temozolomide induces autophagy and antitumor immune responses preclinically and in cancer patients. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 3304. doi:10.1158/1538-7445.AM2013-3304