Introduction

Cancer remains the major cause of death worldwide and the incidence is rising. According to the World Health Organization (WHO) cancer statistics 2012 in Europe, the estimated risk of getting cancer before age 75 are 21.6% for women and 29.7% for men, while rates for cancer mortality before age 75, are 9.2% and 15.6% for women and men, respectively (Ferlay et al. 2014).

Corresponding incidence numbers in Finland are 23.1% for women and 29.1% for men, while mortality rates are considerably lower at 7.6% and 10.7% for women and men, respectively.

Figures are estimated in the absence of other causes of death. The mortality difference is partially explained by the fact that more curable cancer types, prostate and breast cancer, are more common in Finland (Figure 1), but it also reflects the socioeconomical advantages in Finland:

functional health care system, resources for early diagnosis, effective cancer treatments that are based on the latest medical research, and continuously growing repertoire of treatment options.

Worldwide, there were 14.1 million new cancer cases in 2012, and the incidence is expected to rise with over 20 million annual new cases expected by 2025 (Ferlay et al. 2014), all this despite the improvements in cancer prevention. For the first time in history, cancer now causes more deaths, altogether 8.2 million in 2012, than any other particular disease, bypassing even ischaemic heart disease, stroke, and infectious diseases (WHO Global Health Observatory Data Repository, 2012).

Meanwhile, treatment of cancer has taken some major advances in the developed countries.

Unfortunately this progress is yet largely unreachable by the low- and middle-income countries. As seen in Figure 1, three of the top cancer types in Finland, prostate, breast and colon cancer, are already mostly curable in majority of the cases. If comparing historically, this is very much owing to the progress in modern cancer research, since the 5-year survival rates of prostate and breast cancer in Finland in the 1960s were around 30% and 55%, respectively, after which both have increased to around 90% (Pukkala et al. 2011).

Many medical advances account for this progress. Besides earlier cancer diagnosis allowing radical treatments at a less aggressive local stage, also conventional curative therapies have improved owing to novel surgical techniques, effective combinations of chemotherapeutic drugs, and targeted optimally fractionated radiotherapy. Nevertheless, yet disappointing outcomes are seen e.g. with regards to lung, pancreatic and ovarian cancer (Figure 1), and similarly, with advanced metastatic disease of any type. This represents the dilemma in oncology that deals with hundreds, if not thousands, of different genetic disorders of various origins, commonly referred to as

“cancer”. Therefore, it is not expected that there is a magic bullet, a miracle cure for all cancer types, but instead novel modalities together with advances in conventional therapies are gradually increasing our tool box. Combinations of different tools can be then utilized to achieve more cures.

Select tumor types, subtypes, or patients first seem to respond to certain (combinatorial) therapies, which are then taken forward into clinical trial testing in order to determine whether

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the treatment increases survival rates as compared to standard therapy. Eventually, a new form of standard therapy may be assigned, which is then further developed, optimized and revised to improve cure rates and reduce possible adverse reactions. Gradually, along with the progress done in both basic and clinical cancer research, the emotionally and socially challenging historical concept of cancer as a lethal, life-stopping disease could change. To achieve this, however, much work remains to be done and novel treatment options are needed.

Figure 1. Cancer incidence and mortality in Finland. A) Estimated age-standardized rates (ASR[W]) of cancer incidence (blue) and mortality (red) in Finland in 2012, including both genders. Rates represent the number of new cases or deaths per 100,000 persons per year, which are weighted for a standard age structure. Modified from: (Ferlay et al. 2014). B) Development of actual cancer incidence and mortality rates in Finland during 1953–2009. Modified from: (Pukkala et al. 2011).

Biologically, cancer refers to a large group of genetic diseases, which may originate from virtually any cell type and organ in the body. All cancers arise as a result of numerous alterations occurring in the DNA sequence of cells. Consequently, some of the proteins encoded by these cells are differentially expressed or mutated, giving growth advantage and the ability to proliferate in defiance of physiological control. A localized, non-invasive tumor is called benign, whereas malignant tumor refers to a cancer which has acquired the capability to invade or disseminate from the site of primary tumor to other tissues. Metastases spawned by malignant tumors are the main cause of cancer-related deaths in humans (Mehlen and Puisieux 2006).

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Oncolytic viruses are one promising treatment modality for cancer, which has recently gained attention as the first positive clinical phase III results in the Western countries were announced in 2013 (Andtbacka et al. 2013). In addition, two oncolytic adenovirus products have already been approved and are in clinical use in China since 2003 and 2005 (Guo and Xin 2006). Oncolytic adenoviruses are genetically modified to target and replicate only in cancer cells, and thus represent a form of targeted cancer gene therapy. As adenoviruses are originally human pathogens, multiple host immune mechanisms, and also counteractive circuits in the virus, have emerged during evolution. Therefore, it is not surprising that besides replicating in and lysing the infected cancer cells, oncolytic adenoviruses also induce prominent immune reactions at the tumor site. Furthermore, oncolytic viruses can be genetically modified to express immune-stimulating transgenes, which further boost immune responses, directed not only against the virus but also to the host tumor cells (Lichty et al. 2014). Hence, oncolytic virus field has naturally moved towards immunotherapy, aimed at stimulating patient’s own immune system against the mutated altered self, cancer, in order to achieve long-lasting antitumor responses.

Nevertheless, as experimental virotherapy has been around since the mid-19th century, and only now the first approved cancer therapy applications are emerging, it is obvious that obstacles have been encountered. Oncolytic virotherapy is appearing safe approach with over 1,000 cancer gene therapy trials carried out and more than 5,000 cancer patients treated without treatment-related deaths or major limiting toxicity (Ginn et al. 2013). Challenges have lied in the lack of efficacy in clinical trials. This partially reflects the lack of optimal preclinical models to test efficacy, because human adenoviruses do not properly replicate in tissues of other species, forcing researchers to use xenogeneic animal models, i.e. human tumor xenografts in immunodeficient mice, which feature fundamental differences in tumor architecture and impaired immunity. Syrian hamsters have been proposed as a model to circumvent this limitation, but have been found only semi-permissive for replication of human adenovirus (Thomas et al. 2006, Bramante et al. 2014) and represent largely uncharacterized immune system. Therefore, besides developing preclinical testing and more suitable models, reporting and learning from available clinical data is of particular importance.

Both preclinical and clinical evidence suggests that efficacy can be improved, even synergistically, by combining oncolytic immunotherapy with conventional treatment modalities such as chemotherapy and radiotherapy. In preclinical part of this thesis we study combinatorial effects of oncolytic adenoviruses together with radiotherapy and certain chemotherapeutic drugs, and provide mechanistic rationale and show that improved antitumor efficacy can be achieved. In addition, we study the acquired resistance mechanisms against oncolytic adenovirus in ovarian tumors, and reveal relevant pathways, potential tumor marker and targets, which could be utilized in developing countermeasures. In the clinical part, we study altogether 238 patient treatments with oncolytic adenoviruses given in the context of an Advanced Therapy Access Program (ATAP) for patients with metastatic solid tumors progressing after conventional treatments. We demonstrate safety of the approach, and report objective signs of treatment efficacy and antitumor immune responses. In particular, we focus on patients treated, as first-in-humans, with an attractive combination of oncolytic immunotherapy and low-dose chemotherapy that was found synergistic and immunogenic preclinically. Finally, we report finding of a novel serum biomarker that is prognostic and tentatively predicts responsiveness to oncolytic immunotherapy with adenoviruses. Our findings set the stage for testing the combinations and biomarkers in clinical trials, which may ultimately have an impact on cancer therapy in practice.

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