To harness the potential of human pluripotent stem cells (hPSCs), an abundant supply of their progenies is required. Here, hPSC expansion as matrix-independent aggregates in suspension culture was combined with cardiomyogenic differentiation using chemical Wnt pathway modulators. A multiwell screen was scaled up to stirred Erlenmeyer flasks and subsequently to tank bioreactors, applying controlled feeding strategies (batch and cyclic perfusion). Cardiomyogenesis was sensitive to the GSK3 inhibitor CHIR99021 concentration, whereas the aggregate size was no prevailing factor across culture platforms. However, in bioreactors, the pattern of aggregate formation in the expansion phase dominated subsequent differentiation. Global profiling revealed a culture-dependent expression of BMP agonists/antagonists, suggesting their decisive role in cell-fate determination. Furthermore, metallothionein was discovered as a potentially stress-related marker in hPSCs. In 100 ml bioreactors, the production of 40 million predominantly ventricular-like cardiomyocytes (up to 85% purity) was enabled that were directly applicable to bioartificial cardiac tissue formation.

Radiotherapy (RT) is still one of the standard cancer therapies, with up to two third of all cancer patients with solid tumors being irradiated in the course of their disease. The aim of using ionizing radiation in fractionated treatment schedules was always to achieve local tumor control by inducing DNA damage which can be repaired by surrounding normal tissue but leads to cell death in tumor cells. Meanwhile, it is known that RT also has immunological effects reshaping the tumor microenvironment. Nevertheless, RT alone often fails to elicit potent antitumor immune responses as these effects can be immunostimulatory as well as immunosuppressive. Here, we discuss how immunotherapies can be exploited in combined therapies to boost RT-induced antitumor immune responses or to counteract preexisting and RT-mediated immunosuppression to improve local and systemic tumor control. Furthermore, we highlight some parameters of radioimmunotherapies (RITs) which are under investigation for potential optimizations and how RIT approaches are tested in first phases II and III trials. Finally, we discuss how RT might affect normal and cancer stem cells.

In addition to locally control the tumor, hypofractionated radiotherapy (RT) aims particularly to activate immune cells in the RT-modified microenvironment. Therefore we examined, whether hypofractionated RT can activate dendritic cells (DCs), induce immune cell infiltration in tumors and how the chronology of immune cell migration into tumors occurs to gain knowledge for future definition of radiation breaks and inclusion of immunotherapy. Colorectal cancer treatments offer only limited survival benefit and immunobiological principles for additional therapies need to be explored with pre-clinical models. The impact of hypofractionated RT on CT26 colon cancer tumor cell death, migration of DCs towards supernatants (SN) and activation of DCs by SN was analyzed. The subcutaneous tumor of a BALB/c-CT26 mouse model was locally irradiated with 2x5Gy, the tumor volume was monitored and the infiltration of immune cells in the tumor was determined by flow cytometry daily. Hypofractionated RT induced a mixture of apoptotic and necrotic CT26 cells which is known to be in particular immunogenic. DCs that migrated towards supernatants of CT26 cells only upregulated the activation markers CD80 and CD86 when in contact with SN of irradiated tumor cells. After hypofractionated RT the tumor outgrowth was significantly retarded and in the irradiated tumors an increased infiltration of macrophages (CD11bhigh/F4-80+) and DCs (MHC-II+), but only between day 5 and 10 after the first irradiation takes place. While CD4+ T cells migrated into non-irradiated and irradiated tumors, CD8+ T cells were only found in tumors that had been irradiated and they were highly increased at day 8 after the first irradiation. Myeloid-derived suppressor cells and regulatory T cells show regular turnover in irradiated and non-irradiated tumors. Tumor cell-specific anti-IgM antibodies were enhanced in the serum of animals with irradiated tumors. We conclude that hypofractionated RT suffices to activate DCs and to induce infiltration of innate and adaptive immune cells into solid colorectal tumors. However, the presence of immune cells in the tumor which are beneficial for anti-tumor immune responses is timely restricted. These findings should be considered when innovative multimodal tumor treatment protocols of distinct RT with immune therapies are designed and clinically implemented.

Mild hyperthermia (HT) (41.5 °C for 30–60 min) has been shown in various cell culture systems, preclinical and clinical models to be a very potent radiosensitiser. Recent research suggests that local HT application in combination with standard tumour therapies such as radiotherapy (RT) and/or chemotherapy may not only improve local tumour control but also lead to systemic and immune mediated anti-tumour responses. Melanoma has been proven to be rather radioresistant and mostly only the addition of immunotherapy is capable of inducing beneficial anti-melanoma responses. This work therefore focuses on whether HT increases the immunogenic potential of B16-F10 mouse melanoma cells in combination with RT. The in vitro experiments revealed that combination of RT with HT resulted in an increased percentage of apoptotic and necrotic melanoma cells and an increased release of the danger signal heat shock protein 70 (Hsp70) and high mobility group box protein 1 (HMGB1). HT alone was also capable of inducing this release. We set up local irradiation and heating procedures of B16-F10 tumour-bearing C57/BL6 mice and revealed that the tumour growth of tumours treated with RT plus HT was significantly retarded compared to tumours treated only with RT. This combined treatment generated a beneficial tumour microenvironment by enhancing the infiltration of CD11c + /MHCII + /CD86+ dendritic cells, CD8+ T cells, and NK cells, and decreasing that of regulatory T cells and myeloid-derived suppressor cells. We conclude that HT in combination with RT has an immune-stimulating potential that might result in anti-tumour immunity.