Abstract
The treatment of breast cancer by radiotherapy can be complemented by hyperthermia. Little is known about how the immune phenotype of tumor cells is changed thereby, also in terms of a dependence on the heating method. We developed a sterile closed-loop system, using either a warm-water bath or a microwave at 2.45 GHz to examine the impact of ex vivo hyperthermia on cell death, the release of HSP70, and the expression of immune checkpoint molecules (ICMs) on MCF-7 and MDA-MB-231 breast cancer cells by multicolor flow cytometry and ELISA. Heating was performed between 39 and 44 °C. Numerical process simulations identified temperature distributions. Additionally, irradiation with 2 × 5 Gy or 5 × 2 Gy was applied. We observed a release of HSP70 after hyperthermia at all examined temperatures and independently of the heating method, but microwave heating was more effective in cell killing, and microwave heating with and without radiotherapy increased subsequent HSP70 concentrations. Adding hyperthermia to radiotherapy, dynamically or individually, affected the expression of the ICM PD-L1, PD-L2, HVEM, ICOS-L, CD137-L, OX40-L, CD27-L, and EGFR on breast cancer cells. Well-characterized pre-clinical heating systems are mandatory to screen the immune phenotype of tumor cells in clinically relevant settings to define immune matrices for therapy adaption.
Highlights
Tumors have developed a multitude of mechanisms to evade immune surveillance [1].Breast cancer is a heterogeneous disease and represents the second most common cause of cancer deaths among women
Focus was given to the analyses of the inactivation efficiency by monitoring the percentage of apoptotic and necrotic tumor cells, and on the analyses of the immunogenic potential of treated tumor cells through the detection of the danger signal heat shock protein 70 (HSP70) and the surface expression of several immune activatory and immune suppressive checkpoint molecules
We identified in comparative tests on closed-loop cell media treatment that HT does match to RT because of its radiosensitising properties, and because of its influence on the immune phenotype of breast cancer cells
Summary
Tumors have developed a multitude of mechanisms to evade immune surveillance [1].Breast cancer is a heterogeneous disease and represents the second most common cause of cancer deaths among women. Only modest response rates to immunotherapies such as immune checkpoint inhibitors (ICIs) are observed. Just recently, it has been shown that a combination of the ICI pembrolizumab and radiotherapy (RT) is safe and demonstrates encouraging activity in patients with poor-prognosis, metastatic, triple-negative breast cancer [3]. Besides the activation of the immune systems following radiation of cancer cells by, e.g., stimulating the release of danger signals, immune suppressive pathways, such as the increased expression of immune checkpoint molecules (ICMs) are observed [4]. A big challenge in innovative radiation oncology is the identification of patients who do respond well to distinct combinations of radiation protocols with selected immunotherapies [5]. Knowledge about the immune phenotype of cancer cells per se and following treatment with, e.g., RT is mandatory
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