Therapeutic effect of local hyperthermia treatment combined with anti-CD200 immunotherapy in EMT6 breast cancer

Abstract

Breast cancer is the leading cancer-causing death in women worldwide. Conventional cancer treatments have different therapeutic effects and commonly associated with side effects. Combinatorial therapies are typically used to overcome such limitation and reduce the likelihood of cancer recurrence. Hyperthermia (HT) is a less invasive cancer treatment ideal to treat superficial tumour and has shown promising outcomes in combination with the established immunotherapy. In EMT6 breast tumour, an elevated expression of CD200 has been associated with cancer progression. It was reported that the blocking of CD200/CD200R signalling resulted in an enhanced anti-tumour response. In light of this, the present study aimed to investigate the therapeutic effects of local HT in combination with anti-CD200 blockade therapy. Local HT protocol was established prior to the application of combined treatment. Breast tumours were induced by inoculating EMT6 cells subcutaneously at the right flank of Balb/c mice. On day seven post-inoculation, tumours were subjected to local HT treatment by near infrared radiation for three days. Tumour progression and mice survival were monitored and the influence on the immune response was evaluated. Results demonstrated that HT treatment reduced tumour progression and increased the median survival of tumour-bearing mice. Immunohistochemical analysis revealed a large area of necrotized tumour and there was an increase of Hsp70 expression observed around the treated tumour. This observation was accompanied with a significant reduction of proliferating cells when compared to the untreated tumour. Flow cytometry analysis of the lymph node showed an increase of dendritic cells activation and infiltration in treated mice. A higher level of IFN-γ, a pro-inflammatory cytokine but lower IL-10, an anti-inflammatory cytokine, were detected in blood of treated mice, as compared to untreated mice. In another experiment, mice were intraperitoneally injected with anti-CD200 antibody for blockade therapy in addition to HT. Following combined treatment, it was demonstrated that tumour progression was further delayed and the mice survival were greatly improved. In addition, an increase in the number of activated CD8 T cells were observed in the draining lymph nodes of the mice, along with infiltration of T cells, NK cells and B cells into the tumour. In contrast, tumour-infiltrated regulatory T cells and myeloid-derived suppressor cells were largely diminished from the tumour. Taken together, the present findings reveal the potential of combining local HT and anti-CD200 blockade in improving immunological response against breast tumour.