利用5-氟尿嘧啶调节肿瘤微环境以增加乳腺癌骨转移中的T细胞浸润

Bone metastases are a frequent complication of advanced breast cancer, against which immune checkpoint inhibitors are not effective. We previously identified that PD-1 + T cells in a mouse model of bone metastases are inactivated, potentially because of the expansion of PD-L1 + myeloid-derived suppressor cells (MDSC) in bones. 5-Fluorouracil (5-FU) can decrease MDSC in thymoma and colon cancer models. Thus, we aim to determine the efficacy of 5-FU in breast cancer bone metastases as well as its potential side effects.Balb/C mice were inoculated in the left cardiac ventricle with 4T1 breast cancer cells to cause bone metastases within 10 days. A treatment with 4 inoculations of 5-FU (50 mg/kg) prevented splenomegaly and the expansion of PMN- and M-MDSC in bones, as assessed by flow cytometry. Treatments with doses of 10 or 25 mg/kg were not as efficient against MDSC. Also, while these lower doses tended to increase T cell infiltration, 50 mg/kg decreased it, suggesting toxicity. Thus, we modulated the number of inoculations at 50 mg/kg. Two and three inoculations decreased the splenomegaly and histological damage, but also the total number of bone marrow cells. Flow cytometry confirmed these treatments decreased PMN-MDSC, while there was only a trend toward a decrease in M-MDSC in bone metastases. More interestingly, the numbers of CD4 + and CD8 + T cells were increased, especially naïve and central memory T cells. On radiographs, the bone destruction caused by cancer cells was also decreased. However, a single inoculation did not cause any long-lasting effects on MDSC and T cells, and the osteolysis area was not decreased. As 5-FU sharply decreased the number of bone marrow cells in bone metastases, we sought to assess potential side effects. Three inoculations of 5-FU in healthy mice decreased neutrophils, monocytes, B cells, and NK cells in bones 3 days after the last inoculation, while T cells were still increased. However, these effects were transient since a week later, the mice had recovered. We evaluated levels of hematopoietic stem cells (HSC) and myeloid and lymphocytic progenitor cells (MPC, LPC). Surprisingly, in these conditions, 5-FU transiently increased the number of HSC and LPC, which could explain the rapid recovery, while MPC remained stable. Cancer treatments can affect bones and increase the risk of fracture, so we assessed the bone 3D microarchitecture using microCT. Two and three inoculations of 5-FU did not cause any changes in the trabecular bone volume, or the thickness or number of the trabeculae. Surprisingly, histomorphometry analysis did not reveal any changes in the number of osteoclasts, the myeloid cells dedicated to bone resorption. Thus, 5-FU could be used to decrease MDSC and increase T cells in bone metastases, turning them into a hot microenvironment to increase the efficacy of immunotherapies in breast cancer patients.