Comparative effects of proton and photon irradiation on the molecular and cellular profiles of triple-negative breast cancer: the crucial impact of VEGFC on tumor microenvironment remodeling

Metastatic triple-negative breast cancers (TNBC) are among the most aggressive types of breast cancer and are often treated with adjuvant radiotherapy and chemotherapy. Despite initial efficacy, relapses are common, leading to poor prognosis. Understanding the response of tumor microenvironment to radiotherapy is crucial, particularly comparing photon (X) and proton (P) radiotherapy due to proton radiations reduced side effects. MethodsWe investigated the effects of single and multiple X and P irradiations on various cell types within the tumor microenvironment, including vascular and lymphatic endothelial cells, fibroblasts, and TNBC tumor cells. VEGFC, a key factor in lymphatic vessel formation and metastasis, was a primary focus. We used protein arrays to evaluate the effects of irradiation and examined the impact of VEGFC inactivation on the sensitivity to X and P radiation. Additionally, we tested tumor-forming capabilities of irradiated cells and assessed the impact of genetic or therapeutic VEGFC inhibition on TNBC growth. Transcriptomic and proteomic analyses further characterized the differences between X and P tumors, providing deeper insights into their distinct molecular profiles. ResultsBoth X and P irradiations caused a transient increase in VEGFC levels, along with other pro-angiogenic, pro-lymphangiogenic, and pro-fibrotic factors, such as angiopoietin 2, artemin, endostatin, IGFBP2, serpinE1, PDGFA, and DPPIV. Endothelial cells exposed to multiple rounds of radiation showed enhanced proliferation but lost the ability to form pseudo vessels, indicating an endothelial-mesenchymal transition. Tumor cells lacking VEGFC were more sensitive to radiation, and anti-VEGFC antibodies significantly suppressed TNBC cells proliferation, both naive and multi-irradiated. Tumor xenografts formed by multi-irradiated cells grew larger in nude mice, particularly following proton irradiation, while X-irradiated tumors exhibited a more pro-lymphangiogenic phenotype compared to P-irradiated tumors. ConclusionsOur findings show that while P multi-irradiated TNBC cells form larger tumors, X multi-irradiated tumors are more aggressive, with elevated expression of genes linked to angiogenesis, lymphangiogenesis, and endothelial-mesenchymal transition. Targeting VEGFC during photon or proton radiotherapy could reduce metastasis and improve TNBC prognosis.