The Study Of The Mechanism Of 2-hexyl-4-pentylenic Acid On The Inhibition Of Lung Metastasis In Breast Cancer By Regulating Macrophages Phenotype

BackgroundBreast cancer and its lung metastasis have become a major public health problem that seriously endangers women’s health.Advanced breast cancer will metastasize to the distal tissue,of which lung metastasis is one of the most common types of breast cancer metastasis.The survival rate of most breast cancer patients with lung metastasis is significantly lower,so finding treatment measures that can inhibit breast cancer lung metastasis is a hot spot in this field.Breast tumor tissue is infiltrated with a large number of macrophages,usually exhibiting a M2 phenotype that promotes tumor growth,angiogenesis,and metastasis.Macrophages have strong plasticity and their M1 phenotype has anti-tumor effects.Therefore,seeking safe and effective drugs to induce macrophage polarization towards M1 type in tumor tissue is crucial for inhibiting lung metastasis of breast tumors.2-Hexyl-4-pentyne acid(HPTA),a derivative of valproic acid,is a novel HDACi that can inhibit the growth of breast tumors.However,it has not been reported whether HPTA can inhibit lung metastasis of breast tumors by promoting macrophage infiltration in tumor tissue and inducing polarization towards M1 type.Therefore,this study aims to establish a spontaneous lung metastasis model of in situ breast tumors in mice to investigate whether HPTA can inhibit the mechanism of breast tumor lung metastasis by inducing macrophage polarization towards M1 in tumor tissue.The aim is to provide a reliable theoretical basis and experimental support for the clinical treatment of breast tumors and their lung metastasis.MethodsAt the animal level,by establishing a 4T1-Luc mouse model of spontaneous lung metastasis of in situ breast tumors,the effects of HPTA on in situ tumor growth and lung metastasis tumor formation,as well as possible related immune mechanisms,were explored.H&E staining was used to observe the pathological morphology of in situ tumors and lung metastatic tumors in mice;Observing the lung metastasis of breast tumors using Bouin’s staining and lung biopsy imaging;Immunohistochemistry(IHC)was used to detect the proliferation and angiogenesis of tumor cells in situ and lung metastatic tumor tissues,as well as the infiltration of myeloid derived cells,macrophages,and CD8+T lymphocytes;Real time fluorescence quantitative PCR(RT qPRC)was used to detect M1 and M2 macrophage functional markers and related T cell chemokines in situ and lung metastatic tumor tissues;Protein blotting assay(WB)was used to detect functional biomarkers of macrophages in situ and lung metastatic tumor tissues.At the cellular level,the scratch test confirmed that HPTA inhibited the metastasis of breast cancer cells by inducing macrophages to polarize toward M1 type.Results1.HPTA can inhibit the growth and angiogenesis of in situ breast tumors,while promoting the formation of vacuolar necrotic structures in tumor tissue2.HPTA can reduce the metastasis of in situ breast tumors to the lungs,while inhibiting cell proliferation and angiogenesis in lung metastatic tumor tissues,and prolonging the survival period of lung metastatic mice3.After HPTA treatment,the positive expression of F4/80 and CD11b in mouse in situ tumor and lung metastatic tumor tissues significantly increased,indicating that myeloid derived macrophages may be involved in HPTA mediated inhibition of mouse breast tumor growth and lung metastasis.4.HPTA can inhibit the expression of M2 phenotype markers such as CD206,IL-10,Arg-1,etc.in macrophages of in situ and lung metastatic tumor tissues,and promote M1 phenotype markers such as CD80 and TNF-α The expression of equality5.HPTA can up regulate the expression of CXCL9/10/11,the chemokine of T cells,and promote the infiltration of CD8+T lymphocytes and the expression of granzyme B,the functional marker of CD8+ T lymphocytes in mice with in situ breast tumors and lung metastases6.At the cellular level,HPTA can induce macrophages to polarize toward M1 type,thereby inhibiting the metastasis of breast cancer cellsConclusions1.Through the mouse breast cancer spontaneous lung metastasis model,HPTA can not only inhibit the growth of primary breast tumors,but also inhibit the metastasis of tumors to the lungs and prolong the survival of mice.2.HPTA can promote the polarization of macrophages to M1-type in primary tumor and lung metastatic tumor,and the T cell chemokine CXCL9/10/11 secreted by M1-type macrophages may participate in the recruitment of CD8+T lymphocytes in tumor.3.At the cellular level,HPTA could inhibit the metastasis of breast cancer by regulating the polarization of macrophages to M1-type.