Exploratory Study On The Mechanism Of Bone Metastasis In Lung Cancer And Breast Cancer

Background and purposeBone is a common metastatic site for advanced malignant tumors.The incidence of bone metastasis in lung cancer and breast cancer is 40% and 70%,respectively.Patients with bone metastases often have severe pain,pathological fractures,spinal nerve compression,hypercalcemia and other skeletal-related events(SREs)that severely shorten the overall survival of patients.Lung cancer is the most common malignant tumor with the highest morbidity and mortality.Previous studies have shown that calpyphosine(CAPS1)is closely related to bone metastasis in non-small cell lung cancer.CAPS1 is dependent on cAMP protein kinase A and may act through the PI3K-AKT-mTOR signal pathway.Whether CAPS1 plays a role through the PI3K-AKT-mTOR signaling pathway and its mechanism remains to be further studied.At present,the mechanism of CAPS1 in bone metastasis of lung cancer has not been studied.Breast cancer has the highest morbidity and the second mortality in women.Breast cancer with bone metastasis(BM)or bone marrow metastasis(BMM)seriously endangers women’s health.Abnormal activation of Notch signaling pathway is closely related to the occurrence,development,invasion and metastasis of various malignant tumors,such as breast cancer.The aim of this study was to explore the underlying mechanism of CAPS1 in lung cancer bone metastasis through PI3K-AKT-mTOR signaling pathways and study the characteristics and mechanism of bone metastasis or bone marrow metastasis in breast cancer.It provides a theoretical basis for targeted therapy and precise treatment of patients with bone metastasis,and is of great significance for guiding clinical treatment and improving prognosis.Methods and Materials1.Detect the expression of CAPS1 in OS,Paracancer,NSCLC and BM tissue samples;construct a CAPS1 overexpression vector packaged with lentivirus and screen for CAPS1 overexpression H460 cell line(H460-CAPS1);2.Use WB to detect the expression of CAPS1 in H460-CAPS1 and H460-puro cell lines;conduct cloning formation experiments,CCK-8,apoptosis experiments,wound healing tests,Transwell experiments to detect changes in cell function;3.Use WB to detect the expression of mTOR and CAPS1 in H460-CAPS1 and H460-puro cell lines;Use WB to detect the expression levels of PI3 K,AKT,mTOR and CAPS1 after treated with PI3 K,AKT and mTOR inhibitors,respectively;To determine the possible site of CAPS1 in the PI3K-AKT-mTOR pathway;4.Clinical materials were collected and used to analyze features and survival in BM and BMM groups.5.Bone puncture tissue specimens of 2 patients were collected to detect the DNA sequences by exome sequencing technology in each group.The Kaplan–Meier plotter(http://kmplot.com)and WB were used to analyze and detect the significantly enriched genes in sequencing results.Results1.The expression of CAPS1 in primary tissue and bone metastasis tissue of NSCLC is higher than that of osteosarcoma and adjacent tissues;2.CAPS1 was overexpressed in H460-CAPS1 cells.Clonal formation assay,CCK-8 assay,apoptosis assay,wound healing test and Transwell assay showed that the proliferation,invasion and migration of H460-CAPS1 cells were enhanced;3.The expression of mTOR in H460-CAPS1 cells was higher than that in the control group.Treated H460-CAPS1 and H460-puro cells with PI3 K inhibitors and the expression of CAPS1 decreased in H460-CAPS1 cells.Treated with AKT and mTOR inhibitors,respectively,there was no significant change in the expression level of CAPS1;4.239 BC patients were collected,including 25 cases of BMM and 214 cases of BM.In BMM group,16 cases had pathological reports of BM lesions and the subtype is estrogen receptor(ER)positive of each case(100%).In BM group,121 cases had pathological reports of BM lesions and 88 cases were ER positive(73%).The median survival in BMM group(10.00 ± 3.82 months)was significantly shorter than BM group(25.00 ± 2.89 months)(p<0.05).Both in BMM and BM groups,there was no significant differences in overall survival between the local treatment plus systemic therapy group and systemic therapy-only group;5.Whole exome sequencing results showed abnormalities in various pathways and the Notch signal may play an important role.Conclusion1.CAPS1 can enhance cell proliferation,migration and promote metastasis of non-small cell lung cancer;It is located downstream of PI3 K and may play a role through PI3K-AKT-mTOR pathway.2.Patients with ER positive of BM lesions are more prone to BMM.The BMM was associated with decreased overall survival.Breast cancer with BM or BMM is the result of multiple signals and pathways abnormalities,especially the Notch signaling pathway.