signature=b54bb12e143f5f843cb67edeeea0635b,Abstract B54: Non-malignant cells from metastatic niches ...

摘要：

Metastatic lesions remain the main cause of mortality in breast cancer (BrCa) and their resistance to systemic therapies has been mainly attributed to mutations in BrCa cells, while the non-mutually exclusive role of the metastatic microenvironment has been largely overlooked. To address the role of the local microenvironment of metastatic sites in conferring drug resistance to BrCa cells, we assembled heterotypic in vitro three-dimensional (3D) tissue cultures comprised of estrogen receptor-positive (ER+) BrCa cells and non-malignant cells from organs frequently targeted by metastatic disease. BrCa cells (MCF7, T47D, and ZR75-1) expressing the firefly luciferase gene were grown in extracellular matrices (collagen type I and Matrigel) alone or with nonmalignant cells from the respective metastatic target tissues, and exposed to clinically achievable concentrations of FDA-approved antineoplastic agents widely used for the treatment of BrCa. Interestingly, BrCa cell response to antiestrogens (hydroxytamoxifen (4-OHT), fulvestrant or raloxifene) in 3D conditions was marked by acinar differentiation of tumor spheroids into structures that resemble normal breast epithelium. Co-culture of BrCa cells with immortalized accessory cells from the bone (BMSCs and osteoblasts); brain (astrocytes); liver (hepatocytes); and lung (fibroblasts) in 3D (but not 2D) conditions, inhibited the antiestrogen-induced acinar differentiation and conferred resistance to 4-OHT, raloxifene, and fulvestrant at clinically relevant doses. The results obtained with immortalized human BMSCs were further confirmed in co-cultures of BrCa cells with primary human BMSCs isolated from bone metastatic site. MCF7 cells injected s.c. in nude mice together with BMSCs (heterotypic xenografts) had decreased response to tamoxifen compared with monotypic xenografts (MCF7 alone). Furthermore, immortalized BMSCs or metastatic patient-derived BMSCs induced MCF7 cell resistance to several chemotherapeutic agents including vinca alkaloids (vincristine, vinblastin and vinorelbin) and taxanes (docetaxel, paclitaxel, cabazitaxel). Notably, primary BMSCs from healthy donors and brain astrocytes isolated from cancer-free neurosurgical specimens did not induce BrCa cell resistance to anti-estrogens or chemotherapeutic agents. BMSCs induced in MCF7 cells downregulation of TGFβ2; upregulation of a transcriptional signature enriched for genes associated with high-grade tumors, including genes involved in IGF1R, IL6 and EGFR superfamily members; NF-kappaB and other antiapoptotic pathways; and elevated c-Myc protein levels. The co-cultures of MCF7 cells with BMSCs had increased sensitivity to inhibitors of c -myc, IGF1R, IL-6 and JAK/STAT pathways when compared to the respective mono-cultures. Our results indicate that local nonmalignant cells present in the metastatic niche can function as "accessories" to the tumor, conferring BrCa cell resistance to antiestrogens and chemotherapeutic agents in 3D tissue cultures and xenograft models. Preclinical modeling of the composition and 3D architecture of metastatic lesions can help elucidate the mechanisms of stroma-induced drug resistance and may reveal new therapeutic targets for refractory BrCa patients with metastatic disease.

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