Curcumin Suppresses M2 Macrophage-derived Paclitaxel Chemoresistance through Inhibition of PI3K-AKT/STAT3 Signaling.

Breast cancer is the leading cancer in women worldwide. The development of chemoresistance that leads to recurrence and mortality remains a major concern. M2-type tumor-associated macrophages (TAMs), present in the breast tumor microenvironment, secrete various cytokines and growth factors that induce chemoresistance. Curcumin, isolated from Curcuma longa, is known to sensitize cancer cells and increase the efficacy of standard chemotherapeutic agents. However, the effect of curcumin on the chemoresistancegenerating ability of M2 TAMs is not known. The study aimed to determine whether curcumin could modulate M2 macrophages and suppress their ability to induce resistance to paclitaxel in breast cancer cells. THP-1 cells were differentiated to M2 macrophages using PMA and IL-4/IL-13 in the presence or absence of curcumin in vitro. The effect of the conditioned medium of M2 macrophages on inducing resistance towards paclitaxel in MCF-7 or MDA-MB-231 cells was analyzed by cell proliferation assay, cell cycle analysis, wound healing and transwell migration assays. RT-PCR analysis was used to determine the mRNA expression of anti-inflammatory cytokines in M2 macrophages. The effect of curcumin on TGF-β, pAKT, and pSTAT3 in M2 macrophages was analyzed by western blotting. Our data revealed that the M2 macrophages polarized in the presence of curcumin lacked the ability to generate chemoresistance to paclitaxel in breast cancer cell lines. Transcriptomic analysis revealed the expression of TGF-β to be highest amongst M2 macrophage-secreted cytokines. We observed that purified recombinant TGF-β generated chemoresistance in breast cancer cells. We found that curcumin treatment abrogated the expression of TGF-β in M2 macrophages and suppressed their ability to induce chemoresistance in breast cancer cells. STITCH analysis showed strong interaction between curcumin and AKT/STAT3 pathway. Mechanistically, curcumin inhibited PI3K/AKT/STAT3 signaling in M2 macrophages. Western blot analysis revealed that M2 TAM CM, but not curcumin-treated macrophage CM, activated COX2/NF-κB in breast cancer cells. Our results showed that curcumin reduced the chemoresistance-generating ability of M2 TAMs. The study has revealed a non-cancer cell-autonomous mechanism by which curcumin partly overcomes the chemoresistance of paclitaxel in breast cancer.

Deswal B ，Bagchi U ，Kapoor S 《-》

Inhibition of STAT3 by 2-Methoxyestradiol suppresses M2 polarization and protumoral functions of macrophages in breast cancer.

Breast cancer metastasis remains the leading cause of cancer-related deaths in women worldwide. Infiltration of tumor-associated macrophages (TAMs) in the tumor stroma is known to be correlated with reduced overall survival. The inhibitors of TAMs are sought after for reprogramming the tumor microenvironment. Signal transducer and activator of transcription 3 (STAT3) is well known to contribute in pro-tumoral properties of TAMs. 2-Methoxyestradiol (2ME2), a potent anticancer and antiangiogenic agent, has been in clinical trials for treatment of breast cancer. Here, we investigated the potential of 2ME2 in modulating the pro-tumoral effects of TAMs in breast cancer. THP-1-derived macrophages were polarized to macrophages with or without 2ME2. The effect of 2ME2 on macrophage surface markers and anti-inflammatory genes was determined by Western blotting, flow cytometry, immunofluorescence, qRT‒PCR. The concentration of cytokines secreted by cells was monitored by ELISA. The effect of M2 macrophages on malignant properties of breast cancer cells was determined using colony formation, wound healing, transwell, and gelatin zymography assays. An orthotopic model of breast cancer was used to determine the effect of 2ME2 on macrophage polarization and metastasis in vivo. First, our study found that polarization of monocytes to alternatively activated M2 macrophages is associated with the reorganization of the microtubule cytoskeleton. At lower concentrations, 2ME2 treatment depolymerized microtubules and reduced the expression of CD206 and CD163, suggesting that it inhibits the polarization of macrophages to M2 phenotype. However, the M1 polarization was not significantly affected at these concentrations. Importantly, 2ME2 inhibited the expression of several anti-inflammatory cytokines and growth factors, including CCL18, TGF-β, IL-10, FNT, arginase, CXCL12, MMP9, and VEGF-A, and hindered the metastasis-promoting effects of M2 macrophages. Concurrently, 2ME2 treatment reduced the expression of CD163 in tumors and inhibited lung metastasis in the orthotopic breast cancer model. Mechanistically, 2ME2 treatment reduced the phosphorylation and nuclear translocation of STAT3, an effect which was abrogated by colivelin. Our study presents novel findings on mechanism of 2ME2 from the perspective of its effects on the polarization of the TAMs via the STAT3 signaling in breast cancer. Altogether, the data supports further clinical investigation of 2ME2 and its derivatives as therapeutic agents to modulate the tumor microenvironment and immune response in breast carcinoma.

Deswal B ，Bagchi U ，Santra MK ，Garg M ，Kapoor S ... -  《BMC CANCER》

Taraxacum mongolicum extract inhibited malignant phenotype of triple-negative breast cancer cells in tumor-associated macrophages microenvironment through suppressing IL-10 / STAT3 / PD-L1 signaling pathways.

Triple-negative breast cancer (TNBC) is the most aggressive and the worst prognosis breast cancer with limited treatment options. Taraxacum mongolicum (also called dandelion) is a traditional Chinese medicine has been used to treat mastitis, breast abscess, and hyperplasia of mammary glands since ancient times. In modern pharmacological research, dandelion has been proven with anti-breast cancer activities. We previously reported that dandelion extract could induce apoptosis in TNBC cells. However, its anti-tumor effects and mechanisms in the tumor microenvironment have not yet been elucidated. Tumor-associated macrophages (TAMs) play an important role in regulating the interaction between tumor cells and the immune system. The present study aimed to investigate the effects and mechanisms of dandelion extract on TNBC cells under the microenvironment of TAMs, as well as its influence on the polarization of M2 macrophages. M2 macrophages were induced by phorbol-12-myristate 13-acetate (PMA) and interleukin 4 (IL-4), and verified by flow cytometry, quantitative RT-PCR (qRT-PCR), Western blotting, and ELISA. MDA-MB-231 and MDA-MB-468 TNBC cells were co-cultured with the supernatant of M2 macrophage which providing the TAMs microenvironment. The antitumor activity of dandelion extract in TNBC cells was evaluated by MTT assay. The invasive and migratory capacity of TNBC cells was measured by transwell assays. The expression of protein and gene was assessed by Western blotting and qRT-PCR, respectively. TAMs microenvironment promoted the proliferation, migration, and invasion of TNBC cells. However, dandelion extract inhibited the malignant property of MDA-MB-231 and MDA-MB-468 cells induced by TAMs. Both of TAMs and IL-10 caused STAT3 activation and PD-L1 higher expression, the immunosuppressive molecules in TNBC cells, and this effect can be attenuated by IL-10 neutralizing antibody. Dandelion extract exerted inhibition on STAT3 and PD-L1 in TNBC cells under TAMs microenvironment. Furthermore, in M2 macrophages, dandelion extract remarkably promoted the expression of M1-like marker TNF-α, IL-8, and iNOS, but reduced M2-like marker IL-10, CD206, Arginase-1, and TGF-β. Dandelion extract inhibited the proliferation, migration and invasion of TNBC cells in TAMs microenvironment through suppressing IL-10/STAT3/PD-L1 immunosuppressive signaling pathway. Furthermore, dandelion extract promoted the polarization of macrophages from M2 to M1 phenotype. Thus, our results indicated that dandelion may serve as a promising therapeutic strategy for TNBC by modulating tumor immune microenvironment.

Deng XX ，Jiao YN ，Hao HF ，Xue D ，Bai CC ，Han SY ... -  《-》

Increased drug resistance in breast cancer by tumor-associated macrophages through IL-10/STAT3/bcl-2 signaling pathway.

Yang C ，He L ，He P ，Liu Y ，Wang W ，He Y ，Du Y ，Gao F ... -  《-》

Tumor-associated macrophages secrete CC-chemokine ligand 2 and induce tamoxifen resistance by activating PI3K/Akt/mTOR in breast cancer.

Breast cancer is the most prevalent malignancy among women. Although endocrine therapy is effective, the development of endocrine resistance is a major clinical challenge. The tumor microenvironment (TME) promotes tumor malignancy, and tumor-associated macrophages (TAM) within the TME play a crucial role in endocrine resistance. Herein, we aimed to elucidate the relationship between TAM and the endocrine-resistant phenotype of breast cancer. Macrophages were cultured with conditioned medium (CM) from tamoxifen-sensitive (MCF7-S) or -resistant (MCF7-R) MCF7 breast cancer cells. M2 polarization was detected by CD163 immunofluorescence. To determine the effect on endocrine resistance, MCF7 cells were cultured in the supernatant of different TAM, and then treated with tamoxifen. CC-chemokine ligand 2 (CCL2) immunohistochemistry was carried out on pathological sections from 100 patients with invasive estrogen receptor-positive breast cancer. We found that macrophages cultured in the CM of MCF7-S and MCF7-R cells were induced into TAM, with a more obvious M2 polarization in the latter. Tamoxifen resistance was increased by culture in TAM medium. TAM secreted CCL2, which increased endocrine resistance in breast cancer cells through activation of the PI3K/Akt/mTOR signaling pathway. High expression of CCL2 was correlated with infiltration of CD163+macrophages (r = 0.548, P < .001), and patients with high CCL2 expression presented shorter progression-free survival than those with low CCL2 expression (P < .05). We conclude that CCL2 secreted by TAM activates PI3K/Akt/mTOR signaling and promotes an endocrine resistance feedback loop in the TME, suggesting that CCL2 and TAM may be novel therapeutic targets for patients with endocrine-resistant breast cancer.

Li D ，Ji H ，Niu X ，Yin L ，Wang Y ，Gu Y ，Wang J ，Zhou X ，Zhang H ，Zhang Q ... -  《-》