PMID- 30563569 OWN - NLM STAT- MEDLINE DCOM- 20191014 LR - 20200309 IS - 2051-1426 (Electronic) IS - 2051-1426 (Linking) VI - 6 IP - 1 DP - 2018 Dec 18 TI - Tumor cell-released autophagosomes (TRAPs) promote immunosuppression through induction of M2-like macrophages with increased expression of PD-L1. PG - 151 LID - 10.1186/s40425-018-0452-5 [doi] LID - 151 AB - BACKGROUND: Tumor-associated macrophages (TAMs) facilitate tumor progression via establishment of an immunosuppressive tumor microenvironment (TME). However, it is poorly understood how tumor cells could functionally modulate TAMs. Our previous work indicated that tumor cell-released autophagosomes (TRAPs), a type of LC3-II(+) double-membrane extracellular vesicles (EVs) was sufficient to suppress anti-tumor immune responses by inducing IL-10-producing B cells and immune suppressive neutrophils. Here, we hypothesized that TRAPs may participate in regulating macrophage polarization. METHODS: TRAPs isolated from multiple murine tumor cell lines and pleural effusions or ascites of cancer patients were incubated with bone marrow-derived macrophages (BMDMs) and monocytes, respectively. Cellular phenotypes were examined by flow cytometry, ELISA and quantitative PCR. TRAPs treated BMDMs were tested for the ability to suppress T-cell proliferation in vitro, and for promotion of tumor growth in vivo. Transwell chamber and neutralization antibodies were added to ascertain the inhibitory molecules expressed on BMDMs exposed to TRAPs. Knockout mice were used to identify the receptors responsible for TRAPs-induced BMDMs polarization and the signaling mechanism was examined by western blot. Autophagy-deficient tumors were profiled for phenotypic changes of TAMs and IFN-gamma secretion of T cells by flow cytometry. The phenotype of monocytes from pleural effusions or ascites of cancer patients was assessed by flow cytometry. RESULTS: TRAPs converted macrophages into an immunosuppressive M2-like phenotype characterized by the expression of PD-L1 and IL-10. These macrophages inhibited the proliferation of both CD4(+) and CD8(+) T cells in vitro, and promoted tumor growth mainly through PD-L1 in vivo. TRAPs-induced macrophage polarization was dependent on TLR4-mediated MyD88-p38-STAT3 signaling. In vivo studies indicated that disruption of autophagosome formation in B16F10 cells by silencing the autophagy gene Beclin1 resulted in a remarkable delay in tumor growth, which was associated with reduced autophagosome secretion, TAMs reprogramming and enhanced T cell activation. Moreover, the levels of LC3B(+) EVs appeared to correlate significantly with up-regulation of PD-L1 and IL-10 in matched monocytes from effusions or ascites of cancer patients, and TRAPs isolated from these samples could also polarize monocytes to an M2-like phenotype with increased expression of PD-L1, CD163 and IL-10, decreased expression of HLA-DR, and T cell-suppressive function. CONCLUSIONS: These findings suggest the TRAPs-PD-L1 axis as a major driver of immunosuppression in the TME by eliciting macrophage polarization towards an M2-like phenotype, and highlight the potential novel therapeutic approach of simultaneously targeting autophagy and PD-L1. FAU - Wen, Zhi-Fa AU - Wen ZF AD - Department of Microbiology and Immunology, Medical School of Southeast University, 87 Dingjiaqiao Rd, Nanjing, 210009, People's Republic of China. FAU - Liu, Hongxiang AU - Liu H AD - Department of Microbiology and Immunology, Medical School of Southeast University, 87 Dingjiaqiao Rd, Nanjing, 210009, People's Republic of China. FAU - Gao, Rong AU - Gao R AD - Department of Microbiology and Immunology, Medical School of Southeast University, 87 Dingjiaqiao Rd, Nanjing, 210009, People's Republic of China. FAU - Zhou, Meng AU - Zhou M AD - Department of Microbiology and Immunology, Medical School of Southeast University, 87 Dingjiaqiao Rd, Nanjing, 210009, People's Republic of China. FAU - Ma, Jie AU - Ma J AD - Department of Microbiology and Immunology, Medical School of Southeast University, 87 Dingjiaqiao Rd, Nanjing, 210009, People's Republic of China. FAU - Zhang, Yue AU - Zhang Y AD - Department of Microbiology and Immunology, Medical School of Southeast University, 87 Dingjiaqiao Rd, Nanjing, 210009, People's Republic of China. FAU - Zhao, Jinjin AU - Zhao J AD - Department of Microbiology and Immunology, Medical School of Southeast University, 87 Dingjiaqiao Rd, Nanjing, 210009, People's Republic of China. FAU - Chen, Yongqiang AU - Chen Y AD - Department of Microbiology and Immunology, Medical School of Southeast University, 87 Dingjiaqiao Rd, Nanjing, 210009, People's Republic of China. FAU - Zhang, Tianyu AU - Zhang T AD - Department of Microbiology and Immunology, Medical School of Southeast University, 87 Dingjiaqiao Rd, Nanjing, 210009, People's Republic of China. FAU - Huang, Fang AU - Huang F AD - Department of Microbiology and Immunology, Medical School of Southeast University, 87 Dingjiaqiao Rd, Nanjing, 210009, People's Republic of China. FAU - Pan, Ning AU - Pan N AD - Department of Microbiology and Immunology, Medical School of Southeast University, 87 Dingjiaqiao Rd, Nanjing, 210009, People's Republic of China. FAU - Zhang, Jinping AU - Zhang J AD - Institutes of Biology and Medical Sciences, Soochow University, Suzhou, 215123, People's Republic of China. FAU - Fox, Bernard A AU - Fox BA AD - Robert W. Franz Cancer Research Center, Earle A. Chiles Research Institute, Providence Portland Medical Center, 2N81 North Pavilion, 4805 N.E. Glisan St, Portland, OR, 97213, USA. FAU - Hu, Hong-Ming AU - Hu HM AD - Robert W. Franz Cancer Research Center, Earle A. Chiles Research Institute, Providence Portland Medical Center, 2N81 North Pavilion, 4805 N.E. Glisan St, Portland, OR, 97213, USA. hong-ming.hu@providence.org. FAU - Wang, Li-Xin AU - Wang LX AUID- ORCID: 0000-0002-3480-4491 AD - Department of Microbiology and Immunology, Medical School of Southeast University, 87 Dingjiaqiao Rd, Nanjing, 210009, People's Republic of China. lxwang@seu.edu.cn. LA - eng PT - Journal Article PT - Research Support, Non-U.S. Gov't DEP - 20181218 PL - England TA - J Immunother Cancer JT - Journal for immunotherapy of cancer JID - 101620585 RN - 0 (B7-H1 Antigen) RN - 0 (Biomarkers) RN - 0 (CD274 protein, human) RN - 0 (Myeloid Differentiation Factor 88) RN - 0 (Toll-Like Receptor 4) SB - IM MH - Animals MH - Autophagosomes/*immunology/*metabolism/ultrastructure MH - Autophagy MH - B7-H1 Antigen/genetics/*metabolism MH - Biomarkers MH - Cell Line, Tumor MH - Female MH - Humans MH - *Immune Tolerance MH - Immunophenotyping MH - Lymphocyte Activation/immunology MH - Macrophages/*immunology/*metabolism MH - Mice MH - Models, Biological MH - Myeloid Differentiation Factor 88/metabolism MH - Neoplasms/*immunology/*metabolism/pathology MH - Phenotype MH - T-Lymphocyte Subsets/immunology/metabolism MH - Toll-Like Receptor 4/metabolism MH - Tumor Microenvironment/immunology PMC - PMC6299637 OTO - NOTNLM OT - IL-10 OT - MyD88 OT - PD-L1 OT - T cell OT - TAMs OT - Tumor cell-released autophagosomes (TRAPs) OT - Tumor microenvironment COIS- ETHICS APPROVAL: All animal experiments were approved by the Animal Care and Use Committee of Southeast University. All human experiments were approved by the Ethics Committee for Human Studies of Southeast University and performed under protocol 2016ZDKYSB112. Informed consent was obtained from all patients. CONSENT FOR PUBLICATION: All authors provide their consent for publication of the manuscript. COMPETING INTERESTS: Hong-Ming Hu and Bernard A. Fox are co-founders of UbiVac. The other authors declare no competing interests. PUBLISHER'S NOTE: Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations. EDAT- 2018/12/20 06:00 MHDA- 2019/10/15 06:00 PMCR- 2018/12/18 CRDT- 2018/12/20 06:00 PHST- 2018/07/20 00:00 [received] PHST- 2018/11/16 00:00 [accepted] PHST- 2018/12/20 06:00 [entrez] PHST- 2018/12/20 06:00 [pubmed] PHST- 2019/10/15 06:00 [medline] PHST- 2018/12/18 00:00 [pmc-release] AID - 10.1186/s40425-018-0452-5 [pii] AID - 452 [pii] AID - 10.1186/s40425-018-0452-5 [doi] PST - epublish SO - J Immunother Cancer. 2018 Dec 18;6(1):151. doi: 10.1186/s40425-018-0452-5.