We used two different types of chemical UPR inducers, the hypoglycemia-mimicking agent 2-deoxy-D-glucose (2DG) and the axis) sorted by cluster analysis, displayed with 7 samples (axis). control.(TIF) pone.0045845.s003.tif (53K) GUID:?7A02FC51-5813-4EB0-891E-EBB643134D69 Figure S4: Cytotoxicity of single-treatment compound C in unstressed and 2DG-stressed 786-O cells. MTT assay. 786-O cells were treated with compound C for 24 h under normal or 10 mM 2DG stress conditions. Results shown are the means SD of quadruplicate determinations.(TIF) pone.0045845.s004.tif (31K) GUID:?8D115EEA-1F4B-4F38-802C-DF6599C7FF2E Table S1: Summary of 8 samples using microarray analysis. (PDF) pone.0045845.s005.pdf (28K) GUID:?891ED88A-E9C3-4932-BD84-0F99067FE225 Table S2: Expression level of upregulated genes (148 probe sets) in the Glucose Deprivation signature. (PDF) pone.0045845.s006.pdf (31K) GUID:?6F5F74CC-CF40-49B4-9FA3-93F1FCF5EEBF Table S3: Expression level of downregulated genes (98 probe sets) in the Glucose Deprivation signature. (PDF) pone.0045845.s007.pdf (23K) GUID:?7AEE577D-17B2-42D1-A8FA-C2322B2D2AEA Text S1: Supplementary Methods. (PDF) pone.0045845.s008.pdf (81K) GUID:?D75419C5-57B6-4E07-8989-23C30A591BB7 Abstract Inhibiting the unfolded protein response (UPR) can be a therapeutic approach, especially for targeting the tumor microenvironment. Here, we show that compound FR194738 free base C (also known as dorsomorphin), a small-molecule inhibitor of AMP-activated protein kinase (AMPK) and bone morphogenetic protein (BMP) signaling, inhibit the UPR-induced transcription program depending on the glucose deprivation conditions. We found that compound C prevented UPR marker glucose-regulated protein 78 (GRP78) accumulation and exerted enhanced cytotoxicity during glucose deprivation. Gene expression profiling, together with biochemical analysis, revealed that compound C had a unique mode of action to suppress the transcriptional activation of UPR-targeted genes, as compared with the classic UPR inhibitors versipelostatin and biguanides. Surprisingly, the UPR-inhibiting activity of compound C was not associated with either AMPK or BMP signaling inhibition. We further found that combination treatments FR194738 free base of compound C and the classic UPR inhibitors resulted in synergistic cell death with UPR suppression during glucose deprivation. Our findings demonstrate that compound C could be a unique tool for developing a UPR-targeted antitumor therapy. Introduction Glucose deprivation is a common feature of the solid tumor microenvironment and is caused by a combination of FR194738 free base the poorly formed tumor vasculature, uncontrolled proliferation and abnormal energy metabolism of cancer cells. As does hypoxia, glucose deprivation leads to the abnormal accumulation of protein within the endoplasmic reticulum (ER), which triggers the activation of the unfolded protein response (UPR) in tumor cells [1], [2]. The UPR in cancer cells plays an important role in their survival under stress conditions and results in tumor malignancies and COL12A1 in antitumor drug resistance, whereas, in the case of intolerable levels of ER stress, the UPR can contribute to eliciting apoptosis [1], [2], [3]. Thus, the UPR is a potential target of antitumor therapy, and the repression or induction of the UPR by drugs may have therapeutic effects against tumors. The UPR consists of three main signaling pathways initiated by ER membrane-localized stress sensors, PKR-like ER kinase (PERK), activating transcription factor 6 (ATF6) and inositol-requiring 1 (IRE1) [1], [3]. PERK induces the transcription factor activating transcription factor 4 (ATF4) through the phosphorylation of eukaryotic translation initiation factor 2 subunit alpha (eIF2), which also transiently leads to attenuation of global translation [4], [5], [6]. ATF6 becomes an active transcription factor by proteolytic cleavage [7], [8], whereas IRE1 mediates the unconventional splicing of X-box binding protein 1 (XBP1) mRNA, thereby converting it to a potent UPR transcriptional activator [9], [10], [11], [12]. These transcription factors lead to coordinated induction of divergent UPR target genes, such as the ER-resident molecular chaperones glucose-regulated protein 78 and 94 (GRP78 and GRP94), for cell survival [13]. We previously reported.