Kaplan-Meier survival analysis indicated that patients whose tumor expressed a lower level of had a significantly lower overall survival rate (Fig.?1D). Open in a separate window Figure 1. downregulation correlates with disease progression in human PDAC. through regulation of and in autophagic regulation, however, has not been reported. Recent reports have shown that expression of is downregulated in PDAC tissues.31,32 Furthermore, was recently identified as a direct target of exerted a tumor suppression function in PDAC by inducing autophagy-related cell death through targeting the STAT3 pathway. In the current study, we investigated the expression of in different PDAC tissues and its relationship with the prognosis Noscapine of the patients. Further experiments with patient-derived xenograft (PDX) cell systems revealed that functions as a tumor suppressor by inducing autophagy-related cell death through STAT3 signaling pathways. Our findings have unveiled a previously unrecognized mechanism underlying the anticancer effects of against human PDAC. Results downregulation is associated with disease progression in human PDAC Quantification of expression in 92 matched pairs of human PDAC and adjacent normal tissues by quantitative reverse-transcriptase polymerase chain reaction (qRT-PCR) showed that the relative expression levels were significantly lower in PDAC tissues than in the adjacent normal tissues ( 0.01; Fig.?1A). Clinicopathological analyses showed that lower expression in the tumors was significantly correlated with T status and TNM stage (TNM Classification of Malignant Tumors) (Table?S1). The relative expression levels were significantly lower in the advanced tumor than early stage tumor (lower in the III-IV group and T3-T4 group than in the I-II group and T1-T2 group) ( 0.05 and 0.01, respectively; Fig.?1B and ?andC).C). Kaplan-Meier survival NOS3 analysis indicated that patients whose tumor expressed a lower level of had a significantly lower overall survival rate (Fig.?1D). Open in a separate window Figure 1. downregulation correlates with disease progression in human PDAC. (A) Comparison of expression in 92 matched Noscapine pairs of PDAC tissues and corresponding nontumor tissues via qRT-PCR. was used as an internal control. (B) Comparison of expression in stage T1-T2 and stage T3-T4 PDAC tissues. (C) Comparison of expression in stage I-II and stage III-IV PDAC tissues. * 0.05; ** 0.01. (D) Kaplan-Meier curves compared the overall survival rates of PDAC patients whose tumor expressed a low or high level of expression groups. induces cell death in human PDAC cells The effect of on proliferation of PDAC cells was evaluated by immunofluorescent staining for MKI67/Ki-67 in PDX lines MDA-PATC53, MDA-PATC124, and MDA-PATC148. dramatically decreased the MKI67-positive rate in PDAC cells (Fig.?S1A). Consistent with this finding was the observation that significantly impaired the colony-formation capacity of PDAC cells (Fig.?S1B). Assessment of the cell cycle distribution of these cells by propidium iodide (PI) staining and bromodeoxyuridine (BrdU) assay demonstrated that all suppressed proliferation and induced G1 arrest by directly targeting (Fig.?S3) in PDAC cells, which was similar to what we reported in ovarian cancer cells. The 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay was performed to evaluate the impact of on PDAC cell viability. Surprisingly, cell viability significantly decreased in a time-dependent manner in resulted only in blockage of cell viability relative to that of control miRNA-treated cells. Open in a separate window Figure 2. induces cell death in human PDAC cells. (A) MDA-PATC53, MDA-PATC124, and MDA-PATC148 cells were transfected with MIRctrl or for 5 d. Cell viability was measured by the MTT assay every day. (B) After transfection with MIRctrl or 0.01. We performed a trypan blue exclusion assay to confirm whether the induced cell death in a Noscapine time-dependent manner (Fig.?2B). These results reveal that not only inhibited the proliferation of PDAC cells but also induced their death. Cell death caused by is dependent on autophagy To determine the mode of cell death caused by or the apoptosis inducer 5-fluorouracil (5-FU) were exposed to the selective inhibitor of apoptosis N-benzyloxycarbonyl-Val-Ala-Asp-fluoromethyl ketone (z-VAD-FMK). Cell death was analyzed by ANXA5-PI staining followed by flow cytometry. Noscapine Z-VAD-FMK dramatically decreased the ANXA5-positive cell population induced by 5-FU but showed no effect on that induced by (Fig.?3A). The results of the trypan blue exclusion assay also showed that z-VAD-FMK significantly suppressed the apoptotic cell death caused by 5-FU but failed to protect the cells from death caused by (Fig.?3D). To confirm these results, Noscapine we performed Apo-BrdU assays (to detect apoptotic cells) and immunoblotting. As shown in Figure 3G, failed.