Novel Ultrastructural Insights into the Clear-Cell Carcinoma of the Pancreas: A Case Report.

Pancreatic cancer, most frequently as ductal adenocarcinoma (PDAC), is the third leading cause of cancer death. Clear-cell primary adenocarcinoma of the pancreas (CCCP) is a rare, aggressive, still poorly characterized subtype of PDAC. We report here a case of a 65-year-old male presenting with pancreatic neoplasia. A histochemical examination of the tumor showed large cells with clear and abundant intracytoplasmic vacuoles. The clear-cell foamy appearance was not related to the hyperproduction of mucins. Ultrastructural characterization with transmission electron microscopy revealed the massive presence of mitochondria in the clear-cell cytoplasm. The mitochondria showed disordered cristae and various degrees of loss of structural integrity. Immunohistochemistry staining for NADH dehydrogenase [ubiquinone] 1 alpha subcomplex, 4-like 2 (NDUFA4L2) proved specifically negative for the clear-cell tumor. Our ultrastructural and molecular data indicate that the clear-cell nature in CCCP is linked to the accumulation of disrupted mitochondria. We propose that this may impact on the origin and progression of this PDAC subtype.

Morphofunctional analysis of human pancreatic cancer cell lines in 2- and 3-dimensional cultures.

Genetic, transcriptional, and morphological differences have been reported in pancreatic ductal adenocarcinoma (PDAC) cases. We recently found that epithelial or mesenchymal features were enhanced in three-dimensional (3D) cultures compared to two-dimensional (2D) cultures. In this study, we examined the differences in the morphological and functional characteristics of eight PDAC cell lines in 2D and 3D cultures. Most PDAC cells showed similar pleomorphic morphologies in 2D culture. Under 3D culture, PDAC cells with high E-cadherin and low vimentin expression levels (epithelial) formed small round spheres encircled with flat lining cells, whereas those with high vimentin and low E-cadherin expression levels (mesenchymal) formed large grape-like spheres without lining cells and were highly proliferative. In 3D culture, gemcitabine was more effective for the spheres formed by PDAC cells with epithelial features, while abraxane was more effective on those with mesenchymal features. The expression levels of drug transporters were highest PDAC cells with high vimentin expression levels. These findings indicate that PDAC cells possess various levels of epithelial and mesenchymal characteristics. The 3D-culture method is useful for investigating the diversity of PDAC cell lines and may play important roles in the development of personalized early diagnostic methods and anticancer drugs for PDAC.

Three-dimensional culture of a pancreatic cancer cell line, SUIT-58, with air exposure can reflect the intrinsic features of the original tumor through electron microscopy.

Mini-abstract: Application of a three-dimensional culture system with air exposure facilitates the formation of large cell spheres possessing cribriform glands and producing mucin in the collagen gel. Transmission electron microscopy revealed the formation of microvilli and junctional complexes at the apical side of the cell. This study aimed to reproduce the characteristics of original adenocarcinoma tumors in vitro. The pancreatic cell line, SUIT-58, derived from a moderately differentiated adenocarcinoma of metastatic pancreatic cancer was used. The cells have a sheet structure in conventional cell culture without forming glands or exhibiting mucin production in the lumen. First, the necessity of scaffolds to create an adenocarcinoma-like microenvironment for SUIT-58 pancreatic cancer cells was assessed. Compared with conventional culture plates, the use of type I collagen as a scaffold played an important role in the formation of densely congested microvilli, as observed through scanning electron microscopy. As gland formation is one of the features of adenocarcinoma, we also assessed gland formation. Use of a recently developed three-dimensional culture system with air exposure resulted in the formation of large cell spheres possessing cribriform glands, which released mucin into the lumen. Transmission electron microscopy also revealed the formation of microvilli in the lumen of the glands and junctional complex at the intercellular part, which were similar to those observed in xenografts. These findings indicate that an in vitro three-dimensional culture system with air exposure reflects the intrinsic features of the original tumor, suggesting that this culture system could be useful for preliminary research of certain cancers.

Pancreatic Cancer Cells Require the Transcription Factor MYRF to Maintain ER Homeostasis.

Many tumors of endodermal origin are composed of highly secretory cancer cells that must adapt endoplasmic reticulum (ER) activity to enable proper folding of secreted proteins and prevent ER stress. We found that pancreatic ductal adenocarcinomas (PDACs) overexpress the myelin regulatory factor (MYRF), an ER membrane-associated transcription factor (TF) released by self-cleavage. MYRF was expressed in the well-differentiated secretory cancer cells, but not in the poorly differentiated quasi-mesenchymal cells that coexist in the same tumor. MYRF expression was controlled by the epithelial identity TF HNF1B, and it acted to fine-tune the expression of genes encoding highly glycosylated, cysteine-rich secretory proteins, thus preventing ER overload. MYRF-deficient PDAC cells showed signs of ER stress, impaired proliferation, and an inability to form spheroids in vitro, while in vivo they generated highly secretory but poorly proliferating and hypocellular tumors. These data indicate a role of MYRF in the control of ER homeostasis in highly secretory PDAC cells.

Arenobufagin Inhibits the Phosphatidylinositol 3-kinase/Protein Kinase B/Mammalian Target of Rapamycin Pathway and Induces Apoptosis and Autophagy in Pancreatic Cancer Cells.

OBJECTIVE: The aim of the study was to investigate the effects of arenobufagin on pancreatic carcinoma in vitro and in vivo and its molecular mechanism. METHODS: The proliferation of pancreatic cancer cells was detected by the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay. Transmission electron microscopy was used to observe the formation of autophagic vacuoles after arenobufagin treatment. Hoechst 33258 and monodansylcadaverine fluorescence staining were performed to evaluate cell apoptosis and autophagy. Annexin V-fluorescein isothiocyanate/propidium iodide double-staining and JC-1 staining assays were used to evaluate apoptosis-related changes. Reverse-transcription polymerase chain reaction and western blotting were carried out to examine the expression of apoptosis- and autophagy-related markers after arenobufagin treatment. A tumor xenograft nude mouse model was established to evaluate arenobufagin efficacy in vivo. RESULTS: Arenobufagin effectively inhibited the proliferation of SW1990 and BxPC3 cells and induced cell arrest, apoptosis, and autophagy. Arenobufagin upregulated the expression of apoptotic- and autophagy-related proteins while downregulated the expression of phosphatidylinositol 3-kinase family proteins. Furthermore, arenobufagin also exerted inhibitory effects on tumor growth in xenograft nude mice. CONCLUSIONS: Arenobufagin inhibits tumor growth in vivo and in vitro. The mechanism underlying arenobufagin action may involve induction of autophagy and apoptosis through the phosphatidylinositol 3-kinase/protein kinase B/mammalian target of rapamycin pathway.

TGFB1-induced autophagy affects the pattern of pancreatic cancer progression in distinct ways depending on SMAD4 status.

Pancreatic ductal adenocarcinoma (PDAC) is one of the most aggressive and lethal malignancies. Given that macroautophagy/autophagy activation is prevalent in PDAC, the dual roles of autophagy could be involved in PDAC heterogeneity. In this work, we demonstrated that TGFB1 induced autophagic flux through SMAD4-dependent or SMAD4-independent pathways based on a distinct genetic context. In SMAD4-positive PDAC cells, TGFB1-induced autophagy promoted proliferation and inhibited migration by decreasing the nuclear translocation of SMAD4. Conversely, TGFB1-induced autophagy inhibited proliferation and promoted migration in SMAD4-negative cells through the regulation of MAPK/ERK activation. TGFB1 expression also positively correlated with LC3B expression in PDAC specimens. A high level of LC3B was associated with unfavorable overall survival (OS) and disease-free survival (DFS) in SMAD4-negative PDAC patients, although LC3B could not predict OS and DFS for the 110 PDAC patients. Thus, TGFB1-induced autophagy contributed to the different patterns of PDAC progression. This knowledge can aid in improving our understanding of the molecular classification of PDAC and might guide the development of therapeutic strategies for PDAC, especially for SMAD4-negative PDAC.Abbreviations: CDH1: cadherin 1; CDH2: cadherin 2; CI: combination index; CQ: chloroquine; DFS: disease-free survival; EMT: epithelial-to-mesenchymal transition; ERK: extracellular signal-regulated protein kinase; GFP: green fluorescent protein; IHC: immunohistochemistry; MAP1LC3B/LC3B: microtubule associated protein 1 light chain 3 beta; MAPK: mitogen-activated protein kinase; OS: overall survival; PBS: phosphate-buffered saline; PDAC: pancreatic ductal adenocarcinoma; RAP: rapamycin; RFP: red fluorescent protein; RT: room temperature; shRNA: short-hairpin RNA; SQSTM1: sequestosome 1; TCGA: The Cancer Genome Atlas; TEM: transmission electron microscopy; TGFB1: transforming growth factor beta 1; TMA: tissue microarray.

Characterization of the metastatic potential of the floating cell component of MIA PaCa-2, a human pancreatic cancer cell line.

In pancreatic cancer, morphologically and functionally heterogeneous cancer cells reside within the same patient. The heterogeneity is believed to promote metastasis and resistance to chemoradiotherapy. MIA PaCa-2, an established human pancreatic ductal adenocarcinoma (PDAC) cell line, contains round and spindle-shaped adherent cells, as well as, round floating cells. In this study, we aimed to assess if the floating cells might have greater metastatic potential and/or be more resistant to drug-induced apoptosis compared to adherent cells. Time-lapse analysis revealed that the two types of adherent cells transformed bilaterally, and some of the adherent, round cells converted to floating cells. Flow cytometry and electron microscopy showed that approximately 90% of the floating cells were viable. qRT-PCR analysis revealed that floating cells expressed lower levels of integrins and ATP-binding cassette (ABC) transporters than adherent cells. In contrast, except for vimentin, floating cells expressed more epithelial to mesenchymal transition markers than adherent cells. Floating cells included a larger population of G2/M-phase cells, and migration assays revealed a decreased migration ability by floating cells relative to adherent cells. A cell aggregation assay showed that the aggregative properties of the floating cells were lower than those of the adherent cells. In 3D culture, spheres derived from floating cells were more sensitive to anti-cancer drugs, including gemcitabine, 5-FU, and abraxane, than those derived from adherent cells. Expression levels of stemness markers in the spheres derived from floating cells were lower than those derived from adherent cells. Morphological characterization of human PDAC cell lines may help to clarify the series of alterations cancer cells undergo during the metastatic process and may contribute to the development of new PDAC diagnostics and more patient-specific treatments for those with PDAC.

Biphenotypic Differentiation of Pancreatic Cancer in 3-Dimensional Culture.

OBJECTIVE: Pancreatic ductal adenocarcinoma (PDAC) is the third most common cause of cancer death in the United States. Improved characterized models of PDAC are needed for drug screening. METHODS: We grew 4 established pancreatic cancer cell lines in hanging drop cultures to produce spheroids. We also grew organoids from explanted xenografted PDAC and surgically resected primary PDAC. We performed transmission and scanning electron microscopy and compared findings with those of the normal pancreatic duct. We also performed single-cell cloning to determine the potential options for differentiation. RESULTS: Spheroids contained tight junctions and desmosomes but lacked zymogen granules, as expected. The former features were present in normal pancreatic duct but absent from PDAC cell lines grown in standard 2-dimensional culture. Spheroids functionally excluded macromolecules in whole mounts. Cells on the surface of PDAC spheroids were carpeted by microvilli except for rare cells with prominent stereocilia. Carpets of microvilli were also seen in low passage organoids produced from xenografts and surgically resected human PDAC, in addition to normal human pancreatic duct. We performed single-cell cloning and resulting spheroids produced both cell phenotypes at the same approximate ratios as those from bulk cultures. CONCLUSIONS: Pancreatic cancer spheroids/organoids are capable of biphenotypic differentiation.

Tumor selective uptake of drug-nanodiamond complexes improves therapeutic outcome in pancreatic cancer.

Pancreatic ductal adenocarcinoma (PDAC) is one of the leading causes of cancer-related deaths and novel treatment approaches are urgently needed. Here we show that poly(ethylene glycol)-functionalized nanodiamonds loaded with doxorubicin (ND-PEG-DOX) afforded a considerable improvement over free drug in an orthotopic pancreatic xenograft model. ND-PEG-DOX complexes were also superior to free DOX in 3-dimensional (3D) tumor spheroids of PDAC. ND-PEG showed no cytotoxicity towards macrophages, and histopathological analysis showed no abnormalities of major organs upon in vivo administration of ND-PEG-DOX. These results provide evidence that ND-mediated drug delivery may serve as a means of improving the therapeutic outcome in PDAC.

TKS5-positive invadopodia-like structures in human tumor surgical specimens.

Invadopodia, cancer cell protrusions with proteolytic activity, are functionally associated with active remodeling of the extracellular matrix. Here, we show that the invadopodia-related protein TKS5 is expressed in human pancreatic adenocarcinoma lines, and demonstrate that pancreatic cancer cells depend on TKS5 for invadopodia formation and function. Immunofluorescence staining of human pancreatic cancer cells reveals that TKS5 is a marker of mature and immature invadopodia. We also analyze the co-staining patterns of TKS5 and the commonly used invadopodia marker Cortactin, and find only partial co-localization of these two proteins at invadopodia, with a large fraction of TKS5-positive invadopodia lacking detectable levels of Cortactin. Whereas compelling evidence exist on the role of invadopodia as mediators of invasive migration in cultured cells and in animal models of cancer, these structures have never been detected inside human tumors. Here, using antibodies against TKS5 and Cortactin, we describe for the first time structures strongly resembling invadopodia in various paraffin-embedded human tumor surgical specimens from pancreas and other organs. Our results strongly suggest that invadopodia are present inside human tumors, and warrants further investigation on their regulation and occurrence in surgical specimens, and on the value of TKS5 antibodies as pathological research and diagnostic tools.

Delivery of miR-212 by chimeric peptide-condensed supramolecular nanoparticles enhances the sensitivity of pancreatic ductal adenocarcinoma to doxorubicin.

Pancreatic ductal adenocarcinoma (PDAC) is a destructive cancer with poor prognosis. Both novel therapeutic targets and approaches are needed to improve the overall survival of PDAC patients. MicroRNA-212 (miR-212) has been reported as a tumor suppressor in multiple cancers, but its definitive role and exact mechanism in the progression of pancreatic cancer is unclear. In this study, we developed a new chimeric peptide (PL-1) composed of plectin-1-targeted PDAC-specific and arginine-rich RNA-binding motifs which could condense miRNA to self-assemble supramolecular nanoparticles. These nanoparticles could deliver miR-212 into PDAC cells specifically and efficiently which also showed good stability in RNase and serum. Moreover, we demonstrated that PL-1/miR-212 nanoparticles could dramatically enhance the chemotherapeutic effect of doxorubicin for PDAC both in vitro and in vivo. In terms of mechanism, combined miR-212 intervention by PL-1/miR-212 nanoparticles resulted in obvious decrease of USP9X expression (ubiquitin specific peptidase 9, X-linked, USP9X) and eventually enhanced the doxorubicin induced apoptosis and autophagy of PDAC cells. These findings provide a new promising anti-cancer strategy via PL-1/miR-212 nanoparticles and identify miR-212/USP9X as a new potential target for future systemic therapy against human PDAC.

iRGD-guided Tumor-penetrating Nanocomplexes for Therapeutic siRNA Delivery to Pancreatic Cancer.

Pancreatic cancer is one of the leading causes of cancer-related death, with 5-year survival of 8.5%. The lack of significant progress in improving therapy reflects our inability to overcome the desmoplastic stromal barrier in pancreatic ductal adenocarcinoma (PDAC) as well as a paucity of new approaches targeting its genetic underpinnings. RNA interference holds promise in targeting key mutations driving PDAC; however, a nucleic acid delivery vehicle that homes to PDAC and breaches the stroma does not yet exist. Noting that the cyclic peptide iRGD mediates tumor targeting and penetration through interactions with αvß3/5 integrins and neuropilin-1, we hypothesized that "tandem" peptides combining a cell-penetrating peptide and iRGD can encapsulate siRNA to form tumor-penetrating nanocomplexes (TPN) capable of delivering siRNA to PDAC. The use of directly conjugated iRGD is justified by receptor expression patterns in human PDAC biopsies. In this work, we optimize iRGD TPNs with polyethylene glycol (PEG)-peptide conjugates for systemic delivery to sites of disease. We show that TPNs effectively knockdown siRNA targets in PDAC cell lines and in an immunocompetent genetically engineered mouse model of PDAC. Furthermore, we validate their tumor-penetrating ability in three-dimensional organoids and autochthonous tumors. In murine therapeutic trials, TPNs delivering anti-Kras siRNA significantly delay tumor growth. Thus, iRGD TPNs hold promise in treating PDAC by not only overcoming physical barriers to therapy, but by leveraging the stroma to achieve knockdown of the gold-standard genetic target. Moreover, the modular construction of this delivery platform allows for facile adaptation to future genetic target candidates in pancreatic cancer. Mol Cancer Ther; 17(11); 2377-88. ©2018 AACR.

Inhibition of endoplasmic-reticulum-stress-mediated autophagy enhances the effectiveness of chemotherapeutics on pancreatic cancer.

BACKGROUND: Endoplasmic reticulum (ER) stress and its consequent unfolded protein response (UPR) are believed to be associated with progression, survival and chemoresistance of a variety of tumor cells through multiple cellular processes, including autophagy. Therefore, the ER stress-autophagy pathway presents a potential molecular target for therapeutic intervention. The objective of this study was to evaluate the therapeutic efficacy of ER stress and autophagy modulators in the context of pancreatic ductal adenocarcinoma (PDAC). METHODS: We first targeted IRE1α, an important regulator of the UPR, through STF-083010 treatment in PDAC cell lines in vitro. Chloroquine was then used to target autophagy and an optimal combination treatment was developed using chloroquine, sunitinib and gemcitabine. Apoptosis was analyzed using TUNEL assay, autophagy was estimated using lysotracker staining and electron microscopy, and UPR was analyzed using anti-GRP78 immunostaining and XBP1 splicing. Transplantation of PDAC derived KPCP1 and Panc02 cells in mouse pancreas were performed to study treatment efficacy in vivo. RESULTS: Suppression of the IRE1α by STF-083010 alone resulted in increased lysosomes and reduced viability of PDAC cells. Chloroquine treatment alone inhibited downstream autophagy but was insufficient in reducing PDAC cell growth. However, combining STF-083010 and chloroquine had additive anti-tumor efficacy when used with gemcitabine. Sunitinib alone caused abnormal maturation of the autolysosomes with increased intracellular multivesicular bodies and increased apoptosis evident in PDAC cells. Sunitinib showed a synergistic effect with chloroquine in reducing in vitro PDAC cell viability and significantly increased the efficacy of gemcitabine in human and murine PDAC cell lines. The anti-proliferative effect of gemcitabine was significantly increased when used in combination with sunitinib and/or chloroquine in both in vitro and in vivo PDAC models. The addition of sunitinib and/or chloroquine to gemcitabine, resulted in a significantly increased survival of the animals without noticeably increased toxicity. Sunitinib, gemcitabine and chloroquine treated mice showed a significant reduction of GRP78 expression, reduced cell proliferation and increased apoptosis in pancreas, compatible with a tumor response. CONCLUSIONS: Sunitinib combined with chloroquine reduces tumor growth through suppression of autophagy and increased apoptosis. Co-administration of modulators of ER stress-mediated autophagy with chemotherapy presents a novel therapeutic approach in PDAC.

Facile assembly of upconversion nanoparticle-based micelles for active targeted dual-mode imaging in pancreatic cancer.

BACKGROUND: Pancreatic cancer remains the leading cause of cancer-related deaths, the existence of cancer stem cells and lack of highly efficient early detection may account for the poor survival rate. Gadolinium ion-doped upconversion nanoparticles (UCNPs) provide opportunities for combining fluorescent with magnetic resonance imaging, and they can improve the diagnostic efficacy of early pancreatic cancer. In addition, as one transmembrane glycoprotein overexpressed on the pancreatic cancer stem cells, CD326 may act as a promising target. In this study, we developed a facile strategy for developing anti-human CD326-grafted UCNPs-based micelles and performed the corresponding characterizations. After conducting in vitro and vivo toxicology experiments, we also examined the active targeting capability of the micelles upon dual-mode imaging in vivo. RESULTS: We found that the micelles owned superior imaging properties and long-time stability based on multiple characterizations. By performing in vitro and vivo toxicology assay, the micelles had good biocompatibility. We observed more cellular uptake of the micelles with the help of anti-human CD326 grafted onto the micelles. Furthermore, we successfully concluded that CD326-conjugated micelles endowed promising active targeting ability by conducting dual-mode imaging in human pancreatic cancer xenograft mouse model. CONCLUSIONS: With good biocompatibility and excellent imaging properties of the micelles, our results uncover efficient active homing of those micelles after intravenous injection, and undoubtedly demonstrate the as-obtained micelles holds great potential for early pancreatic cancer diagnosis in the future and would pave the way for the following biomedical applications.

A RhoA-FRET Biosensor Mouse for Intravital Imaging in Normal Tissue Homeostasis and Disease Contexts.

The small GTPase RhoA is involved in a variety of fundamental processes in normal tissue. Spatiotemporal control of RhoA is thought to govern mechanosensing, growth, and motility of cells, while its deregulation is associated with disease development. Here, we describe the generation of a RhoA-fluorescence resonance energy transfer (FRET) biosensor mouse and its utility for monitoring real-time activity of RhoA in a variety of native tissues in vivo. We assess changes in RhoA activity during mechanosensing of osteocytes within the bone and during neutrophil migration. We also demonstrate spatiotemporal order of RhoA activity within crypt cells of the small intestine and during different stages of mammary gestation. Subsequently, we reveal co-option of RhoA activity in both invasive breast and pancreatic cancers, and we assess drug targeting in these disease settings, illustrating the potential for utilizing this mouse to study RhoA activity in vivo in real time.

"Stealth dissemination" of macrophage-tumor cell fusions cultured from blood of patients with pancreatic ductal adenocarcinoma.

Here we describe isolation and characterization of macrophage-tumor cell fusions (MTFs) from the blood of pancreatic ductal adenocarcinoma (PDAC) patients. The MTFs were generally aneuploidy, and immunophenotypic characterizations showed that the MTFs express markers characteristic of PDAC and stem cells, as well as M2-polarized macrophages. Single cell RNASeq analyses showed that the MTFs express many transcripts implicated in cancer progression, LINE1 retrotransposons, and very high levels of several long non-coding transcripts involved in metastasis (such as MALAT1). When cultured MTFs were transplanted orthotopically into mouse pancreas, they grew as obvious well-differentiated islands of cells, but they also disseminated widely throughout multiple tissues in "stealth" fashion. They were found distributed throughout multiple organs at 4, 8, or 12 weeks after transplantation (including liver, spleen, lung), occurring as single cells or small groups of cells, without formation of obvious tumors or any apparent progression over the 4 to 12 week period. We suggest that MTFs form continually during PDAC development, and that they disseminate early in cancer progression, forming "niches" at distant sites for subsequent colonization by metastasis-initiating cells.

MicroRNA-155 Controls Exosome Synthesis and Promotes Gemcitabine Resistance in Pancreatic Ductal Adenocarcinoma.

The cancer drug gemcitabine (GEM) is a key drug for treating pancreatic ductal adenocarcinoma (PDAC), but PDAC cells develop chemoresistance after long-term administration. Since the tolerance was immediately spread to every PDAC tissue in a patient, it is assumed that some certain efficient mechanisms underlay in the development of chemoresistance. Changes in the levels of particular microRNAs or alterations in intercellular communication play a dominant role in chemoresistance development, and recent data also suggest that exosomes play an important role in this process. In this study, we revealed that the loop conferred chemoresistance in PDAC cells. The loop was as follows; 1, The long-term exposure of GEM increased miR-155 expression in PDAC cells. 2, The increase of miR-155 induced two different functions; exosome secretion and chemoresistance ability via facilitating the anti-apoptotic activity. 3, Exosome deliver the miR-155 into the other PDAC cells and induce the following function. The target therapy to miR-155 or the exosome secretion effectively attenuated the chemoresistance, and these results were validated with both clinical samples and in vivo experiments. This mechanism represents a novel therapeutic target in GEM treatment to PDAC.

miR-138-5p suppresses autophagy in pancreatic cancer by targeting SIRT1.

The role of microRNA in the aberrant autophagy that occurs in pancreatic cancer remains controversial. Because hypoxia is known to induce autophagy, we screened for differentially expressed microRNAs using a miRNA microarray with pancreatic cancer cells cultured under normoxic and hypoxic conditions. We found that miR-138-5p was among the most downregulated miRNA in hypoxia-stimulated cells, and that overexpression of miR-138-5p substantially reduced expression of autophagy markers. In addition, western blot and immunofluorescence analyses and electron microscopy revealed that miR-138-5p inhibited autophagy in pancreatic cancer cells and blocked serum starvation-induced autophagic flux independently of the typical autophagic signaling pathway. miR-138-5p had no effect on ATG3, ATG5, or ATG7, three primary autophagy-associated genes. Instead, miR-138-5p specifically targeted the SIRT1 3' untranslated region and suppressed autophagy by reducing the level of SIRT1, which acetylates FoxO1 and regulates autophagy via FoxO1/Rab7. SIRT1 or Rab7 knockdown blocked the SIRT1/FoxO1/Rab7 axis and suppressed autophagic inhibition by miR-138-5p. Finally, we found that miR-138-5p inhibited autophagy and tumor growth in vivo. These results indicate that miR-138-5p suppresses autophagy in pancreatic cancer by targeting SIRT1.