Purpose: Gemcitabine (GEM)-based chemotherapy is the first-line treatment for locally advanced pancreatic cancer. GEM resistance, however, remains a significant clinical challenge. Here, we investigated whether exosomes derived from GEM-resistant pancreatic cancer stem cells (CSCs) mediate cell-cell communication between cells that are sensitive or resistant to GEM and, by doing so, regulate drug resistance. Methods: GEM-sensitive BxPC-3-derived Bx_S and PANC-1 pancreatic cancer cells were cultured with exosomes extracted from CSCs isolated from GEM-resistant BxPC-3-derived Bx_R cells (Bx_R-CSC). The effect of exosomes on drug resistance, cell cycle progression, apoptosis and miRNA expression was evaluated in Bx_S and PANC-1 cells. Relevant miRNAs associated with GEM resistance were identified and the role of miR-210 in conferring drug resistance was examined in vitro and in vivo. Results: Bx_R-CSC-derived exosomes induced GEM resistance, inhibited GEM-induced cell cycle arrest, antagonized GEM-induced apoptosis, and promoted tube formation and cell migration in Bx_S and PANC-1 cells. Elevated miR-210 expression levels were detected in Bx_R-CSCs and Bx_R-CSC-derived exosomes compared to those in Bx_S-CSCs and Bx_S-CSC-derived exosomes. In addition, increased expression levels of miR-210 were observed in Bx_S and PANC-1 cells cultured with Bx_R-CSC-derived exosomes upon exposure to GEM in a dose-dependent manner. Also, a series of biological changes was observed in Bx_S cells after transfection with miR-210 mimics, including activation of the mammalian target of rapamycin (mTOR) signaling pathway, and these changes were similar to those triggered by Bx_R-CSC-derived exosomes. Conclusions: Our findings suggest that exosomes derived from GEM-resistant pancreatic cancer stem cells mediate the horizontal transfer of drug-resistant traits to GEM-sensitive pancreatic cancer cells by delivering miR-210.

Background: Gemcitabine (GEM) resistance remains a significant clinical challenge in pancreatic cancer treatment. Here, we investigated the therapeutic utility of everolimus (Evr), an inhibitor of mammalian target of rapamycin (mTOR), in targeting the Warburg effect to overcome GEM resistance in pancreatic cancer. Methods: The effect of Evr and/or mTOR overexpression or GEM on cell viability, migration, apoptosis, and glucose metabolism (Warburg effect) was evaluated in GEM-sensitive (GEM_sen) and GEM-resistant (GEM_res) pancreatic cancer cells. Results: We demonstrated that the upregulation of mTOR enhanced cell viability and favored the Warburg effect in pancreatic cancer cells via the regulation of PI3K/AKT/mTOR signaling. However, this effect was counteracted by Evr, which inhibited aerobic glycolysis by reducing the levels of glucose, lactic acid, and adenosine triphosphate and suppressing the expression of glucose transporter 1, lactate dehydrogenase-B, hexokinase 2, and pyruvate kinase M2 in GEM_sen and GEM_res cells. Evr also promoted apoptosis by upregulating the pro-apoptotic proteins Bax and cytochrome-c and downregulating the anti-apoptotic protein Bcl-2. GEM was minimally effective in suppressing GEM_res cell activity, but the therapeutic effectiveness of Evr against pancreatic cancer growth was greater in GEM_res cells than that in GEM_sen cells. In vivo studies confirmed that while GEM failed to inhibit the progression of GEM_res tumors, Evr significantly decreased the volume of GEM_res tumors while suppressing tumor cell proliferation and enhancing tumor apoptosis in the presence of GEM. Conclusions: Evr treatment may be a promising strategy to target the growth and activity of GEM-resistant pancreatic cancer cells by regulating glucose metabolism via inactivation of PI3K/AKT/mTOR signaling.

Pancreatic cancer (PC) is a serious threat to human health, with most patients diagnosed at the advanced stages of the disease. Treatment with gemcitabine (GEM) leads to PC GEM resistance. In addition, cancer stem cell (CSC)-derived exosomes play an important role in cancer progression. We aimed to investigate the role and mechanism of action of PC stem cell-derived exosomes in PC drug resistance and progression. CSC-derived exosomes increased the proportion of F4/80+/CD86 + cells and levels of M2 polarization factors. miR-210 is expressed in CSC-derived exosomes. Thus, following co-culture, miR-210 was taken up by macrophages. Transfection or the addition of miR-210 mimics increased the proportion of F4/80+/CD206 + cells and levels of M2 polarization factors. Further, the miR-210 targets inhibited the levels of FGFRL1. The FGFRL1 overexpression plasmid also inhibited miR-210-mediated M2 polarization. After co-culture of THP-M2 cells with PC cells and treatment with GEM, the survival rate, migration rate, and levels of MDR, YB-1, BCRP, p-PI3K, p-AKT, and p-mTOR in PC cells increased. And THP-M2 increased the tumor volume and MDR, YB-1, BCRP, p-PI3K, p-AKT, and p-mTOR levels. Overall, miR-210 from PC stem cell-derived exosome targets and inhibits FGFRL1 to promote macrophage M2 polarization, which activates the p-PI3K/p-AKT/p-mTOR pathway and increases GEM resistance.

Gemcitabine (GEM) resistance remains a challenging clinical issue to overcome in chemotherapy against pancreatic cancer. We previously demonstrated that miR-210 derived from pancreatic cancer stem cells enhanced the GEM-resistant properties of pancreatic cancer cells, thus identifying miR-210 as an oncogenic miRNA. Herein, we report the existence of an upstream effector that acts as a competing endogenous RNA (ceRNA) to miR-210. Bioinformatic screening was performed to identify lncRNAs with a binding relationship to miR-210. Overexpression and interference vectors were constructed to demonstrate the effect of ceRNA activity in pancreatic cell behavior, both in vitro and in vivo. DLEU2L (deleted in lymphocytic leukemia 2-like), which is expressed at low levels in pancreatic cancer tissues, was shown to exhibit a binding relationship with miR-210-3p. Overexpression of DLEU2L and silencing of miR-210-3p suppressed the proliferation, migration, and invasion of pancreatic cancer cells while promoting apoptosis. These effects occurred via the inhibition of the Warburg effect (aerobic glycolysis) and AKT/mTOR signaling. In addition, we showed that BRCA2 is a target gene of miR-210-3p, and the downregulation of miR-210-3p by DLEU2L effectively induced an upregulation of BRCA2 via the ceRNA mechanism. In vivo, DLEU2L overexpression and miR-210-3p interference suppressed pancreatic tumor progression, consistent with the results of in vitro studies. The findings of our study establish DLEU2L as a ceRNA to miR-210-3p and reveal the critical role of the DLEU2L/miR-210-3p crosstalk in targeting GEM resistance.