Yoshifumi Iwagami

miR-320c regulates gemcitabine-resistance in pancreatic cancer via SMARCC1

Background:Gemcitabine-based chemotherapy is the standard treatment for pancreatic cancer. However, the issue of resistance remains unresolved. The aim of this study was to identify microRNAs (miRNAs) that govern the resistance to gemcitabine in pancreatic cancer.Methods:miRNA microarray analysis using gemcitabine-resistant clones of MiaPaCa2 (MiaPaCa2-RGs), PSN1 (PSN1-RGs), and their parental cells (MiaPaCa2-P, PSN1-P) was conducted. Changes in the anti-cancer effects of gemcitabine were studied after gain/loss-of-function analysis of the candidate miRNA. Further assessment of the putative target gene was performed in vitro and in 66 pancreatic cancer clinical samples.Results:miR-320c expression was significantly higher in MiaPaCa2-RGs and PSN1-RGs than in their parental cells. miR-320c induced resistance to gemcitabine in MiaPaCa2. Further experiments showed that miR-320c-related resistance to gemcitabine was mediated through SMARCC1, a core subunit of the switch/sucrose nonfermentable (SWI/SNF) chromatin remodeling complex. In addition, clinical examination revealed that only SMARCC1-positive patients benefited from gemcitabine therapy with regard to survival after recurrence (P=0.0463).Conclusion:The results indicate that miR-320c regulates the resistance of pancreatic cancer cells to gemcitabine through SMARCC1, suggesting that miR-320c/SMARCC1 could be suitable for prediction of the clinical response and potential therapeutic target in pancreatic cancer patients on gemcitabine-based therapy.

Involvement of microRNA-181b in the gemcitabine resistance of pancreatic cancer cells

BACKGROUND/OBJECTIVES MicroRNAs (miRs) have been shown to regulate the sensitivity to several chemotherapeutic agents in various types of cancers. MiR-181b is one of such regulators, yet its importance in pancreatic cancer is not determined so far. The aim of this study was to investigate the relationship between microRNA (miR)-181b expression and gemcitabine resistance in pancreatic cancer cells. METHODS The effects of overexpression or knockdown of miR-181b on four pancreatic cancer cell lines exposed to gemcitabine were examined. The induction of apoptosis and the changes of the cylindromatosis (CYLD) protein were examined. Furthermore, the effect of small interference RNA for CYLD (siCYLD) on cell viability and the relationship between CYLD and nuclear factor kappa B (NF-κB) were investigated. RESULTS The expression of miR-181b was higher in BxPC3, Panc1 and PSN1 cells compared with MiaPaCa2 cells. Pre-miR-181b transfection into MiaPaCa2 cells increased their gemcitabine resistance, whereas anti-miR-181b transfection into the other pancreatic cancer cell lines reduced their resistance to gemcitabine and led to the induction of apoptosis. The protein levels of CYLD were increased by anti-miR-181b in Panc1 and PSN1 cells. Inhibition of CYLD increased the NF-κB activity and gemcitabine resistance in Panc1 and PSN1 cells. CONCLUSIONS The present study demonstrated that miR-181b was associated with the resistance of pancreatic cancer cells to gemcitabine, and verified that miR-181b enhances the activity of NF-κB by inhibiting CYLD, leading to the resistance to gemcitabine. Our results suggest that miR-181b is a potential target for decreasing gemcitabine resistance.

A novel preoperative predictor of pancreatic fistula using computed tomography after distal pancreatectomy with staple closure

PurposeA thick pancreas has proven to be a conspicuous predictor of pancreatic fistula (PF) following distal pancreatectomy (DP) using staples. Other predictors for this serious surgical complication currently remain obscure. This study sought to identify novel predictors of PF following DP.MethodsOne hundred and twenty-two patients were retrospectively assessed to determine the correlation between PF occurrence and the clinicopathological findings and radiologic data from preoperative computed tomography (CT). CT assessments included the thickness of the pancreas (TP) and pancreatic CT number (pancreatic index; PI), calculated by dividing the pancreatic CT by the splenic CT density.ResultsTwenty-four patients (19.7%) developed a clinically relevant PF. TP was identified as an independent risk factor for PF in multivariate analyses (odds ratio 1.17; P = 0.0095). In subgroup analyses, a lower PI in a thick pancreas was a significant predictor of PF (P = 0.032). The combination of these two prediction parameters, known as the TP-to-PI ratio (TPIR), showed a significantly better prediction ability than TP alone (area under the receiver operating characteristic curve for the incidence of PF, TPIR 0.80 vs. TP 0.69; P = 0.037).ConclusionCombining the CT number with TP substantially improves the prediction ability for the incidence of PF following DP with staple use.

MicroRNA-181b-5p, ETS1, and the c-Met pathway exacerbate the prognosis of pancreatic ductal adenocarcinoma after radiation therapy

Preoperative chemoradiation therapy (CRT) for pancreatic ductal adenocarcinoma (PDAC) has emerged as a reasonable strategy that shows good prognostic impact. However, after preoperative CRT, resected specimens show remnant tumor cells, which indicate that some tumor cells had acquired or were selected for resistance to CRT. Recently, two oncological mechanisms, the EMT and the presence of CSCs, were reported to be associated with resistance in various cancers. Previous reports showed that HGF could induce EMT in PDAC cells; moreover, the HGF receptor, c‐Met, was identified as a dominant pancreatic CSC marker. However, the clinical significance of c‐Met expression remains unclear. So, we hypothesized that remnant PDAC tissue after CRT might harbor cells with high c‐Met expression, and these cells may exacerbate patients’ prognosis. In the immunohistochemical analysis, we showed that preoperative CRT was significantly associated with high c‐Met expression; moreover, high c‐Met expression was a significant marker of a dismal prognosis. Next, we investigated mechanisms of c‐Met upregulation in PDAC cells. We established GEM‐resistant and radioresistant PDAC cells to analyze the transcriptome involved in c‐Met expression. The microarray data for the established radiation‐resistant PDAC cells indicated miR‐181b‐5p downregulation, which targets ETS1, one of the transcription factors for c‐Met, and it was shown that radiation exposure induced c‐Met expression through ETS1 increase by the suppression of miR‐181b‐5p. These results suggested that targeting these mechanisms may promote the development of a novel multidisciplinary treatment strategy for improving preoperative CRT efficiency.

Layer-by-layer cell coating technique using extracellular matrix facilitates rapid fabrication and function of pancreatic β-cell spheroids

Tissue engineering of insulin-secreting cells using alternatives to islet transplantation has been fueled by the development of available materials and fabrication techniques. We have established a cell coating technique that enables the cell surface to be coated with extracellular matrix based on the concept of a layer-by-layer (LbL) assembly. The present study evaluated whether this technique is beneficial for fabricating pancreatic β-cell spheroids using a mouse β-cell line. The well-structured and dense spheroids could immediately be constructed by the LbL-coated cells. In the functional analysis, spheroids with the LbL-coated cells had greater insulin secretion ability with increased expression of the insulin and glucose transporter 2 genes versus spheroids with non-coated cells. In addition, we found that the expression of connexin 36, a gap junction molecule, was upregulated by the LbL cell coating. When spheroids with the LbL-coated cells were syngeneically transplanted in diabetic mice, blood glucose levels immediately decreased and glucose sensitivity significantly improved after intraperitoneal glucose stimulation compared to spheroids with non-coated cells. This cell coating technique would be a clinically applicable approach for fabricating pancreatic β-cell spheroids and treating type 1 diabetes mellitus.

Prolonged Neoadjuvant Therapy for Locally Advanced Pancreatic Cancer

Purpose: The purpose of this study was to investigate if prolonged neoadjuvant therapy, for locally advanced pancreatic cancer could be used to identify those patients who could benefit from resection surgery. Methods: Thirty-four patients with locally advanced pancreatic cancer invading the adjacent major arteries underwent chemoradiotherapy, followed by 6 months of systemic chemotherapy, unless their tumor was already resectable. After this combination treatment, surgical resection was performed on those patients with tumors judged to be at least borderline resectable. Results: Reevaluation after chemoradiotherapy revealed that, at that stage, surgery was only possible for one case; resection was successfully performed. Following 6 months of systemic chemotherapy, a further 7 patients were diagnosed with borderline resectable lesions; 4 underwent surgery with no postoperative complications. The median survival time of the patients following surgery was 3.63 years; this was reduced to approximately a third in patients not undergoing surgery (1.01 years; p = 0.0005). Conclusions: For patients with locally advanced pancreatic cancer, prolonged neoadjuvant therapy was successfully used to select candidates that could benefit from surgical resection of their tumor. Resection significantly increased the long-term survival rate.

Vitamin D Supplementation is a Promising Therapy for Pancreatic Ductal Adenocarcinoma in Conjunction with Current Chemoradiation Therapy

BackgroundThe cancer-associated fibroblasts (CAFs) in pancreatic ductal adenocarcinoma (PDAC) are well known to play a dominant role in distant metastasis. Nevertheless, the effect on CAFs with current chemoradiation therapies remains uncertain.ObjectiveThis study aimed to reveal the role of CAFs under current chemoradiation therapy (CRT) and investigate the factors regulating CAFs.Methodsα-SMA-positive cells in 86 resected PDAC specimens with/without preoperative CRT were evaluated by immunohistochemistry. Various factors, including the plasma levels of vitamin D, were investigated for association with the number of CAFs or distant metastasis-free survival (DMFS). Human pancreatic satellite cells (hPSCs) extracted from clinical specimens were used to validate the factors.ResultsAll PDAC samples contained CAFs but the number varied widely. Multivariate analysis for DMFS indicated a larger number of CAFs was a significant risk factor. Univariate analysis for the number of CAFs identified two clinical factors: preoperative CRT and lower plasma levels of vitamin D. In subgroup analysis, the higher plasma level of vitamin D was a dominant factor for longer DMFS in PDAC patients after preoperative CRT. These results were validated by using extracted hPSCs. Irradiation activated stromal cells into CAFs facilitating malignant characteristics of PDAC and the change was inhibited by vitamin D supplementation in vitro.ConclusionIn conjunction with established current therapies, vitamin D supplementation may be an effective treatment for PDAC patients by inactivating CAFs.

Short- and Long-term Outcomes of De Novo Liver Transplant Patients Treated With Once-daily Prolonged-release Tacrolimus

Background Tacrolimus is the key immunosuppressive drug for liver transplantation. Once-daily prolonged-release tacrolimus (TAC-PR) exhibits good drug adherence but has difficulty controlling the trough level in the early phase of liver transplantation. The aim of this study was to compare the feasibility and efficacy of immediately starting oral TAC-PR versus traditional twice-daily tacrolimus (TAC-BID) in de novo liver transplantation recipients. Methods The study included 28 patients treated with conventional TAC-BID and 60 patients treated with TAC-PR (median follow-up 70.5 months). Short-term and long-term outcomes were compared. Results Patient characteristics were similar except for the incidence of hepatocellular carcinoma and type of graft. Dose adjustment was more frequently required for TAC-PR than TAC-BID (73.3% vs 42.9%, P = 0.006), but trough levels of TAC during the first 3 months after liver transplantation were controlled well in both groups. The rate of acute cellular rejection and long-term renal function were similar in both groups. In both groups, renal function worsened during the first 6 months after transplantation and remained stable until the end of the follow-up period. The 1-year, 3-year, and 5-year survival rates were 96.4%, 85.7%, and 85.7% for TAC-BID and 96.7%, 94.8%, and 94.8% for TAC-PR, respectively. The overall survival curve for TAC-PR was not inferior to that of TAC-BID. Conclusions The TAC-PR protocol was feasible and effective with strict adjustment.

Fatty Acid-Mediated Stromal Reprogramming of Pancreatic Stellate Cells Induces Inflammation and Fibrosis That Fuels Pancreatic Cancer

Objectives Pancreatic ductal adenocarcinoma is one of the deadliest diseases worldwide. Fatty acids (FAs) have properties that affect both cancer cells and tumor environment. We assessed the effects of FAs on malignant characteristics in a pancreatic cancer and pancreatic stellate cell (PSC) coculture model. This study aimed to clarify the FA signature of PSC-derived inflammation and fibrosis in vitro and in a clinicopathological analysis. Methods The in vitro model involved coculture of the human pancreatic cancer cell lines PANC-1 and MIA PaCa-2 with human PSCs. Clinical histological samples were analyzed to characterize the surgical margins of samples from patients who received distal pancreatectomies. Results The pancreatic cancer cells took up lipids from the culture media. Saturated and unsaturated FAs were required to induce inflammatory responses in human PSCs, and the cocultures showed fibrotic changes. Clinical samples from pancreatic ductal adenocarcinoma patients had more fatty and fibrotic changes in the normal tissue in the surgical margins than samples from noncancer patients. Conclusions Inflammation and fibrosis levels were increased in pancreatic cancer specimens, supporting the in vitro observations and suggesting that PSCs contribute to pancreatic carcinogenesis. Pancreatic stellate cells thus represent a potential therapeutic target for suppressing stromal changes in pancreatic cancer.

Clinical Impact of Preoperative Sarcopenia on the Postoperative Outcomes After Pancreas Transplantation

BackgroundThe importance of evaluating sarcopenia is increasingly being recognized in the field of transplantation because sarcopenia can have an adverse effect on the treatment outcomes. However, the clinical significance of preoperative sarcopenia on the postoperative outcomes following pancreas transplantation (PTx) has been largely unknown. The objective of this study was to investigate the role of preoperative sarcopenia in predicting the postoperative outcomes following PTx in recipients with type 1 diabetes mellitus (T1D).MethodsForty-one recipients with severe T1D who underwent PTx were retrospectively reviewed. The psoas muscle mass index (PMI) and intramuscular adipose tissue content (IMAC), as determined by preoperative computed tomography, were substituted for the quantity and quality of skeletal muscle for the definition of sarcopenia, respectively. Gender-specific quartiles were generated, and PMI lower than the first quantile or IMAC higher than the third quantile was considered to represent sarcopenia. The postoperative outcomes included postoperative surgical complications and pancreas graft survival.ResultsSarcopenia was identified in 11 recipients according to both the PMI and IMAC stratifications. The multivariate analyses revealed that high IMAC was independently associated with the development of postoperative surgical complications (odds ratio, 9.35; p = 0.016). In addition, the recipients with high IMAC showed unfavorable graft survival compared to those with normal IMAC (log-rank test; p = 0.038). In contrast, low PMI was not significantly associated with the postoperative outcomes.ConclusionsOur data suggested that preoperative sarcopenia, especially a decline in the quality of skeletal muscle, predicted poorer postoperative outcomes in T1D recipients undergoing PTx.