Chemical Biopsy for GNMT as Noninvasive and Tumorigenesis-Relevant Diagnosis of Liver Cancer.

Early diagnosis of hepatocellular carcinoma (HCC) is difficult; the lack of convenient biomarker-based diagnostic modalities renders high-risk HCC patients burdened by life-long periodical examinations. Here, a new chemical biopsy approach was developed for noninvasive diagnosis of HCC using urine samples. Bioinformatic screening for tumor suppressors yielded glycine N-methyltransferase (GNMT) as a biomarker with clinical relevance to HCC tumorigenesis. A liquid chromatography-mass spectrometry (LC-MS)-based chemical biopsy detecting nonradioactive 13C-sarcosine from 13C-glycine was designed to noninvasively assess liver GNMT activity extrahepatically. 13C-Sarcosine showed a strong correlation with GNMT in normal and cancerous liver cells. In an autochthonous animal model developing visible cancer nodules at 17 weeks, the urinary 13C-sarcosine chemical biopsy exhibited notable changes as early as 8 weeks, showing significant correlations with liver GNMT and molecular pathological changes. Our chemical biopsy approach should facilitate early and noninvasive diagnosis of HCC, with direct relevance to tumorigenesis, which can be straightforwardly applied to other diseases.

Artemisiae Iwayomogii Herba Inhibits Growth, Motility, and the PI3K/AKT/mTOR Signaling Pathway in Hepatocellular Carcinoma Cells.

Artemisia gmelinii (Artemisia iwayomogi) has been used in traditional medicine to cure various infectious diseases such as cholecystitis, hepatitis, and jaundice. In this study, the Artemisiae Iwayomogii Herba ethanol extract was investigated for the ability to inhibit growth of hepatocellular carcinoma and its underlying mechanism involved. The antiproliferative effect of Artemisiae Iwayomogii Herba ethanol extract was evaluated using cell viability and proliferation assays. The effect of Artemisiae Iwayomogii Herba ethanol extract on apoptosis was measured using western blotting, terminal deoxynucleotidyl transferase-mediated dUTP-biotin end labeling staining, JC-1 staining, cytochrome c release, immunohistochemistry, and immunofluorescence in ex vivo mouse xenografts. Artemisiae Iwayomogii Herba ethanol extract inhibited hepatocellular carcinoma cell growth and proliferation in a dose-dependent manner. The apoptotic effect of Artemisiae Iwayomogii Herba ethanol extract was observed via increased levels of cleaved caspase-3 and cleaved PARP, as well as elevated numbers of terminal deoxynucleotidyl transferase-mediated dUTP-biotin end labeling-positive apoptotic cells. Artemisiae Iwayomogii Herba ethanol extract also decreased XIAP and Mcl-1 expression via loss of mitochondrial membrane potential. Additionally, Artemisiae Iwayomogii Herba ethanol extract inhibited hepatocellular carcinoma cell invasion and migration. In the ex vivo model, Artemisiae Iwayomogii Herba ethanol extract significantly inhibited tumor cell proliferation and increased the number of apoptotic cells with more activated cleaved caspase-3. A mechanistic study revealed that Artemisiae Iwayomogii Herba ethanol extract effectively suppressed the PI3K/AKT/mTOR signaling pathway in hepatocellular carcinoma cells. Our findings demonstrate that Artemisiae Iwayomogii Herba ethanol extract can efficiently induce apoptosis and inhibit the growth, migration, and invasion of human hepatocellular carcinoma cells, and simultaneously block PI3K/AKT/mTOR pathway. We therefore suggest Artemisiae Iwayomogii Herba ethanol extract as a novel natural agent for prevention and therapy of hepatocellular carcinoma.

Systemic and Local Metabolic Alterations in Sleep-Deprivation-Induced Stress: A Multiplatform Mass-Spectrometry-Based Lipidomics and Metabolomics Approach.

Sleep deprivation (SD) is known to be associated with metabolic disorders and chronic diseases. Complex metabolic alterations induced by SD at omics scale and the associated biomarker candidates have been proposed. However, in vivo systemic and local metabolic shift patterns of the metabolome and lipidome in acute and chronic partial SD models remain to be elucidated. In the present study, the serum, hypothalamus, and hippocampus CA1 of sleep-deprived rats (SD rats) from acute and chronic sleep restriction models were analyzed using three different omics platforms for the discovery and mechanistic assessment of systemic and local SD-induced dysregulated metabolites. We found a similar pattern of systemic metabolome alterations between two models, for which the area under the curve (AUC) of receiver operating characteristic curves was AUC = 0.847 and 0.930 with the pseudotargeted and untargeted metabolomics approach, respectively. However, SD-induced systemic lipidome alterations were significantly different and appeared to be model-dependent (AUC = 0.374). Comprehensive pathway analysis of the altered lipidome and metabolome in the hypothalamus indicated the abnormal behavior of eight metabolic and lipid metabolic pathways. The metabolic alterations of the hippocampus CA1 was subtle in two SD models. Collectively, these results extend our understanding of the quality of sleep and suggest metabolic targets in developing diagnostic biomarkers for better SD control.

Melatonin Synergizes with Sorafenib to Suppress Pancreatic Cancer via Melatonin Receptor and PDGFR-ß/STAT3 Pathway.

BACKGROUND/AIMS: Pancreatic ductal adenocarcinoma (PDAC) is one of the most lethal malignant tumors with poor prognosis. Conventional chemotherapies including gemcitabine have failed owing to weak response and side effects. Hence novel treatment regimens are urgently needed to improve the therapeutic efficacy. In this study, we aimed to assess the anticancer activity of melatonin and sorafenib as a novel therapy against PDAC. METHODS: We used various apoptosis assay and PDAC xenograft model to assess anticancer effect in vitro and in vivo. We applied phospho-receptor tyrosine kinase (RTK) array and phospho-tyrosine kinase array to explore the mechanism of the combined therapy. Western blotting, proximity ligation assay, and immunoprecipitation assay were also performed for validation. RESULTS: Melatonin synergized with sorafenib to suppress the growth of PDAC both in vitro and in vivo. The effect was due to increased apoptosis rate of PDAC cells that was accompanied by mitochondria dysfunction. The enhanced anticancer efficacy by the co-treatment could be explained by blockade of PDGFR-ß/STAT3 signaling pathway and melatonin receptor (MT)-mediated STAT3. CONCLUSIONS: Melatonin reinforces the anticancer activity of sorafenib by downregulation of PDGFR-ß/STAT3 signaling pathway and melatonin receptor (MT)-mediated STAT3. The combination of the two agents might be a potential therapeutic strategy for treating PDAC.

Artemisia Capillaris leaves inhibit cell proliferation and induce apoptosis in hepatocellular carcinoma.

BACKGROUND: Natural product is one of the most important sources of drugs used in pharmaceutical therapeutics. Artemisia capillaris has been traditionally used as a hepatoprotective and anti-inflammatory agent. In this study, we extracted an ethanol fraction (LAC117) from the dried leaves of Artemisia capillaris and identified its anticancer activity and mechanism of action against hepatocellular carcinoma (HCC). METHODS: Anti-proliferative effect of LAC117 was evaluated by MTT assay and BrdU assay. The apoptotic effect of LAC117 on the expression of cleaved PARP and cleaved caspase-3 was evaluated by Western blot and immunohistochemistry from in vivo mouse xenograft, respectively. RESULTS: We found that LAC117 strongly suppressed the growth and proliferation of human HCC cell lines (HepG2 and Huh7). Induction of apoptosis was evidenced by the increases of cleaved caspase-3 and PARP as well as TUNEL-positive cells. Additionally, the pro-apoptotic effect of LAC117 was observed by a decrease in the expression of the XIAP and an increase in cytochrome c releases via mitochondrial membrane potential. Moreover, it significantly inhibited PI3K/AKT pathway in HCC in vivo and in vitro. LAC117 suppressed tumor growth in an ex vivo model as well as in vivo mouse xenograft by inducing apoptosis and inhibiting tumor cell proliferation. CONCLUSIONS: The present study highlights that LAC117 could not only efficiently induce apoptosis, but also inhibit the growth of human HCC cells by blocking the PI3K/AKT signaling pathway, suggesting that LAC117 would be a potentially useful drug candidate against HCC.

An ethyl acetate fraction of Artemisia capillaris (ACE-63) induced apoptosis and anti-angiogenesis via inhibition of PI3K/AKT signaling in hepatocellular carcinoma.

In cancer treatment, herbal medicines may be a good choice because of the reduced risk of adverse side effects. Artemisia capillaris has been recognized as a promising candidate due to its hepatoprotective effects. Herein, we investigated whether A. capillaris-derived fraction (ACE-63) could inhibit the progression of hepatocellular carcinoma (HCC) and its underlying mechanism. In this study, ACE-63 effectively inhibited the growth and proliferation of HCC cells. ACE-63 induced apoptosis, as observed using Terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) staining, which was accompanied with increases in cleaved Poly (ADP-ribose) polymerase (PARP) and caspase-3 in HCC cells. Additionally, the pro-apoptotic effect of ACE-63 was demonstrated by a decrease in the expression of the X-linked inhibitor of apoptosis protein (XIAP) and survivin via a loss of mitochondrial membrane potential. In an ex vivo model, ACE-63 significantly inhibited tumor cell growth and induced apoptosis by increasing the expression of cleaved caspase-3 and DNA fragmentation. In addition, ACE-63 decreased the expression of hypoxia-inducible factor-1α and vascular endothelial growth factor and inhibited tube formation of human umbilical vein endothelial cells. A mechanistic study revealed that ACE-63 effectively suppressed the PI3K/AKT/mTOR signaling pathways, which were observed as a target signaling by phosphokinase array. Taken together, our findings demonstrate that ACE-63 could not only efficiently induce apoptosis but also inhibit the growth/angiogenesis of human HCC cells by blocking the PI3K/AKT/mTOR signaling pathway, suggesting that ACE-63 may be a new chemotherapeutic candidate against HCC.

Tumor Stroma as a Therapeutic Target for Pancreatic Ductal Adenocarcinoma.

Pancreatic ductal adenocarcinoma (PDAC) has a poor prognosis owing to its desmoplastic stroma. Therefore, therapeutic strategies targeting this tumor stroma should be developed. In this study, we describe the heterogeneity of cancer-associated fibroblasts (CAFs) and their diverse roles in the progression, immune evasion, and resistance to treatment of PDAC. We subclassified the spatial distribution and functional activity of CAFs to highlight their effects on prognosis and drug delivery. Extracellular matrix components such as collagen and hyaluronan are described for their roles in tumor behavior and treatment outcomes, implying their potential as therapeutic targets. We also discussed the roles of extracellular matrix (ECM) including matrix metalloproteinases and tissue inhibitors in PDAC progression. Finally, we explored the role of the adaptive and innate immune systems in shaping the PDAC microenvironment and potential therapeutic strategies, with a focus on immune cell subsets, cytokines, and immunosuppressive mechanisms. These insights provide a comprehensive understanding of PDAC and pave the way for the development of prognostic markers and therapeutic interventions.

Potential role of ANGPTL4 in cancer progression, metastasis, and metabolism: a brief review.

Angiopoietin-like 4 (ANGPTL4) has been identified as an adipokine involved in several non-metabolic and metabolic diseases, including angiogenesis, glucose homeostasis, and lipid metabolism. To date, the role of ANGPTL4 in cancer growth and progression, and metastasis, has been variable. Accumulating evidence suggests that proteolytic processing and posttranslational modifications of ANGPTL4 can significantly alter its function, and may contribute to the multiple and conflicting roles of ANGPTL4 in a tissue-dependent manner. With the growing interest in ANGPTL4 in cancer diagnosis and therapy, we aim to provide an up-to-date review of the implications of ANGPTL4 as a biomarker/oncogene in cancer metabolism, metastasis, and the tumor microenvironment (TME). In cancer cells, ANGPTL4 plays an important role in regulating metabolism by altering intracellular glucose, lipid, and amino acid metabolism. We also highlight the knowledge gaps and future prospect of ANGPTL4 in lymphatic metastasis and perineural invasion through various signaling pathways, underscoring its importance in cancer progression and prognosis. Through this review, a better understanding of the role of ANGPTL4 in cancer progression within the TME will provide new insights into other aspects of tumorigenesis and the potential therapeutic value of ANGPTL4.

Acetyl-CoA synthetase 2 contributes to a better prognosis for liver cancer by switching acetate-glucose metabolism.

Acetyl-CoA synthetase 2 (ACSS2)-dependent acetate usage has generally been associated with tumorigenesis and increased malignancy in cancers under nutrient-depleted conditions. However, the nutrient usage and metabolic characteristics of the liver differ from those of other organs; therefore, the mechanism of ACSS2-mediated acetate metabolism may also differ in liver cancer. To elucidate the underlying mechanisms of ACSS2 in liver cancer and acetate metabolism, the relationships between patient acetate uptake and metabolic characteristics and between ACSS2 and tumor malignancies were comprehensively studied in vitro, in vivo and in humans. Clinically, we initially found that ACSS2 expression was decreased in liver cancer patients. Moreover, PET-CT imaging confirmed that lower-grade cancer cells take up more 11C-acetate but less 18F-fluorodeoxyglucose (18F-FDG); however, this trend was reversed in higher-grade cancer. Among liver cancer cells, those with high ACSS2 expression avidly absorbed acetate even in a glucose-sufficient environment, whereas those with low ACSS2 expression did not, thereby showing correlations with their respective ACSS2 expression. Metabolomic isotope tracing in vitro and in vivo revealed greater acetate incorporation, greater lipid anabolic metabolism, and less malignancy in high-ACSS2 tumors. Notably, ACSS2 downregulation in liver cancer cells was associated with increased tumor occurrence in vivo. In human patient cohorts, patients in the low-ACSS2 subgroup exhibited reduced anabolism, increased glycolysis/hypoxia, and poorer prognosis. We demonstrated that acetate uptake by ACSS2 in liver cancer is independent of glucose depletion and contributes to lipid anabolic metabolism and reduced malignancy, thereby leading to a better prognosis for liver cancer patients.

SB365, Pulsatilla saponin D, targets c-Met and exerts antiangiogenic and antitumor activities.

SB365, Pulsatilla saponin D isolated from the root of Pulsatilla koreana, has exhibited potential beneficial effects as a chemopreventive agent for critical health conditions including cancer. However, the molecular mechanisms underlying the activity of SB365 remain unknown. Here, we examined anticancer efficacy of SB365 against gastric cancer and its mechanism of action. SB365 effectively inhibited the growth of gastric cancer cells. Its apoptotic effect was accompanied by increased evidence of cleaved caspase-3 and poly(ADP ribose) polymerase. To elucidate the anticancer mechanism of SB365, we used an array of 42 different receptor tyrosine kinases (RTKs). Of the 42 different phospho-RTKs, SB365 strongly inhibited expression of activated c-mesenchymal-epithelial transition factor (c-Met) in gastric cancer cells. Also, the activation of the c-Met signal cascade components, including Akt and mammalian target of rapamycin, was inhibited by SB365 in a dose-dependent manner. In angiogenesis studies, SB365 inhibited tube formation in hepatocyte growth factor (HGF)-induced human umbilical vein endothelial cells and suppressed microvessel sprouting from the rat aortic ring, ex vivo, and blood vessel formation in the Matrigel plug assay in mice. In xenograft animal models, SB365 significantly delayed tumor growth in a dose-dependent manner. In tumor tissue, SB365 suppressed c-Met signaling, proliferation and angiogenesis and induced apoptosis. These findings suggest that SB365 docks at an allosteric site on c-Met and thereby targets c-Met signaling pathway, cell growth/angiogenesis inhibition and apoptosis induction. Therefore, SB365 may be a novel drug candidate for the treatment of gastric cancer.

Synergistic Renoprotective Effect of Melatonin and Zileuton by Inhibition of Ferroptosis via the AKT/mTOR/NRF2 Signaling in Kidney Injury and Fibrosis.

According to recent evidence, ferroptosis is a major cell death mechanism in the pathogenesis of kidney injury and fibrosis. Despite the renoprotective effects of classical ferroptosis inhibitors, therapeutic approaches targeting kidney ferroptosis remain limited. In this study, we assessed the renoprotective effects of melatonin and zileuton as a novel therapeutic strategy against ferroptosis-mediated kidney injury and fibrosis. First, we identified RSL3-induced ferroptosis in renal tubular epithelial HK-2 and HKC-8 cells. Lipid peroxidation and cell death induced by RSL3 were synergistically mitigated by the combination of melatonin and zileuton. Combination treatment significantly downregulated the expression of ferroptosis-associated proteins, 4-HNE and HO-1, and upregulated the expression of GPX4. The expression levels of p-AKT and p-mTOR also increased, in addition to that of NRF2 in renal tubular epithelial cells. When melatonin (20 mg/kg) and zileuton (20 mg/kg) were administered to a unilateral ureteral obstruction (UUO) mouse model, the combination significantly reduced tubular injury and fibrosis by decreasing the expression of profibrotic markers, such as α-SMA and fibronectin. More importantly, the combination ameliorated the increase in 4-HNE levels and decreased GPX4 expression in UUO mice. Overall, the combination of melatonin and zileuton was found to effectively ameliorate ferroptosis-related kidney injury by upregulating the AKT/mTOR/ NRF2 signaling pathway, suggesting a promising therapeutic strategy for protection against ferroptosis-mediated kidney injury and fibrosis.

A combination of BR101801 and venetoclax enhances antitumor effect in DLBCL cells via c-Myc/Bcl-2/Mcl-1 triple targeting.

Double hit diffuse large B-cell lymphoma (DLBCL) with rearrangement and overexpression of both c-Myc and Bcl-2 responds poorly to standard R-CHOP therapy. In a recent phase I study, Venetoclax (ABT-199) targeting Bcl-2 also exhibited disappointing response rates in patients with relapsed/refractory DLBCL, suggesting that targeting only Bcl-2 is not sufficient for achieving successful efficacy due to the concurrent oncogenic function of c-Myc expression and drug resistance following an increase in Mcl-1. Therefore, co-targeting c-Myc and Mcl-1 could be a key combinatorial strategy to enhance the efficacy of Venetoclax. In this study, BR101801 a novel drug for DLBCL, effectively inhibited DLBCL cell growth/proliferation, induced cell cycle arrest, and markedly inhibited G0/G1 arrest. The apoptotic effect of BR101801 was also observed by increased Cytochrome C, cleaved PARP, and Annexin V-positive cell populations. This anti-cancer effect of BR101801 was confirmed in animal models, where it effectively inhibited tumor growth by reducing the expression of both c-Myc and Mcl-1. Furthermore, BR101801 exhibited a significant synergistic antitumor effect even in late xenograft models when combined with Venetoclax. Our data strongly suggest that c-Myc/Bcl-2/Mcl-1 triple targeting through a combination of BR101801 and Venetoclax could be a potential clinical option for double-hit DLBCL.

Vactosertib, TGF-ß receptor I inhibitor, augments the sensitization of the anti-cancer activity of gemcitabine in pancreatic cancer.

Pancreatic ductal adenocarcinoma (PDAC) exhibits a pronounced extracellular matrix (ECM)-rich response, which is produced by an excessive amount of transforming growth factor ß (TGF-ß), resulting in tumor progression and metastasis. In addition, TGF-ß signaling contributes to rapidly acquired resistance and incomplete response to gemcitabine. Recently, selective inhibitors of the TGF-ß signaling pathway have shown promise in PDAC treatment, particularly as an option for augmenting responses to chemotherapy. Here, we investigated the synergistic anticancer effects of a small-molecule TGF-ß receptor I kinase inhibitor (vactosertib/EW-7197) in the presence of gemcitabine, and its mechanism of action in pancreatic cancer. Vactosertib sensitized pancreatic cancer cells to gemcitabine by synergistically inhibiting their viability. Importantly, the combination of vactosertib and gemcitabine significantly attenuated the expression of major ECM components, including collagens, fibronectin, and α-SMA, in pancreatic cancer compared with gemcitabine alone. This resulted in potent induction of mitochondrial-mediated apoptosis, gemcitabine-mediated cytotoxicity, and inhibition of tumor ECM by vactosertib. Additionally, the combination decreased metastasis through inhibition of migration and invasion, and exhibited synergistic anti-cancer activity by inhibiting the TGF-ß/Smad2 pathway in pancreatic cancer cells. Furthermore, co-treatment significantly suppressed tumor growth in orthotopic models. Therefore, our findings demonstrate that vactosertib synergistically increased the antitumor activity of gemcitabine via inhibition of ECM component production by inhibiting the TGF-ß/Smad2 signaling pathway. This suggests that the combination of vactosertib and gemcitabine may be a potential treatment option for patients with pancreatic cancer.

SB365 inhibits angiogenesis and induces apoptosis of hepatocellular carcinoma through modulation of PI3K/Akt/mTOR signaling pathway.

Identification of small molecules that safely inhibit cancer progression is critical for cancer therapeutics. Saponins exhibit cytostatic and cytotoxic activity against various cancer cells, but the mechanism is not well understood. Here, we investigated whether saponin D (designated SB365), an active component isolated from Pulsatilla koreana, could inhibit the progression of hepatocellular carcinoma (HCC) and considered its mechanism. SB365 strongly suppressed the growth of HCC cells in a dose-dependent manner and induced apoptosis by increasing the proportion of sub G1 apoptotic cells from 8% to 21% through induction of expression of Bax and cleaved caspase-3. In addition, SB365 exhibited potent anti-angiogenic activity and decreased the expression of hypoxia-inducible factor-1α (HIF-1α) and vascular endothelial growth factor, a key molecule for angiogenesis. Furthermore, SB365 suppressed the tube formation and migration of HUVEC, as well as in vivo neovascularization in a mouse Matrigel plug assay. In vivo study showed that SB365 significantly inhibited tumor growth in an HCC xenograft model, inducing apoptosis by increasing the expression of the cleaved caspase-3 and DNA fragmentation. The expressions of vascular endothelial growth factor and CD34 in the tumor tissue were decreased by SB365 treatment. In examining its mechanism, SB365 was found to effectively suppress the phosphorylation of PI3K downstream factors, such as Akt, mTOR and p70S6K both in vitro and in vivo. Our study demonstrates that SB365 not only induces apoptosis but also inhibits cell growth and angiogenesis through modulation of the PI3K/Akt/mTOR pathway in human HCC. We suggest that SB365 may be a new chemotherapeutic candidate against HCC.

Human bone marrow-derived clonal mesenchymal stem cells inhibit inflammation and reduce acute pancreatitis in rats.

BACKGROUND & AIMS: Acute pancreatitis (AP) has a high mortality rate; repetitive AP induces chronic AP and pancreatic adenocarcinoma. Mesenchymal stem cells (MSCs) have immunoregulatory effects and reduce inflammation. We developed a protocol to isolate human bone marrow-derived clonal MSCs (hcMSCs) from bone marrow aspirate and investigated the effects of these cells in rat models of mild and severe AP. METHODS: Mild AP was induced in Sprague-Dawley rats by 3 intraperitoneal injections of cerulein (100 µg/kg), given at 2-hour intervals; severe AP was induced by intraparenchymal injection of 3% sodium taurocholate solution. hcMSCs were labeled with CM-1,1'-dioctadecyl-3,3,3'-tetramethylindo-carbocyanine perchloride and administered to rats through the tail vein. RESULTS: hcMSCs underwent self-renewal and had multipotent differentiation capacities and immunoregulatory functions. Greater numbers of infused hcMSCs were detected in pancreas of rats with mild and severe AP than of control rats. Infused hcMSCs reduced acinar-cell degeneration, pancreatic edema, and inflammatory cell infiltration in each model of pancreatitis. The hcMSCs reduced expression of inflammation mediators and cytokines in rats with mild and severe AP. hcMSCs suppressed the mixed lymphocyte reaction and increased expression of Foxp3(+) (a marker of regulatory T cells) in cultured rat lymph node cells. Rats with mild or severe AP that were given infusions of hcMSCs had reduced numbers of CD3(+) T cells and increased expression of Foxp3(+) in pancreas tissues. CONCLUSIONS: hcMSCs reduced inflammation and damage to pancreatic tissue in a rat model of AP; they reduced levels of cytokines and induced numbers of Foxp3(+) regulatory T cells. hcMSCs might be developed as a cell therapy for pancreatitis.

Discovery of a novel NUAK1 inhibitor against pancreatic cancer.

The novel (nua) kinase family 1 (NUAK1) is an AMPK-related kinase and its expression is associated with tumor malignancy and poor prognosis in several types of cancer, suggesting its potential as a target for cancer therapy. Therefore, the development of NUAK1-targeting inhibitors could improve therapeutic outcomes in cancer. We synthesized KI-301670, a novel NUAK1 inhibitor, and assessed its anticancer effects and mechanism of action in pancreatic cancer. It effectively inhibited pancreatic cancer growth and proliferation, and induced cell cycle arrest, markedly G0/G1 arrest, by increasing the expression of p27 and decreasing expression of p-Rb and E2F1. Additionally, the apoptotic effect of KI-301670 was observed by an increase in cleaved PARP, TUNEL-positive cells, and annexin V cell population, as well as the release of cytochrome c via the loss of mitochondrial membrane potential. KI-301670 inhibited the migration and invasion of pancreatic cancer cells. Mechanistically, KI-301670 effectively inhibited the PI3K/AKT pathway in pancreatic cancer cells. Furthermore, it significantly attenuated tumor growth in a mouse xenograft tumor model. Our results demonstrate that a novel NUAK1 inhibitor, KI-301670, exerts anti-tumor effects by directly suppressing cancer cell growth by affecting the PI3K/AKT pathway, suggesting that it could be a novel therapeutic candidate for pancreatic cancer treatment.

Combination Therapy of the Active KRAS-Targeting Antibody inRas37 and a PI3K Inhibitor in Pancreatic Cancer.

KRAS activating mutations, which are present in more than 90% of pancreatic cancers, drive tumor dependency on the RAS/mitogen-activated protein kinase (MAPK) and phosphoinositide 3-kinase (PI3K)/AKT signaling pathways. Therefore, combined targeting of RAS/MAPK and PI3K/AKT signaling pathways may be required for optimal therapeutic effect in pancreatic cancer. However, the therapeutic efficacy of combined MAPK and PI3K/AKT signaling target inhibitors is unsatisfactory in pancreatic cancer treatment, because it is often accompanied by MAPK pathway reactivation by PI3K/AKT inhibitor. Therefore, we developed an inRas37 antibody, which directly targets the intra-cellularly activated GTP-bound form of oncogenic RAS mutation and investigated its synergistic effect in the presence of the PI3K inhibitor BEZ-235 in pancreatic cancer. In this study, inRas37 remarkably increased the drug response of BEZ-235 to pancreatic cancer cells by inhibiting MAPK reactivation. Moreover, the co-treatment synergistically inhibited cell proliferation, migration, and invasion and exhibited synergistic anticancer activity by inhibiting the MAPK and PI3K pathways. The combined administration of inRas37and BEZ-235 significantly inhibited tumor growth in mouse models. Our results demonstrated that inRas37 synergistically increased the antitumor activity of BEZ-235 by inhibiting MAPK reactivation, suggesting that inRas37 and BEZ-235 co-treatment could be a potential treatment approach for pancreatic cancer patients with KRAS mutations.

Comparative analysis of mesenchymal stem cells cultivated in serum free media.

Stem cells are attractive candidates for the regeneration of tissue and organ. Mesenchymal stem cells (MSCs) have been extensively investigated for their potential applications in regenerative medicine and cell therapy. For developing effective stem cell therapy, the mass production of consistent quality cells is required. The cell culture medium is the most critical aspect of the mass production of qualified stem cells. Classically, fetal bovine serum (FBS) has been used as a culture supplement for MSCs. Due to the undefined and heterologous composition of animal origin components in FBS, efforts to replace animal-derived components with non-animal-derived substances led to safe serum free media (SFM). Adipose derived mesenchymal stem cells (ADSCs) cultivated in SFM provided a more stable population doubling time (PDT) to later passage and more cells in a shorter time compared to FBS containing media. ADSCs cultivated in SFM had lower cellular senescence, lower immunogenicity, and higher genetic stability than ADSCs cultivated in FBS containing media. Differential expression analysis of mRNAs and proteins showed that the expression of genes related with apoptosis, immune response, and inflammatory response were significantly up-regulated in ADSCs cultivated in FBS containing media. ADSCs cultivated in SFM showed similar therapeutic efficacy in an acute pancreatitis mouse model to ADSCs cultivated in FBS containing media. Consideration of clinical trials, not only pre-clinical trial, suggests that cultivation of MSCs using SFM might offer more safe cell therapeutics as well as repeated administration due to low immunogenicity.

A novel DDR1 inhibitor enhances the anticancer activity of gemcitabine in pancreatic cancer.

Pancreatic ductal adenocarcinoma (PDAC) is an extracellular matrix (ECM)-rich carcinoma, which promotes chemoresistance by inhibiting drug diffusion into the tumor. Discoidin domain receptor 1 (DDR1) increases tumor progression and drug resistance by binding to collagen, a major component of tumor ECM. Therefore, DDR1 inhibition may be helpful in cancer therapeutics by increasing drug delivery efficiency and improving drug sensitivity. In this study, we developed a novel DDR1 inhibitor, KI-301690 and investigated whether it could improve the anticancer activity of gemcitabine, a cytotoxic agent widely used for the treatment of pancreatic cancer. KI-301690 synergized with gemcitabine to suppress the growth of pancreatic cancer cells. Importantly, its combination significantly attenuated the expression of major tumor ECM components including collagen, fibronectin, and vimentin compared to gemcitabine alone. Additionally, this combination effectively decreased mitochondrial membrane potential (MMP), thereby inducing apoptosis. Further, the combination synergistically inhibited cell migration and invasion. The enhanced anticancer efficacy of the co-treatment could be explained by the inhibition of DDR1/PYK2/FAK signaling, which significantly reduced tumor growth in a pancreatic xenograft model. Our results demonstrate that KI-301690 can inhibit aberrant ECM expression by DDR1/PYK2/FAK signaling pathway blockade and attenuation of ECM-induced chemoresistance observed in desmoplastic pancreatic tumors, resulting in enhanced antitumor effect through effective induction of gemcitabine apoptosis.

A Serum Marker for Early Pancreatic Cancer With a Possible Link to Diabetes.

BACKGROUND: Pancreatic cancer (PC) has a grim prognosis, and an early diagnostic biomarker has been highly desired. The molecular link between diabetes and PC has not been well established. METHODS: Bioinformatics screening was performed for a serum PC marker. Experiments in cell lines (5 PC and 1 normal cell lines), mouse models, and human tissue staining (37 PC and 10 normal cases) were performed to test asprosin production from PC. Asprosin's diagnostic performance was tested with serums from multi-center cohorts (347 PC, 209 normal, and 55 additional diabetic patients) and evaluated according to PC status, stages, and diabetic status, which was compared with that of CA19-9. RESULTS: Asprosin, a diabetes-related hormone, was found from the bioinformatics screening, and its production from PC was confirmed. Serum asprosin levels from multi-center cohorts yielded an age-adjusted diagnostic area under the curve (AUC) of 0.987 (95% confidence interval [CI] = 0.961 to 0.997), superior to that of CA19-9 (AUC = 0.876, 95% CI = 0.847 to 0.905), and a cut-off of 7.18 ng/mL, at which the validation set exhibited a sensitivity of 0.957 and a specificity of 0.924. Importantly, the performance was maintained in early-stage and non-metastatic PC, consistent with the tissue staining. A slightly lower performance against additional diabetic patients (n = 55) was restored by combining asprosin and CA19-9 (AUC = 0.985, 95% CI = 0.975 to 0.995). CONCLUSIONS: Asprosin is presented as an early-stage PC serum marker that may provide clues for PC-induced diabetes. Larger prospective clinical studies are warranted to solidify its utility.