Mouse Model Of Liver Metastasis Research Articles

Tumor Contrast Enhancement and Whole-Body Elimination of the Manganese-Based Magnetic Resonance Imaging Contrast Agent Mn-PyC3A.

The goals of this study were to compare the efficacy of the new manganese-based magnetic resonance imaging (MRI) contrast agent Mn-PyC3A to the commercial gadolinium-based agents Gd-DOTA and to Gd-EOB-DTPA to detect tumors in murine models of breast cancer and metastatic liver disease, respectively, and to quantify the fractional excretion and elimination of Mn-PyC3A in rats. T1-weighted contrast-enhanced MRI with 0.1 mmol/kg Mn-PyC3A was compared with 0.1 mmol/kg Gd-DOTA in a breast cancer mouse model (n = 8) and to 0.025 mmol/kg Gd-EOB-DTPA in a liver metastasis mouse model (n = 6). The fractional excretion, 1-day biodistribution, and 7-day biodistribution in rats after injection of 2.0 mmol/kg [Mn]Mn-PyC3A or Gd-DOTA were quantified by Mn gamma counting or Gd elemental analysis. Imaging data were compared with a paired t test; biodistribution data were compared with an unpaired t test. The postinjection-preinjection increases in tumor-to-muscle contrast-to-noise ratio (ΔCNR) 3 minutes after injection of Mn-PyC3A and Gd-DOTA (mean ± standard deviation) were 17 ± 3.8 and 20 ± 4.4, respectively (P = 0.34). Liver-to-tumor ΔCNR values at 8 minutes postinjection of Mn-PyC3A and Gd-EOB-DTPA were 28 ± 9.0 and 48 ± 23, respectively (P = 0.11). Mn-PyC3A is eliminated with 85% into the urine and 15% into the feces after administration to rats. The percentage of the injected doses (%ID) of Mn and Gd recovered in tissues after 1 day were 0.32 ± 0.12 and 0.57 ± 0.12, respectively (P = 0.0030), and after 7 days were 0.058 ± 0.051 and 0.19 ± 0.052, respectively (P < 0.0001). Mn-PyC3A provides comparable tumor contrast enhancement to Gd-DOTA in a mouse breast cancer model and is more completely eliminated than Gd-DOTA; partial hepatobiliary elimination of Mn-PyC3A enables conspicuous delayed phase visualization of liver metastases.

E2A attenuates tumor-initiating capacity of colorectal cancer cells via the Wnt/beta-catenin pathway

BackgroundThe E2A gene, which encodes two basic helix–loop–helix transcription factors, E12 and E47, regulates colorectal cancer progression and epithelial-mesenchymal transition. However, whether E2A regulates the tumor-initiating capacity of colorectal cancer is unclear. Thus, we have studied E2A expression in the initiation of colorectal cancer in vivo and in vitro.MethodsImmunohistochemistry and immunoblot were performed to determine protein levels of E2A in colorectal cancer specimens and cells. RNAi was employed to downregulate E2A expression, and the subsequent change in protein level was evaluated by immunoblot. Sphere-forming assay and enumeration of liver metastasis in mouse models were used to identify the tumor formation ability of colorectal cancer cells.ResultsE2A expression in colorectal cancer clinical specimens was inversely associated with patients’ progression-free survival. Functional studies demonstrated that E2A significantly decreased tumor formation in vitro and in vivo. Furthermore, nuclear translocation of beta-catenin and activation of the Wnt/beta-catenin pathway occurred after suppression of E2A in colorectal cancer cells. FoxM1 was identified as a down-stream target by mRNA microarray, implying that FoxM1 plays a main role in determining how E2A regulates the tumor-initiating capacity of colorectal cancer.ConclusionE2A suppresses tumor-initiating capacity by targeting the FoxM1-Wnt/β-catenin pathway.

PLEK2 promotes gallbladder cancer invasion and metastasis through EGFR/CCL2 pathway

BackgroundGallbladder cancer (GBC) is an extremely malignant tumor with a high mortality rate. Little is known about its invasion and metastasis mechanism so far.MethodsTo identify the driver genes in GBC metastasis, we performed a mRNA microarray of metastatic GBC and paired non-tumor samples, and found PLEK2 was markedly upregulated in GBC tissues. Next, the expression of PLEK2 in GBC were examined in a larger cohort of patients by qRT-PCR, western blot and IHC staining. The clinicopathologic correlation of PLEK2 was determined by statistical analyses. The biological involvement of PLEK2 in GBC metastasis and the underlying mechanisms were investigated.ResultsIn this study, we found that PLEK2 had higher expression in GBC tumor tissues compared to non-cancerous adjacent tissues and cholecystolithiasis tissues. The clinicopathologic analyses showed PLEK2 expression was positively correlated with tumor TNM stage, distant metastasis and PLEK2 was an independent predictor of overall survival (OS) in GBC patients. The cellular function assays showed PLEK2 promoted GBC cells migration, invasion and liver metastasis in mouse model via the regulation of epithelial-mesenchymal transition (EMT) process. Our mass spectrum and co-immunoprecipitation (co-IP) assays demonstrated that PLEK2 could interact with the kinase domain of EGFR and suppress EGFR ubiquitination mediated by c-CBL, leading to constitutive activation of EGFR signaling. Furthermore, RNA-sequencing and qRT-PCR results demonstrated chemokine (C-C motif) ligand 2 (CCL2), a target gene downstream of PLEK2/EGFR signaling, mediated the motility-promoting function of PLEK2.ConclusionsOn the basis of these collective data, we propose that PLEK2 promotes the invasion and metastasis of GBC by EGFR/CCL2 pathway and PLEK2 can serve as a potential therapeutic target for GBC treatment.

Programmed cell death ligand 1 (PD-L1) blockade attenuates metastatic colon cancer growth in cAMP-response element-binding protein (CREB)-binding protein (CBP)/β-catenin inhibitor-treated livers.

ABSTRACTImmune checkpoint blockade with specific antibodies can accelerate anti-tumor immunity, resulting in clinical responses in patients with various types of cancer. However, these antibodies achieve only partial tumor regression. Thus, a wide variety of treatment combinations based on programmed death-ligand 1 (PD-L1) pathway inhibition are under development to enhance such therapeutic effects. In this study, the effects of combination treatment using PRI-724, a selective inhibitor of CBP/β-catenin, and an anti-PD-L1 antibody were examined in a mouse model of colon cancer liver metastasis. Mice were inoculated with SL4 colon cancer cells to produce metastatic liver tumors. The combination treatment resulted in regression of tumor growth, whereas monotherapy with each treatment individually failed to exhibit any anti-tumor activity. In addition, co-administration of the inhibitor and antibody induced CD8+CD44lowCD62Llow cells and interferon (IFN)-γ production in CD8+ T-cells in the liver compared with that in control mice. Administration of an anti-CD8 antibody mitigated the anti-tumor effects of the combined treatment of PRI-724 and anti-PD-L1 antibody. In conclusion, targeting CBP/β-catenin, combined with PD-1/PD-L1 immune checkpoint blockade, shows potential as a new therapeutic strategy for treating liver metastasis during colon cancer.

NSD2 circular RNA promotes metastasis of colorectal cancer by targeting miR-199b-5p-mediated DDR1 and JAG1 signalling.

Liver metastasis is the main cause of death in patients with colorectal cancer (CRC). Here, we searched for CRC metastasis-associated circular RNA in a mouse model of liver metastasis of CRC by using RNA (transcriptome)-sequencing. We identified a novel and conserved circular RNA, circ-NSD2, functioning as a promoter of CRC metastasis. Circ-NSD2 expression was elevated in CRC tissues and was markedly increased in advanced stages or metastatic tumours of CRC patients. Gain-of-function and loss-of-function experiments demonstrated that circ-NSD2 promoted migration and metastasis of CRC in vitro and in vivo. Mechanistically, circ-NSD2 acted as a sponge for the tumour suppressor miR-199b-5p and activated DDR1 (discoidin domain receptor tyrosine kinase 1) and JAG1 (Jagged 1) genes, which synergistically helped with cell-matrix interaction, migration and metastasis of CRC cells. Taken together, our findings highlight a novel oncogenic function of circ-NSD2 and uncover a key mechanism for the circ-NSD2/miR-199b-5p/DDR1/JAG1 axis in CRC metastasis, which may serve as a prognostic factor and therapeutic target for antimetastatic therapy in CRC patients. Copyright © 2019 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.

Salinomycin Effectively Eliminates Cancer Stem-Like Cells and Obviates Hepatic Metastasis in Uveal Melanoma

Background: Uveal melanoma (UM) is the most common primary intraocular tumor. Approximately 50% patients with UM manifest single-organ metastasis to liver, which is the major and drect death reason in the UM patients owing to lack effective chemotherapeutic drugs. Consequently, there is a desperate need for new effective treatment modalities. Given that cancer stem-like cells (CSCs) are roots of metastasis, targeting CSCs may be a promising strategy to overcome hepatic metastasis in UM. Salinomycin, which has been identified as a selective inhibitor of CSCs in multiple types of cancer, may be an attractive agent to against CSCs thereby restrain hepatic metastasis in UM. The objective of the study is to explore the antitumor activity of salinomycin on UM and clarify its underlying mechanism. Methods: UM cells were treated with salinomycin, and its effects on cell proliferation, apoptosis, migration, invasion, CSCs population, and the related signal transduction pathways were determined. The in vivo antitumor activity of salinomycin was evaluated in the NOD/SCID UM xenograft model and intrasplenic transplantation liver metastasis mouse model. Findings: We found that salinomycin remarkably obviated growth and survival in UM cell lines and in a UM xenograft mouse model. Meanwhile, salinomycin significantly eliminated CSCs and efficiently hampered hepatic metastasis in UM liver metastasis mouse model. Mechanistically, Twist1 was fundamental for the salinomycin-enabled CSCs elimination and migration/invasion blockage in UM cells. Interpretation: These findings suggest that salinomycin may be a promising therapeutic agent for UM patients with hepatic metastasis. Funding Statement: This study was supported by grants from the Research Foundation of Education Bureau of Guangdong Province, China (Grant cxzd1103 to J. Pan), and Natural Science Foundation of Guangdong province (Grant 2015A030312014 to J. Pan). Present address of W.D., Y.W., and S.W.: Institute of Tumor Pharmacology, College of Pharmacy, Jinan University, Guangzhou, China. Declaration of Interests: The authors declare that they have no competing interests. Ethics Approval Statement: The animal studies were approved by the Sun Yat-sen University Institutional Animal Care and Use Committee.

Anti-claudin-4 extracellular domain antibody enhances the antitumoral effects of chemotherapeutic and antibody drugs in colorectal cancer.

Claudin-4 (CLDN4) is a major epithelial tight junction protein overexpressed in many cancers to maintain the tumor environment. In this report, we aimed to determine the efficacy of targeting CLDN4 in colorectal cancer (CRC) using an anti-CLDN4 extracellular domain antibody, 4D3. CLDN4 was upregulated in CRC metastatic foci. CLDN4 expression in CRC cells was reduced by upregulation of TNFα, which was induced by Clostridium perfringens enterotoxin produced by gut flora. In a nude mouse liver metastasis model, inhibition of metastasis was increased by combination treatment with 5-fluorouracil (FU) and 4D3 compared to that with 5-FU alone. Moreover, combination treatment with 4D3 and anti-epithelial growth factor receptor (EGFR) antibody C225 resulted in more pronounced inhibition of in vitro sphere formation and tumor growth in nude mice compared to that observed with C225 alone. Moreover, the time interval between the administration of 4D3 and that of C225 was important for maximizing the C225-induced inhibition of EGFR phosphorylation. In a nude mouse model, sequential treatment with 4D3 and C225 with a 6-h time interval resulted in more pronounced inhibition of tumor growth than concurrent treatment. These findings suggest that the targeting of CLDN4 enhances the antitumoral effects of chemotherapeutic agents and molecular targeting antibodies when used in combination.

Orthotopic Pancreatic Tumor Mouse Models of Liver Metastasis.

The survival from pancreatic cancer is poor because most patients are diagnosed after the cancer has metastasized. Liver is the most common site of pancreatic cancer metastasis. Orthotopic mouse models of liver metastasis by intrasplenically injecting the pancreatic tumor cells are useful in studying the molecular mechanisms of metastasis and evaluating therapeutic regimens.

HNF6 promotes tumor growth in colorectal cancer and enhances liver metastasis in mouse model.

Hepatocyte nuclear factor 6 (HNF6), as a transcription factor, has been reported to be involved in cell proliferation, carcinogenesis, and tumor metastasis. Here, we demonstrated the role of HNF6 in tumor growth and liver metastasis in colorectal cancer (CRC). Through bioinformatics and clinical samples analysis, we found HNF6 messenger RNA was upregulated both in CRC primary sites and liver metastases, and its high expression indicated poor survival in CRC patients. In vitro studies confirmed that HNF6 promoted cell proliferation and colony formation. What is more, in mouse models, the xenografts grew significantly faster and liver metastasis rate was nearly 45% higher in mice injected with HNF6-overexpressing cells. Further mechanism exploration showed that HNF6 expression affected cell adhesion and conferred resistance to anoikis in CRC cells. Taken together, HNF6 expression was upregulated in CRC and closely correlated with poor survival. HNF6 promoted CRC cell proliferation and tumor growth, and may contribute to liver metastasis via conferring cell resistance to anoikis.

Abstract 1728: Nonalcoholic steatohepatitis (NASH) impairs treatment of intrahepatic metastases with CD4+ T cell dependent RNA vaccine

Abstract Nonalcoholic fatty liver disease (NAFLD) is the most common liver disorder in Western industrialized countries. NAFLD along with its advanced form non-alcoholic steatohepatitis (NASH) are becoming an increasing risk factor for the development of cirrhosis and subsequently for hepatocellular carcinoma (HCC). We showed a selective CD4+ T cell loss in mice fed with methionine-choline deficient (MCD) diet, a model to induce NASH (Ma et al. Nature 2016). This selective loss of CD4 T cells may not only lead to immune escape by tumor cells but also lessen the effectiveness of immunotherapy. Kreiter et al. created a RNA vaccine from B16 melanoma cells encoding the neo-epitope M30, which is presented by MHC class II and recognized by CD4+ T cells. The vaccine showed therapeutic immune response in mouse models of subcutaneous B16-tumors and lung metastases (Kreiter et al. Nature 2015). Here we present an in vivo study of M30 RNA vaccine in a mouse model of B16 melanoma liver metastasis induced by intrahepatic injection. Mice were separated into groups and fed regular chow or MCD diet. B16F10-GFP-Luc melanoma cells were intrahepatically injected and mice were treated with M30 RNA or irrelevant RNA as control via tail vein injection. Intrahepatic tumor growth was monitored by bioluminescence imaging over the course of treatment. Mice were sacrificed and immunomonitoring of tumor surrounding liver tissue and tumor infiltrating lymphocytes (TIL) was performed. Mice treated with M30 RNA vaccine showed reduced tumor growth of intrahepatic metastasis compared to irrelevant RNA. Mice fed MCD diet could not control tumor growth either receiving M30 RNA vaccine treatment or irrelevant RNA. This impaired function of the treatment was related to a loss of CD4+ T cells in the NASH liver and a more immunosuppressive immune cell phenotype in the tumor environment. This study indicates impaired efficacy of cancer immunotherapy due to fatty liver disease and subsequently changed immune response due to loss of CD4+ T cells as the main effector immune cell. As the M30 RNA vaccine reduced tumor growth in non-fatty liver, this data might be considered when planning future immunotherapy treatment strategies for comorbid patients with fatty liver disease as CD4+ T cell based treatment strategies might be impaired in patients with NASH. Citation Format: Bernd Heinrich, Zachary J. Brown, Vormehr Mathias, Qiong Fu, Chi Ma, Su Jong Yu, Ugur Sahin, Tim F. Greten. Nonalcoholic steatohepatitis (NASH) impairs treatment of intrahepatic metastases with CD4+ T cell dependent RNA vaccine [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 1728.

Protective Role of the Hepatic Vagus Nerve against Liver Metastasis in Mice

Objective(s): Although accumulating evidence has shown that the autonomic nervous system is involved in liver pathology, its role in regulating cancer development remains unclear. The purpose of this study was to elucidate its detailed mechanisms. Methods: A mouse model of liver metastasis of colorectal cancer was used. To elucidate the potential mechanisms involved, we examined the effect of selective hepatic vagotomy on the survival rate and liver-to-body weight. We further evaluated the possible involvement of the hepatic sympathetic nerve fibers in this model. Results: The mortality rate and the liver-to-body weight ratio after cancer inoculation were significantly higher in the vagotomized mice than in the sham-operated mice. The vagotomized mice exhibited a transient decrease in hepatic norepinephrine levels following cancer inoculation. Interestingly, the vagotomy-induced exacerbation of liver metastasis was attenuated by supplementary norepinephrine or phenylephrine, a selective α₁-adrenoceptor agonist, but not by clonidine, a selective α₂-adrenoceptor agonist. Conclusion: Collectively, these results suggest that the hepatic vagus nerve may play a protective role against liver metastasis. Hepatic sympathetic nerves may also be involved as a protective efferent loop, possibly acting through the α₁-adrenoceptor.

Chemotherapy can promote liver metastasis by enhancing metastatic niche formation in mice

BackgroundSome chemotherapeutic agents have been reported to promote lung metastasis. However, there have been no reports regarding chemotherapy-induced liver metastasis. We hypothesized that chemotherapy might also enhance liver metastasis. The present study aimed to create a chemotherapy-enhanced liver metastasis mouse model and investigate its mechanism. Materials and methodsMice were pretreated with cisplatin, vincristine, or saline by intraperitoneal injection. Next, B16F10 mouse melanoma cells and BE(2)-C human neuroblastoma cells were injected into the spleens of C57BL/6 and BALB/c nu/nu mice, respectively, to induce experimental liver metastasis, and the number of liver nodules was determined. We also analyzed the effect of chemotherapy on changes of the liver tissue regarding representative metastasis-promoting factors using real-time quantitative polymerase chain reaction and immunohistochemical and histological analysis. ResultsCisplatin increased the number of nodules by 4.7-fold in the B16F10 liver metastasis model. Vincristine increased the number of nodules by 3.8-fold in the BE(2)-C liver metastasis model. Cisplatin increased mRNA levels of matrix-metalloproteinase (MMP)-2 and periostin, while vincristine increased MMP-9 and S100A8/9 levels in liver tissues. Cisplatin induced fibrosis, whereas vincristine induced neutrophil recruitment in liver tissues according to histological and immunohistochemical analysis. ConclusionsWe concluded that cisplatin or vincristine could enhance liver metastasis of mouse melanoma cells or human neuroblastoma cells, respectively. In addition, the mRNA expression of MMP-2 and periostin, or MMP-9 and S100A8/9 is increased by cisplatin or vincristine pretreatment, possibly resulting in fibrosis or neutrophil recruitment, respectively. These niche factors might be associated with increased liver metastasis.

Impact of associating liver partition and portal vein occlusion for staged hepatectomy on tumor growth in a mouse model of liver metastasis

BackgroundThe impact of associating liver partition and portal vein occlusion for staged hepatectomy (ALPPS) on tumor growth activity was investigated. MethodsA BALB/c mouse model (male, 8–10 weeks old) of liver metastasis labeled by red fluorescent protein was established. Changes in future liver remnant (FLR) volumes, tumor growth activity, and levels of cytokines and growth factors in liver tissues during the treatment period were compared among the models involving ALPPS, portal vein ligation (PVL), or sham operation. ResultsThe ratio of the FLR volume to body weight at 24 h after the procedure was greater for ALPPS (4.45 ± 0.12 × 10−2) than for PVL (3.79 ± 0.12 × 10−2; P = 0.003) and sham operation (3.18 ± 0.16 × 10−2; P < 0.001). No differences in tumor progression in the FLR were observed at any time point after the procedures. Within the deportalized liver (DL), although tumor progression was observed during a later period after ALPPS (9 days postoperative) and PVL (12 days postoperative), no acceleration of tumor growth after ALPPS was observed in an early period similar to PVL. ConclusionALPPS induces a rapid increase in FLR volume and avoids remnant tumor progression during the early postoperative period.

MiR-146a targets c-met and abolishes colorectal cancer liver metastasis

A major complication of colorectal cancer (CRC), one of the most frequent and deadly types of cancer, is disease progression via liver metastases. At this stage, very few treatment options are available for patients, and the disease remains incurable. Herein, we used a well-established mouse model of CRC liver metastasis (CLM) to identify new regulators of this process. Using serial transplantation of murine MC38 adenocarcinoma cells, we obtained liver metastatic variants that displayed extremely strong colonization abilities. Using these newly established cell lines, we performed gene expression arrays and microRNA (miR) profiling. Comparative and predictive analyses between the two arrays showed higher expression of c-met and concomitant reduction of miR-146a in the mestastatic variants. In CRC patients, expression levels of both c-met and miR-146a were similar between primary tumors and liver metastases. Interestingly, we identified c-met as a new target for miR-146a, as miR-146a was able to impede c-met translation. Of relevance, overexpression of miR-146a in metastatic clones showed reduced in vitro malignancy and abolished the development of primary tumor and liver metastases. Our results document a new mechanism for c-met regulation in CLM and highlight the crucial role of miR-146a in suppressing tumorigenesis.

Influence of the Twist gene on the invasion and metastasis of colon cancer.

The present study investigated the role of the Twist gene in epithelial-mesenchymal transition (EMT) and its effects on the invasion and metastasis of malignant tumors. In vitro, we transfected SW480, HCT116 and HT29 cells with recombinant plasmids, pTracer-CMV/BSD-Twist and pGenesil1.2-Twist-shRNA, to influence expression of Twist. The transfection efficacy of the plasmids in the cell lines was confirmed by flow cytometry. The relative mRNA and protein expression levels of Twist, E-cadherin and vimentin in the transfected cells were detected by RT-PCR and western blotting, respectively. In addition, migration and invasion were assessed by Transwell assays. In vivo, we established a xenogenic liver metastasis mouse model by intrasplenic injection with transfected SW480, HCT116 or HT29 human colon cancer cells and used hematoxylin and eosin (H&E) staining to demonstrate the effective establishment of the model. The relative mRNA levels of Twist and vimentin were detected by RT-PCR. In vitro, RT-PCR and western blotting showed higher relative mRNA and protein expression levels of Twist and vimentin in cell lines transfected with the recombinant, highly expressed Twist plasmid than in non-transfected cell lines (P<0.05), while E-cadherin was inhibited (P<0.05). After transfection with the plasmid pGenesil1.2-Twist-shRNA, the relative mRNA and protein levels of Twist and vimentin were markedly inhibited in the HCT116 cells (P<0.05), and the levels of E-cadherin were not changed (P>0.05), along with inhibition of the migration and invasion abilities of the cell line (P<0.01). In vivo, relative mRNA levels of Twist and vimentin in both the liver and spleen of the mouse model were higher in the groups that were injected with one of the three cell lines transfected with pTracer-CMV/BSD-Twist than in the groups injected with cells transfected with pGenesil1.2-Twist-shRNA (P<0.05). In conclusion, upregulation of Twist gene expression can promote EMT molecular events. Interfering with the Twist gene can effectively silence Twist gene expression in HCT116 cells and consequently inhibit colon cancer cell migration and invasion.

Polymeric micelles loaded with (1,2-diaminocyclohexane)platinum(II) against colorectal cancer

BackgroundWe investigated the potential of nanomedicine in loading the oxaliplatin parent complex (1,2-diaminocyclohexane)platinum(II)-loaded polymeric micelles (DACHPt/m) against multiple liver metastases from colon cancer in a mouse model. Materials and methodsThe efficacy of DACHPt/m or oxaliplatin (on days 14 and 21 after inoculation of tumor cells) was evaluated in a mouse model of liver metastasis for murine colon adenocarcinoma C26 cells. In vivo antitumor effects were evaluated by recording the number of liver metastases and weights of metastatic livers after treatment (day 28). The accumulation of drugs in tumors and liver parenchyma was analyzed using ion coupled plasma-mass spectrometry 24 h after administration of DACHPt/m or oxaliplatin (n = 5). We assessed renal and hepatic toxicities through changes in creatinine, aspartate transaminase, and alanine transaminase on the last day of the antitumor activity experiment. ResultsMice receiving DACHPt/m had significantly fewer metastatic nodules (P = 0.038) and lower liver weights (P = 0.038) than those receiving oxaliplatin. The accumulation of DACHPt/m in the metastatic liver was significantly higher than that of oxaliplatin, whereas the distribution of micelles in healthy liver tissues was limited. Mice treated with DACHPt/m also showed significantly lower serum creatinine levels than those treated with oxaliplatin (P = 0.007), whereas serum aspartate transaminase and alanine transaminase levels for both drugs were not different. ConclusionsHigh levels of DACHPt/m accumulate in metastatic livers, producing a strong antitumor effect without severe adverse effects. DACHPt/m is a safe approach for managing liver metastasis from colorectal cancer.

Inhibition of PRDM14 expression in pancreatic cancer suppresses cancer stem-like properties and liver metastasis in mice.

Pancreatic cancer is one of the most lethal types of cancer, with aggressive properties characterized by metastasis, recurrence and drug resistance. Cancer stem cells are considered to be responsible for these properties. PRDM14, a transcriptional regulator that maintains pluripotency in embryonic stem cells, is overexpressed in some cancers. Here, we assessed PRDM14 expression and the effects of PRDM14 knockdown on cancer stem-like phenotypes in pancreatic cancer. We observed that PRDM14 protein was overexpressed in pancreatic cancer tissues compared with normal pancreatic tissues. Using lentiviral shRNA-transduced pancreatic cancer cells, we found that PRDM14 knockdown decreased sphere formation, number of side population and cell surface marker-positive cells and subcutaneous xenograft tumors and liver metastasis in mice. This was accompanied by upregulation of some microRNAs (miRNAs), including miR-125a-3p. miR-125a-3p, a tumor suppressor that is down-regulated in pancreatic cancer, has been suggested to regulate the expression of the Src-family kinase, Fyn. In PRDM14-knockdown cells, Fyn was expressed at lower levels and downstream proteins were less activated. These changes were considered to cause suppression of the above cancer phenotypes. In addition, we used small interfering RNA (siRNA)-based therapy targeting PRDM14 in a mouse model of liver metastasis induced using MIA-PaCa2 cells, and this treatment significantly decreased metastasis and in vitro migration. Taken together, these results suggest that targeting the overexpression of PRDM14 suppresses cancer stem-like phenotypes, including liver metastasis, via miRNA regulation and siRNA-based therapy targeting it shows promise as a treatment for patients with pancreatic cancer.

Selumetinib Inhibits Melanoma Metastasis to Mouse Liver via Suppression of EMT-targeted Genes.

We investigated the therapeutic effects of a mitogen-activated protein (MEK) inhibitor, selumetinib, in a hepatic melanoma metastasis model and studied its possible mechanism of action. Melanoma cell lines were exposed to selumetinib under different experimental conditions. We established a mouse model of liver metastasis and treated mice orally with vehicle or selumetinib and then evaluated metastasis progress. Growth inhibition was observed in melanoma cells as a consequence of G1-phase cell-cycle arrest and the subsequent induction of apoptosis in a dose- and time-dependent manner. Mice with established liver metastases that were treated with selumetinib exhibited significantly less tumor progression than vehicle-treated mice. c-Myc expression in metastasized liver tissues were suppressed by selumetinib. Moreover, oral treatment with selumetinib modulated expression of epithelial-to-mesenchymal transition- and metastasis-related genes, including integrin alpha-5 (ITGA5), jagged 1 (JAG1), zinc finger E-box-binding homeobox 1 (ZEB1), NOTCH, and serpin peptidase inhibitor clade E (SERPINE1). We established a mouse model of hepatic metastasis using a human melanoma cell line, such models are essential in elucidating the therapeutic effects of anti-metastatic drugs. Our data suggest the possibility that selumetinib presents a new strategy to treat liver metastasis in patients with melanoma by suppressing epithelial-to-mesenchymal transition-related genes.

Antihypoxic Potentiation of Standard Therapy for Experimental Colorectal Liver Metastasis through Myo-Inositol Trispyrophosphate.

Tumor hypoxia activates hypoxia-inducible factors (Hifs), which induce a range of malignant changes including vascular abnormalities. Here, we determine whether inhibition of the hypoxic tumor response through myo-inositol trispyrophosphate (ITPP), a compound with antihypoxic properties, is able to cause prolonged vascular normalization that can be exploited to improve standard-of-care treatment. We tested ITPP on two syngeneic orthotopic mouse models of lethal colorectal cancer liver metastasis. Tumors were monitored by MRI and analyzed for the hypoxic response and their malignant potential. A Hif activator and in vitro assays were used to define the working mode of ITPP. Hypoxic response and vasculature were re-evaluated 4 weeks after treatment. Finally, we determined survival following ITPP monotherapy, FOLFOX monotherapy, FOLFOX plus Vegf antibody, and FOLFOX plus ITPP, both overlapping and sequential. ITPP reduced tumor load, efficiently inhibited the hypoxic response, and improved survival. These effects were lost when mice were pretreated with a Hif activator. Its immediate effects on the hypoxic response, including an apparent normalization of tumor vasculature, persisted for at least 4 weeks after treatment cessation. Compared with FOLFOX alone, Vegf antibody combined with FOLFOX prolonged survival by <30%, whereas ITPP combined with FOLFOX extended survival by >140%, regardless of whether FOLFOX was given in overlap or after ITPP exposure. Our findings reveal a truly antihypoxic mechanism for ITPP and demonstrate the capacity of this nontoxic compound to potentiate the efficacy of existing anticancer treatment in a way amenable to clinical translation. Clin Cancer Res; 22(23); 5887-97. ©2016 AACR.

Expression of metastasis suppressor gene AES driven by a Yin Yang (YY) element in a CpG island promoter and transcription factor YY2.

We recently found that the product of the AES gene functions as a metastasis suppressor of colorectal cancer (CRC) in both humans and mice. Expression of amino‐terminal enhancer of split (AES) protein is significantly decreased in liver metastatic lesions compared with primary colon tumors. To investigate its downregulation mechanism in metastases, we searched for transcriptional regulators of AES in human CRC and found that its expression is reduced mainly by transcriptional dysregulation and, in some cases, by additional haploidization of its coding gene. The AES promoter‐enhancer is in a typical CpG island, and contains a Yin‐Yang transcription factor recognition sequence (YY element). In human epithelial cells of normal colon and primary tumors, transcription factor YY2, a member of the YY family, binds directly to the YY element, and stimulates expression of AES. In a transplantation mouse model of liver metastases, however, expression of Yy2 (and therefore of Aes) is downregulated. In human CRC metastases to the liver, the levels of AES protein are correlated with those of YY2. In addition, we noticed copy‐number reduction for the AES coding gene in chromosome 19p13.3 in 12% (5/42) of human CRC cell lines. We excluded other mechanisms such as point or indel mutations in the coding or regulatory regions of the AES gene, CpG methylation in the AES promoter enhancer, expression of microRNAs, and chromatin histone modifications. These results indicate that Aes may belong to a novel family of metastasis suppressors with a CpG‐island promoter enhancer, and it is regulated transcriptionally.