[Show abstract][Hide abstract] ABSTRACT: Recent work has identified dysfunctional Hippo signaling to be involved in maintenance and progression of various human cancers, although data on clear cell renal cell carcinoma (ccRCC) have been limited. Here, we provide evidence implicating aberrant Hippo signaling in ccRCC proliferation, invasiveness, and metastatic potential. Nuclear overexpression of the Hippo target Yes-associated protein (YAP) was found in a subset of patients with ccRCC. Immunostaining was particularly prominent at the tumor margins and highlighted neoplastic cells invading the tumor-adjacent stroma. Short hairpin RNA-mediated knockdown of YAP significantly inhibited proliferation, migration, and anchorage-independent growth of ccRCC cells in soft agar and led to significantly reduced murine xenograft growth. Microarray analysis of YAP knockdown versus mock-transduced ccRCC cells revealed down-regulation of endothelin 1, endothelin 2, cysteine-rich, angiogenic inducer, 61 (CYR61), and c-Myc in ccRCC cells as well as up-regulation of the cell adhesion molecule cadherin 6. Signaling pathway impact analysis revealed activation of the p53 signaling and cell cycle pathways as well as inhibition of mitogen-activated protein kinase signaling on YAP down-regulation. Our data suggest CYR61 and c-Myc as well as signaling through the endothelin axis as bona fide downstream effectors of YAP and establish aberrant Hippo signaling as a potential therapeutic target in ccRCC.
[Show abstract][Hide abstract] ABSTRACT: The evolutionarily conserved Hedgehog (Hh) signaling pathway is essential for correct embryogenesis and is misregulated in several malignancies. In cell culture, Hh-sensitive cells display a striking dependence on cell density with active Hh signaling requiring cell-to-cell contact. As the Hippo/YAP system is tightly linked to cell density control and contact inhibition, we investigated the cross-talk between the two pathways. Our data reveal that the suppression of Hh signaling in the absence of cellular contacts is independent of primary cilia and is mediated by the YAP oncogene. Overexpression of YAP blocks Hh signaling whereas RNA interference-mediated knockdown of YAP enhances Hh/GLI activity. Despite this negative regulation, Hh signaling promotes YAP activity through post-transcriptional mechanisms, resulting in a negative feedback loop. In vivo, we found strong nuclear YAP immunoreactivity restricted to compartments with low Hh pathway activity in human and mouse pancreatic cancer. Finally, we identified protease-activated receptors (PARs) as molecules being able to override the inverse Hippo/Hh regulation, potentially giving tumors a mechanism to utilize both oncogenic pathways in parallel.Oncogenesis (2014) 3, e112; doi:10.1038/oncsis.2014.27; published online 11 August 2014.
[Show abstract][Hide abstract] ABSTRACT: Due to an extensive use of modern imaging, incidental pancreatic cysts are increasingly diagnosed these days. Fortunately, comprehensive research over the past years has remarkably improved our pathogenetic and clinical understanding of pancreatic cysts that, as we know, are in majority harmless. However, mucinous cysts including intraductal papillary mucinous neoplasms, mucinous cystic neoplasms, as well as solid pseudopapillary neoplasms harbor relevant potential for developing into a lethal invasive cancer and may therefore require immediate surgical resection or at least close surveillance. In order to allow an optimized clinical management, it is crucial to gather reliable information about entity as well as biologic behavior of every cyst detected. Unfortunately, in the absence of reliable biomarkers and by just applying currently available diagnostic means such as clinical and radiologic criteria or cyst fluid cytology, there is still a risk for incorrect preoperative diagnoses. This may be followed by inappropriate treatment possibly resulting in severe morbidity or even mortality.
In this review article, we summarize some of the salient recent advances in molecular diagnostics of pancreatic cysts. Herein, we put particular focus on the emerging field of biomarker research in pancreatic cyst fluid based on protein, DNA, and microRNA analyses.
Langenbeck s Archives of Surgery 09/2013; · 1.89 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Introduction: Despite being the second most common malignancy of the pancreas, pancreatic neuroendocrine tumors (PNET) have long been understudied due to their low incidence and heterogeneous clinical presentation. Emerging data from a Phase III trial demonstrates improved progression-free survival of patients with advanced PNET on treatment with sunitinib . Areas covered: This article reviews the role of sunitinib, a multitargeted tyrosine kinase inhibitor with potent antiangiogenic and antitumor effects, in the clinical management of PNET. Furthermore, the authors also discuss the pharmacokinetics and pharmacodynamics as well as other clinically relevant aspects regarding sunitinib. Expert opinion: A recent Phase III clinical trial of sunitinib demonstrated significant improvement of progression-free survival in patients with advanced or metastatic well-differentiated PNET that led to its approval in several countries, including Europe and United States. This marks a significant step forward in the clinical management of this disease and spurs hopes to further improve overall survival in this once difficult-to-treat set of patients in the coming years. Fields of future interest will include evaluation of combinatorial regimens, including conventional cytotoxic agents as well as additional targeted drugs in order to overcome resistance to sunitinib.
Expert Opinion on Drug Metabolism & Toxicology 04/2013; · 2.94 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: PURPOSE: High-throughput chemosensitivity testing of low-passage cancer cell lines can be used to prioritize agents for personalized chemotherapy. However, generating cell lines from primary cancers is difficult, because contaminating stromal cells overgrow the malignant cells. EXPERIMENTAL DESIGN: We produced a series of hypoxanthine phosphoribosyl transferase (hprt)-null immunodeficient mice. During growth of human cancers in these mice, hprt-null murine stromal cells replace their human counterparts. RESULTS: Pancreatic and ovarian cancers explanted from these mice were grown in selection media to produce pure human cancer cell lines. We screened one cell line with a 3,131-drug panel and identified seventy-seven FDA approved drugs with activity, including two novel drugs to which the cell line was uniquely sensitive. Xenografts of this carcinoma were selectively responsive to both drugs. CONCLUSIONS: Chemotherapy can be personalized using patient-specific cell lines derived in biochemically selectable mice.
Clinical Cancer Research 01/2013; · 7.84 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Management of patients suffering from metastatic malignant melanoma and brain metastasis remains challenging in routine clinical practice. The inhibitory anti-CTLA-4 antibody ipilimumab has recently been approved as second-line therapeutic option for melanoma patients. Increasing evidence suggests distinct therapeutic activity on central nervous system metastases, although this continues to be actively debated. Here, we present the case of a patient suffering from metastatic melanoma, including symptomatic brain metastasis, who showed a partial response to ipilimumab in extracranial tumor manifestations, while the disease was progressing intracranially. Subsequently, intracranial disease progression could be managed by local irradiation. An overview of currently available literature on the efficacy of ipilimumab in melanoma patients with central nervous system metastases is provided.
[Show abstract][Hide abstract] ABSTRACT: Introduction: Pancreatic neuroendocrine tumors (PNET) represent the second most common primary malignancy of the pancreas. Until recently, therapeutic options for advanced PNET have been limited. Areas covered: A recently published Phase III clinical trial demonstrated striking therapeutic activity of the mTOR inhibitor everolimus in advanced PNET and led to its approval for this indication by the FDA. This review discusses this landmark discovery in the context of currently available therapeutic options, pathophysiology and molecular genetics of PNET. Expert opinion: The approval of everolimus for the treatment of PNET marks a major step forward in the clinical management of this disease and represents a notable example of the successful translation of a targeted therapy that was initially developed based on findings at the lab bench, into everyday clinical practice. These results encourage hopes that the overall therapeutic efficacy of such approaches can be further enhanced by the introduction of combinatorial regimens, simultaneously targeting more than one oncogenic signaling pathway, as well as by stratification of patients based on the individual genetic setup of their tumors.
Expert Opinion on Pharmacotherapy 08/2012; 13(14):2073-84. · 2.86 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: It is a challenging task to distinguish between benign and malignant lesions in patients with biliary strictures. Here we analyze whether determination of target gene mRNA levels in intraductal brush cytology specimens may be used to improve the diagnosis of bile duct carcinoma.
Brush cytology specimens from 119 patients with biliary strictures (malignant: n = 72; benign: n = 47) were analyzed in a retrospective cohort study. mRNA of IGF-II mRNA-binding protein 3 (IGF2BP3), homeobox B7 (HOXB7), Forkhead box M1 (FOXM1), kinesin family member 2C (KIF2C) and serine/threonine kinase NEK2 was determined by semi-quantitative RT-PCR using the ΔCt method.
IGF2BP3 (p<0.0001), HOXB7 (p<0.0001), and NEK2 (p<0.0001) mRNA expression levels were significantly increased in patients with cholangiocarcinoma or pancreatic cancer. Median ΔCt values differed by 3.5 cycles (IGF2BP3), 2.8 cycles (HOXB7) and 1.3 cycles (NEK2) corresponding to 11-fold, 7-fold and 2.5-fold increased mRNA levels in malignant versus benign samples. Sensitivity to detect biliary cancer was 76.4% for IGF2BP3 (80.9% specificity); 72.2% for HOXB7 (78.7% specificity) and 65.3% for NEK2 (72.3% specificity), whereas routine cytology reached only 43.1% sensitivity (85.4% specificity). Diagnostic precision was further improved, when all three molecular markers were assessed in combination (77.8% sensitivity, 87.2% specificity) and achieved 87.5% sensitivity and 87.2% specificity when molecular markers were combined with routine cytology.
Our data suggest that measuring IGF2BP3, HOXB7 and NEK2 mRNA levels by RT-PCR in addition to cytology has the potential to improve detection of malignant biliary disorders from brush cytology specimens.
PLoS ONE 01/2012; 7(8):e42141. · 3.73 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Pancreatic cancer is one of the most lethal of human malignancies, and potent therapeutic options are lacking. Inhibition of cell cycle progression through pharmacological blockade of cyclin-dependent kinases (CDK) has been suggested as a potential treatment option for human cancers with deregulated cell cycle control. Dinaciclib (SCH727965) is a novel small molecule multi-CDK inhibitor with low nanomolar potency against CDK1, CDK2, CDK5 and CDK9 that has shown favorable toxicity and efficacy in preliminary mouse experiments, and has been well tolerated in Phase I clinical trials. In the current study, the therapeutic efficacy of SCH727965 on human pancreatic cancer cells was tested using in vitro and in vivo model systems. Treatment with SCH727965 significantly reduced in vitro cell growth, motility and colony formation in soft agar of MIAPaCa-2 and Pa20C cells. These phenotypic changes were accompanied by marked reduction of phosphorylation of Retinoblastoma (Rb) and reduced activation of RalA. Single agent therapy with SCH727965 (40 mg/kg i.p. twice weekly) for 4 weeks significantly reduced subcutaneous tumor growth in 10/10 (100%) of tested low-passage human pancreatic cancer xenografts. Treatment of low passage pancreatic cancer xenografts with a combination of SCH727965 and gemcitabine was significantly more effective than either agent alone. Gene Set Enrichment Analysis identified overrepresentation of the Notch and Transforming Growth Factor-β (TGF-β) signaling pathways in the xenografts least responsive to SCH727965 treatment. Treatment with the cyclin-dependent kinase inhibitor SCH727965 alone or in combination is a highly promising novel experimental therapeutic strategy against pancreatic cancer.
Cancer biology & therapy 10/2011; 12(7):598-609. · 3.29 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Ligand-dependent activation of the Hedgehog (Hh) signaling pathway has been implicated in both tumor initiation and metastasis of pancreatic ductal adenocarcinoma (PDAC). Prior studies in genetically engineered mouse models (GEMMs) have assessed the role of Hh signaling by cell autonomous expression of a constitutively active Gli2 within epithelial cells. On the contrary, aberrant pathway reactivation in the human exocrine pancreas occurs principally as a consequence of Sonic Hh ligand (Shh) overexpression from epithelial cells. To recapitulate the cognate pathophysiology of Hh signaling observed in the human pancreas, we examined GEMM where Hh ligand is conditionally overexpressed within the mature exocrine pancreas using a tamoxifen-inducible Elastase-Cre promoter (Ela-CreERT2;LSL-mShh). We also facilitated potential cell autonomous epithelial responsiveness to secreted Hh ligand by generating compound transgenic mice with concomitant expression of the Hh receptor Smoothened (Ela-CreERT2;LSL-mShh;LSL-mSmo). Of interest, none of these mice developed intraductal precursor lesions or PDAC during the follow-up period of up to 12 months after tamoxifen induction. Instead, all animals demonstrated marked expansion of stromal cells, consistent with the previously described epithelial-to-stromal paracrine Hh signaling. Hh responsiveness was mirrored by the expression of primary cilia within the expanded mesenchymal compartment and the absence within mature acinar cells. In the absence of cooperating mutations, Hh ligand overexpression in the mature exocrine pancreas is insufficient to induce neoplasia, even when epithelial cells coexpress the Smo receptor. This autochthonous model serves as a platform for studying epithelial stromal interactions in pancreatic carcinogenesis.
Neoplasia (New York, N.Y.) 10/2011; 13(10):923-30. · 5.48 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: An inactivating germline mutation in BRCA2 is the most common known genetic basis for familial pancreatic cancer (FPC), accounting for 5-10% of inherited cases. A genetically engineered mouse model of pancreatic ductal adenocarcinoma (PDAC) arising on the backdrop of Brca2 deficiency is likely to elucidate valuable diagnostic and therapeutic insights for FPC. Both Brca2 alleles were conditionally deleted during development within the pancreatic epithelium by generating Pdx1-Cre; Brca2(f/f) (CB) mice; in addition, triple transgenic Pdx1-Cre; Brca2(f/f); LSL-Trp53(R172H) (CBP) mice were generated, in order to determine the impact of p53 deregulation on Brca2-deficient carcinogenesis. Both CB and CBP mice developed non-invasive ductal precursor lesions (murine pancreatic intraepithelial neoplasia or mPanIN), although these were observed at an earlier time point (5 versus 8 months) and with higher prevalence in CBP mice. A minority of CB mice (15%) developed invasive and metastatic PDAC at a latency of 15 months or greater; in contrast, CBP mice of comparable age uniformly developed PDAC with variable histological features. Mortality in the absence of neoplasia in CB and CBP mice was associated with profound loss of pancreatic parenchyma, consistent with progressive elimination of Brca2-deficient cells. Widespread DNA damage, as evidenced by overexpression of the phosphorylated histone H(2)AX(Ser139), was observed in the non-neoplastic exocrine pancreas, as well as in the mPanIN and PDAC lesions of Brca2-deficient mice, independent of p53 status. Loss of Brca2 function predisposes the exocrine pancreas to profound DNA damage, and the frequency of invasive neoplasia is accentuated by the concomitant deregulation of p53.
Cancer biology & therapy 06/2011; 11(11):959-68. · 3.29 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Pancreatic cancer is a devastating disease associated with near uniform mortality. Usually diagnosed at advanced, metastatic stages when surgical resection with curative intention is not possible any more, most patients succumb to progressive disease after a few months. Despite recent advances in understanding pancreatic carcinogenesis and continuous efforts in translational research, so far these results failed to translate into clinically relevant improvements of patient survival.
Preclinical evaluation of drug candidates and novel therapeutic strategies rely on in vitro and in vivo model systems to predict response in patients. This article reviews mouse models of pancreatic cancer, their respective applications in translational research and discusses their potential to predict clinical responses in patients.
This article provides a profound overview of individual strength as well as of shortcomings of mouse models of pancreatic cancer currently available for translational research.
Considerable progress in designing mouse models of pancreatic cancer has been made over the last decade and several xenograft as well as genetically engineered mouse models faithfully recapitulating human disease development has been developed. Taken together, these newly developed in vivo model systems provide powerful tools likely to boost preclinical evaluation and bench-to-bedside transition of novel therapeutic approaches directed against this dire malady.
Expert Opinion on Drug Discovery 01/2011; 6(1):33-48. · 2.30 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Although activating mutations in RAS oncogenes are known to result in aberrant signaling through multiple pathways, the role of microRNAs (miRNAs) in the Ras oncogenic program remains poorly characterized. Here we demonstrate that Ras activation leads to repression of the miR-143/145 cluster in cells of human, murine, and zebrafish origin. Loss of miR-143/145 expression is observed frequently in KRAS mutant pancreatic cancers, and restoration of these miRNAs abrogates tumorigenesis. miR-143/145 down-regulation requires the Ras-responsive element-binding protein (RREB1), which represses the miR-143/145 promoter. Additionally, KRAS and RREB1 are targets of miR-143/miR-145, revealing a feed-forward mechanism that potentiates Ras signaling.
Genes & development 12/2010; 24(24):2754-9. · 12.08 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Recent evidence has demonstrated that aberrant reactivation of the Hedgehog signaling pathway contributes to tumor initiation and progression in various human malignancies, including pancreatic cancer; therefore, the Hedgehog pathway has emerged as a promising novel therapeutic target. Initial translational studies conducted using cyclopamine, a small-molecule inhibitor of the Smoothened (SMO) component of the Hedgehog pathway, demonstrated that pharmacological blockade of aberrant Hedgehog signaling has the potential to inhibit tumor initiation, progression and metastatic spread. This concept has been corroborated using different compounds in various preclinical models of pancreatic cancer and other malignancies; several of these studies suggest possible therapeutic synergisms of Hedgehog inhibitors with established antineoplastic agents. This review provides a concise overview of translational studies assessing the use of Hedgehog inhibitors as novel therapeutic strategy for cancer, particularly pancreatic cancer.
Current opinion in investigational drugs (London, England: 2000) 12/2010; 11(12):1387-98. · 3.55 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Pancreatic cancer has both genetic and epigenetic underlying causes. The importance of epigenetic alterations in the formation
and maintenance of malignant tumors has become apparent in the last decade, with accumulating evidence suggesting this is
probably the most common clonal aberration in human neoplasia. Identifying epigenetic alterations in pancreatic cancer has
not only enhanced our understanding of pancreatic cancer biology, but has also opened up avenues for the development of early
detection and novel therapeutic strategies.
In this chapter, an overview of the current literature on epigenetic alterations found in pancreatic cancer is presented and
discussed in the light of potential therapeutic applicability as well as pointing out possible future directions of studies
combining global genetic and epigenetic analyses.
[Show abstract][Hide abstract] ABSTRACT: Esophageal adenocarcinoma (EAC) arises in the backdrop of reflux-induced metaplastic phenomenon known as Barrett esophagus. The prognosis of advanced EAC is dismal, and there is an urgent need for identifying molecular targets for therapy. Serial Analysis of Gene Expression (SAGE) was performed on metachronous mucosal biopsies from a patient who underwent progression to EAC during endoscopic surveillance. SAGE confirmed significant upregulation of Axl "tags" during the multistep progression of Barrett esophagus to EAC. In a cohort of 92 surgically resected EACs, Axl overexpression was associated with shortened median survival on both univariate (p < 0.004) and multivariate (p < 0.036) analysis. Genetic knockdown of Axl receptor tyrosine kinase (RTK) function was enabled in two EAC lines (OE33 and JH-EsoAd1) using lentiviral short hairpin RNA (shRNA). Genetic knockdown of Axl in EAC cell lines inhibited invasion, migration, and in vivo engraftment, which was accompanied by downregulation in the activity of the Ral GTPase proteins (RalA and RalB). Restoration of Ral activation rescued the transformed phenotype of EAC cell lines, suggesting a novel effector mechanism for Axl in cancer cells. Pharmacological inhibition of Axl was enabled using a small molecule antagonist, R428 (Rigel Pharmaceuticals). Pharmacological inhibition of Axl with R428 in EAC cell lines significantly reduced anchorage-independent growth, invasion and migration. Blockade of Axl function abrogated phosphorylation of ERBB2 (Her-2/neu) at the Tyr877 residue, indicative of receptor crosstalk. Axl RTK is an adverse prognostic factor in EAC. The availability of small molecule inhibitors of Axl function provides a tractable strategy for molecular therapy of established EAC.
Cancer biology & therapy 11/2010; 10(10):1009-18. · 3.29 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Curcumin or diferuloylmethane is a yellow polyphenol extracted from the rhizome of turmeric (Curcuma longa). A large volume (several hundreds) of published reports has established the anticancer and chemopreventative properties of curcumin in preclinical models of every known major cancer type. Nevertheless, the clinical translation of curcumin has been significantly hampered due to its poor systemic bioavailability, which mandates that patients consume up to 8 to 10 g of the free drug orally each day to achieve detectable levels in circulation. We have engineered a polymeric nanoparticle encapsulated curcumin formulation (NanoCurc) that shows remarkably higher systemic bioavailability in plasma and tissues compared with free curcumin upon parenteral administration. In xenograft models of human pancreatic cancer established in athymic mice, administration of parenteral NanoCurc significantly inhibits primary tumor growth in both subcutaneous and orthotopic settings. The combination of parenteral NanoCurc with gemcitabine results in enhanced tumor growth inhibition versus either single agent, suggesting an additive therapeutic influence in vivo. Furthermore, this combination completely abrogates systemic metastases in orthotopic pancreatic cancer xenograft models. Tumor growth inhibition is accompanied by significant reduction in activation of nuclear factor-kappaB, as well as significant reduction in expression of matrix metalloproteinase-9 and cyclin D1, in xenografts treated with NanoCurc and gemcitabine. NanoCurc is a promising new formulation that is able to overcome a major impediment for the clinical translation of curcumin to cancer patients by improving systemic bioavailability, and by extension, therapeutic efficacy.
Molecular Cancer Therapeutics 08/2010; 9(8):2255-64. · 5.60 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Cyclin-dependent kinase 5 (CDK5), a neuronal kinase that functions in migration, has been found to be activated in some human cancers in which it has been implicated in promoting metastasis. In this study, we investigated the role of CDK5 in pancreatic cancers in which metastatic disease is most common at diagnosis. CDK5 was widely active in pancreatic cancer cells. Functional ablation significantly inhibited invasion, migration, and anchorage-independent growth in vitro, and orthotopic tumor formation and systemic metastases in vivo. CDK5 blockade resulted in the profound inhibition of Ras signaling through its critical effectors RalA and RalB. Conversely, restoring Ral function rescued the effects of CDK5 inhibition in pancreatic cancer cells. Our findings identify CDK5 as a pharmacologically tractable target to degrade Ras signaling in pancreatic cancer.
Cancer Research 06/2010; 70(11):4460-9. · 8.65 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Intraductal papillary mucinous neoplasms (IPMNs) are one of the three known curable precursor lesions of invasive pancreatic ductal adenocarcinoma, an almost uniformly fatal disease. Cell lines from IPMNs and their invasive counterparts should be valuable to identify gene mutations critical to IPMN carcinogenesis, and permit high-throughput screening to identify drugs that cause regression of these lesions. To advance the study of the biological features of IPMNs, we attempted in vivo and in vitro growth of selected IPMNs based on the hypothesis that IPMNs could be grown in the most severely immunodeficient mice. We examined 14 cases by implanting them into nude, severe combined immunodeficient (SCID), and NOD/SCID/IL2Rgamma(null) (NOG) mice, in addition to direct culture, to generate tumor xenografts and cell lines. One sample was directly cultured only. Thirteen tumors were implanted into the three types of mice, including 10 tumors implanted into the triple immunodeficient NOG mice, in which the majority (8 of 10) grew. This included five IPMNs lacking an invasive component. One of the explanted IPMNs, with an associated invasive carcinoma, was successfully established as a cell line. Tumorigenicity was confirmed by growth in soft agar, growth in immunodeficient mice, and the homozygous deletion of p16/cdkn2a. Epithelial differentiation of the cell line was documented by cytokeratin expression. Patient origin was confirmed using DNA fingerprinting. Most non-invasive IPMNs grow in NOG mice. We successfully established one IPMN cell line, and plan to use it to clarify the molecular pathogenesis of IPMNs.