miR-431 Promotes Metastasis of Pancreatic Neuroendocrine Tumors by Targeting DAB2 Interacting Protein, a Ras GTPase Activating Protein Tumor Suppressor.

The incidence of pancreatic neuroendocrine tumor (PNET) is increasing, and it presents with various clinical manifestations and an unfavorable survival rate. A better understanding of the drivers of PNET tumorigenesis is urgently needed. Distinct miRNA signatures have been identified for different stages of tumorigenesis in both human and mouse PNETs. The functions of these miRNAs are poorly understood. miR-431 is the most up-regulated miRNA in the metastatic signature. However, it is unknown whether miR-431 contributes to metastasis of PNETs. Herein, we show that miR-431 overexpression activates Ras/extracellular signal-regulated kinase (Erk) signaling and promotes epithelial-mesenchymal transition, migration/invasion in vitro, and metastasis in both xenograft and spontaneous mouse models of PNET. Treatment of PNET cells with Erk inhibitor or locked nucleic acids sequestering miR-431 inhibits invasion. Four target prediction modules and dual-luciferase reporter assays were used to identify potential mRNA targets of miR-431. A Ras GTPase activating protein tumor suppressor (RasGAP), DAB2 interacting protein (DAB2IP), was discovered as an miR-431 target. Overexpression of DAB2IP's rat homolog, but not its mutant defective in Ras GTPase activating protein activity, reverses miR-431's effect on promoting invasion, Erk phosphorylation, and epithelial-mesenchymal transition of PNETs. Taken together, miR-431 silences DAB2IP to active Ras/Erk and promote metastasis of PNETs. miR-431 may be targeted to manage metastatic PNETs.

Function and clinical relevance of RHAMM isoforms in pancreatic tumor progression.

The receptor for hyaluronic acid-mediated motility (RHAMM) is upregulated in various cancers. We previously screened genes upregulated in human hepatocellular carcinomas for their metastatic function in a mouse model of pancreatic neuroendocrine tumor (PNET) and identified that human RHAMMB promoted liver metastasis. It was unknown whether RHAMMB is upregulated in pancreatic cancer or contributes to its progression. In this study, we found that RHAMM protein was frequently upregulated in human PNETs. We investigated alternative splicing isoforms, RHAMMA and RHAMMB, by RNA-Seq analysis of primary PNETs and liver metastases. RHAMMB, but not RHAMMA, was significantly upregulated in liver metastases. RHAMMB was crucial for in vivo metastatic capacity of mouse and human PNETs. RHAMMA, carrying an extra 15-amino acid-stretch, did not promote metastasis in spontaneous and experimental metastasis mouse models. Moreover, RHAMMB was substantially higher than RHAMMA in pancreatic ductal adenocarcinoma (PDAC). RHAMMB, but not RHAMMA, correlated with both higher EGFR expression and poorer survival of PDAC patients. Knockdown of EGFR abolished RHAMMB-driven PNET metastasis. Altogether, our findings suggest a clinically relevant function of RHAMMB, but not RHAMMA, in promoting PNET metastasis in part through EGFR signaling. RHAMMB can thus serve as a prognostic factor for pancreatic cancer.

Receptor for hyaluronan-mediated motility isoform B promotes liver metastasis in a mouse model of multistep tumorigenesis and a tail vein assay for metastasis.

The gene encoding the receptor for hyaluronan-mediated motility (RHAMM) is overexpressed in many human cancers. However, it is unclear whether RHAMM plays a causal role in tumor initiation or progression. Using somatic gene transfer in a mouse model of islet cell tumorigenesis, we demonstrate that RHAMM isoform B (RHAMM(B)) promotes tumor growth and metastases to lymph nodes and the liver. The propensity of RHAMM(B)-expressing cells to metastasize to the liver was confirmed using an experimental metastasis assay in which cells were injected into the tail vein of immunodeficient mice. However, RHAMM(B) did not increase cell migration or proliferation in culture. In initial efforts to identify signaling pathways activated by RHAMM(B), we found that RHAMM(B) induced phosphorylation of epidermal growth factor receptor (EGFR), Erk1/2, and STAT3 and conferred susceptibility to apoptosis after treatment with an EGFR inhibitor, gefitinib. Taken together, the results indicate that RHAMM(B) promotes hepatic metastasis by islet tumor cells, perhaps through growth factor receptor-mediated signaling.

Defining the ATM-mediated barrier to tumorigenesis in somatic mammary cells following ErbB2 activation.

p53, apoptosis, and senescence are frequently activated in preneoplastic lesions and are barriers to progression to malignancy. These barriers have been suggested to result from an ATM-mediated DNA damage response (DDR), which may follow oncogene-induced hyperproliferation and ensuing DNA replication stress. To elucidate the currently untested role of DDR in breast cancer initiation, we examined the effect of oncogene expression in several murine models of breast cancer. We did not observe a detectable DDR in early hyperplastic lesions arising in transgenic mice expressing several different oncogenes. However, DDR signaling was strongly induced in preneoplastic lesions arising from individual mammary cells transduced in vivo by retroviruses expressing either PyMT or ErbB2. Thus, activation of an oncogene after normal tissue development causes a DDR. Furthermore, in this somatic ErbB2 tumor model, ATM, and thus DDR, is required for p53 stabilization, apoptosis, and senescence. In palpable tumors in this model, p53 stabilization and apoptosis are lost, but unexpectedly senescence remains in many tumor cells. Thus, this murine model fully recapitulates early DDR signaling; the eventual suppression of its endpoints in tumorigenesis provides compelling evidence that ErbB2-induced aberrant mammary cell proliferation leads to an ATM-mediated DDR that activates apoptosis and senescence, and at least the former must be overcome to progress to malignancy. This in vivo study also uncovers an unexpected effect of ErbB2 activation previously known for its prosurvival roles, and suggests that protection of the ATM-mediated DDR-p53 signaling pathway may be important in breast cancer prevention.

Bcl-xL is translocated to the nucleus via CtBP2 to epigenetically promote metastasis.

Besides its mitochondria-based anti-apoptotic role, Bcl-xL also travels to the nucleus to promote cancer metastasis by upregulating global histone H3 trimethyl Lys4 (H3K4me3) and TGFß transcription. How Bcl-xL is translocated into the nucleus and how nuclear Bcl-xL regulates H3K4me3 modification are not understood. Here, we report that C-terminal Binding Protein 2 (CtBP2) binds Bcl-xL via its N-terminus and translocates Bcl-xL into the nucleus. Knockdown of CtBP2 by shRNA decreases the nuclear portion of Bcl-xL and reverses Bcl-xL-induced cell migration and metastasis in mouse models. Furthermore, knockout of CtBP2 suppresses Bcl-xL transcription. The binding between Bcl-xL and CtBP2 is required for their interaction with MLL1, a histone H3K4 methyltransferase. Pharmacologic inhibition of MLL1 enzymatic activity reverses Bcl-xL-induced H3K4me3 and TGFß mRNA upregulation as well as cell invasion. Moreover, cleavage under targets and release using nuclease (CUT&RUN) coupled with next generation sequencing reveals that H3K4me3 modifications are particularly enriched in the promotor region of genes encoding TGFß and its signaling pathway in the cancer cells overexpressing Bcl-xL. Altogether, the metastatic function of Bcl-xL is mediated by its interaction with CtBP2 and MLL1.

cProSite: A web based interactive platform for online proteomics, phosphoproteomics, and genomics data analysis.

We developed cProSite, a website that provides online genomics, proteomics, and phosphoproteomics analysis for the data of The National Cancer Institute's Clinical Proteomic Tumor Analysis Consortium (CPTAC). This tool focuses on comparisons and correlations between different proteins and mRNAs of tumors and normal tissues. Our website is designed with biologists and clinicians in mind, with a user-friendly environment and fast search engine. The search results of cProSite can be used for clinical data validation and provide useful strategic information to identify drug targets at proteomic, phosphoproteomic, or genomic levels. The site is available at http://cprosite.ccr.cancer.gov.

Targeting the Pro-survival Protein BCL-2 to Prevent Breast Cancer.

Current chemopreventive strategies require 3-5 years of continuous treatment and have the concerns of significant side effects; therefore, new chemopreventive agents that require shorter and safer treatments are urgently needed. In this study, we developed a new murine model of breast cancer that mimics human breast cancer initiation and is ideal for testing the efficacy of chemopreventive therapeutics. In this model, introduction of lentivirus carrying a PIK3CA gene mutant commonly found in breast cancers infects a small number of the mammary cells, leading to atypia first and then to ductal carcinomas that are positive for both estrogen receptor and progesterone receptor. Venetoclax is a BH3 mimetic that blocks the anti-apoptotic protein BCL-2 and has efficacy in treating breast cancer. We found that venetoclax treatment of atypia-bearing mice delayed the progression to tumors, improved overall survival, and reduced pulmonary metastasis. Therefore, prophylactic treatment to inhibit the pro-survival protein BCL-2 may provide an alternative to the currently available regimens in breast cancer prevention. PREVENTION RELEVANCE: This study demonstrates that prophylactic treatment with the BCL2-specific antagonist venetoclax prevents breast cancer initiated by a mutated and activated PIK3CA, the most common breast oncogene.

Low-dose carbon monoxide suppresses metastatic progression of disseminated cancer cells.

Low-dose carbon monoxide (CO) is under investigation in clinical trials to treat non-cancerous diseases and has an excellent safety profile. Due to early detection and cancer awareness, an increasing number of cancer patients are diagnosed at early stages, when potentially curative surgical resection can be done. However, many patients ultimately experience recurrence. Here, we evaluate the therapeutic effect of CO on metastatic cancer progression. We show that 250 ppm CO inhibits the migration of multiple types of cancer cell lines, including breast, pancreatic, colon, prostate, liver, and lung cancer and reduces the ability to adhere to fibronectin. We demonstrate that in mouse models, 250 ppm inhaled CO inhibits lung metastasis of breast cancer and liver metastasis of pancreatic cancer. Moreover, low-dose CO suppresses recurrence and increases survival after surgical removal of primary pancreatic cancer in mice. Mechanistically, low-dose CO blocks transcription of heme importers, leading to diminished intracellular heme levels and a heme-regulated enzyme, cytochrome P4501B1 (CYP1B1). Either supplementing heme or overexpressing CYP1B1 reverses the anti-migration effect of low-dose CO. Taken together, low-dose CO therapy inhibits cell migration, reduces adhesion to fibronectin, prevents disseminated cancer cells from expanding into gross metastases, and improves survival in pre-clinical mouse models of metastasis.

STAT5 confers lactogenic properties in breast tumorigenesis and restricts metastatic potential.

Signal transducer and activator of transcription 5 (STAT5) promotes cell survival and instigates breast tumor formation, and in the normal breast it also drives alveolar differentiation and lactogenesis. However, whether STAT5 drives a differentiated phenotype in breast tumorigenesis and therefore impacts cancer spread and metastasis is unclear. We found in two genetically engineered mouse models of breast cancer that constitutively activated Stat5a (Stat5aca) caused precancerous mammary epithelial cells to become lactogenic and evolve into tumors with diminished potential to metastasize. We also showed that STAT5aca reduced the migratory and invasive ability of human breast cancer cell lines in vitro. Furthermore, we demonstrated that STAT5aca overexpression in human breast cancer cells lowered their metastatic burden in xenografted mice. Moreover, RPPA, Western blotting, and studies of ChIPseq data identified several EMT drivers regulated by STAT5. In addition, bioinformatic studies detected a correlation between STAT5 activity and better prognosis of breast cancer patients. Together, we conclude that STAT5 activation during mammary tumorigenesis specifies a tumor phenotype of lactogenic differentiation, suppresses EMT, and diminishes potential for subsequent metastasis.

Metastatic pancreatic neuroendocrine tumors feature elevated T cell infiltration.

Pancreatic neuroendocrine tumors (PNETs) are malignancies arising from the islets of Langerhans. Therapeutic options are limited for the over 50% of patients who present with metastatic disease. We aimed to identify mechanisms to remodel the PNET tumor microenvironment (TME) to ultimately enhance susceptibility to immunotherapy. The TMEs of localized and metastatic PNETs were investigated using an approach that combines RNA-Seq, cancer and T cell profiling, and pharmacologic perturbations. RNA-Seq analysis indicated that the primary tumors of metastatic PNETs showed significant activation of inflammatory and immune-related pathways. We determined that metastatic PNETs featured increased numbers of tumor-infiltrating T cells compared with localized tumors. T cells isolated from both localized and metastatic PNETs showed evidence of recruitment and antigen-dependent activation, suggestive of an immune-permissive microenvironment. A computational analysis suggested that vorinostat, a histone deacetylase inhibitor, may perturb the transcriptomic signature of metastatic PNETs. Treatment of PNET cell lines with vorinostat increased chemokine CCR5 expression by NF-κB activation. Vorinostat treatment of patient-derived metastatic PNET tissues augmented recruitment of autologous T cells, and this augmentation was substantiated in a mouse model of PNET. Pharmacologic induction of chemokine expression may represent a promising approach for enhancing the immunogenicity of metastatic PNET TMEs.

RHAMMB-mediated bifunctional nanotherapy targeting Bcl-xL and mitochondria for pancreatic neuroendocrine tumor treatment.

The incidence of pancreatic neuroendocrine tumor (PNET) has continued to rise. Due to their indolent feature, PNET patients often present with incurable, metastatic diseases. Novel therapies are urgently needed. We have previously shown that Receptor for Hyaluronic Acid-Mediated Motility isoform B (RHAMMB) and Bcl-xL are upregulated in PNETs and both of them promote PNET metastasis. Because RHAMM protein is undetectable in most adult tissues, we hypothesized that RHAMMB could be a gateway for nanomedicine delivery into PNETs. To test this, we developed a RHAMMB-targeting nanoparticle (NP). Inside this NP, we assembled small interfering RNA (siRNA) against Bcl-xL (siBcl-xL) and mitochondria-fusing peptide KLA. We demonstrated that RHAMMB-positive PNETs picked up the RHAMMB-targeting NPs. siBcl-xL or KLA alone killed only 30% of PNET cells. In contrast, a synergistic killing effect was achieved with the co-delivery of siBcl-xL and KLA peptide in vitro. Unexpectedly, siBcl-xL induced cell death before reducing Bcl-xL protein levels. The systemically injected RHAMMB-targeting NPs carrying siBcl-xL and KLA peptide significantly reduced tumor burden in mice bearing RHAMMB-positive PNETs. Together, these findings indicate that the RHAMMB-targeting nanotherapy serves as a promising drug delivery system for PNET and possibly other malignancies with upregulated RHAMMB. The combination of siBcl-xL and KLA peptide can be a therapy for PNET treatment.

High levels of truncated RHAMM cooperate with dysfunctional p53 to accelerate the progression of pancreatic cancer.

Pancreatic cancer has the lowest survival rate out of all types of cancer. Pancreatic cancer patients are often diagnosed at advanced stages, hence an urgent need for a better therapeutic development of this devastating disease. Receptor for hyaluronan-mediated motility (RHAMM), not expressed in adult normal pancreas, has been suggested as a prognostic factor and a potential therapeutic target for pancreatic ductal adenocarcinoma (PDAC) and pancreatic neuroendocrine tumor (PNET). In this study, we initially sought to determine whether genetic deletion of RHAMM would slow down pancreatic cancer progression using Rhamm-/- mice. However, we found that Rhamm-/- mice expressed a truncated HMMRΔexon8-16 protein at higher abundance levels than wild-type RHAMM. While HMMRΔexon8-16 did not enable malignant progression of pancreatic intraepithelial neoplasia in p48-Cre; LSL-KRASG12D mice, it accelerated the formation of invasive PDAC and shortened the survival of p48-Cre; LSL-KRASG12D mice with heterozygous p53 knockout. KrasG12D PDAC mice with homozygous p53 knockout mice died around 10 weeks, and the effect of HMMRΔexon8-16 was not apparent in these short lifespan mice. In addition, HMMRΔexon8-16 shortened the survival of PNET-bearing RIP-Tag mice, which had inactivated p53. In our analysis of TCGA dataset, pancreatic cancer patients with mutant TP53 or loss of one copy of TP53 had higher RHAMM expression, which, combined, predicted worse outcomes. Taken together, by collaborating with dysfunctional p53, high levels of HMMRΔexon8-16 , which lacks the centrosome targeting domain and degrons for interaction with the Anaphase-Promoting Complex (APC), accelerated pancreatic cancer progression.

Assessing tumor progression factors by somatic gene transfer into a mouse model: Bcl-xL promotes islet tumor cell invasion.

Tumors develop through multiple stages, implicating multiple effectors, but the tools to assess how candidate genes contribute to stepwise tumor progression have been limited. We have developed a novel system in which progression of phenotypes in a mouse model of pancreatic islet cell tumorigenesis can be used to measure the effects of genes introduced by cell-type-specific infection with retroviral vectors. In this system, bitransgenic mice, in which the rat insulin promoter (RIP) drives expression of both the SV40 T antigen (RIP-Tag) and the receptor for subgroup A avian leukosis virus (RIP-tva), are infected with avian viral vectors carrying cDNAs encoding candidate progression factors. Like RIP-Tag mice, RIP-Tag; RIP-tva bitransgenic mice develop isolated carcinomas by approximately 14 wk of age, after progression through well-defined stages that are similar to aspects of human tumor progression, including hyperplasia, angiogenesis, adenoma, and invasive carcinoma. When avian retroviral vectors carrying a green fluorescent protein marker were introduced into RIP-Tag; RIP-tva mice by intra-cardiac injection at the hyperplastic or early dysplastic stage of tumorigenesis, approximately 20% of the TVA-positive cells were infected and expressed green fluorescent proteins as measured by flow cytometry. Similar infection with vectors carrying cDNA encoding either of two progression factors, a dominant-negative version of cadherin 1 (dnE-cad) or Bcl-xL, accelerated the formation of islet tumors with invasive properties and pancreatic lymph node metastasis. To begin studying the mechanism by which Bcl-xL, an anti-apoptotic protein, promotes invasion and metastasis, RIP-Tag; RIP-tva pancreatic islet tumor cells were infected in vitro with RCASBP-Bcl-xL. Although no changes were observed in rates of proliferation or apoptosis, Bcl-xL altered cell morphology, remodeled the actin cytoskeleton, and down-regulated cadherin 1; it also induced cell migration and invasion, as evaluated using two-chamber transwell assays. In addition, myosin Va was identified as a novel Bcl-xL-interacting protein that might mediate the effects of Bcl-xL on tumor cell migration and invasion.

Expression of the Receptor for Hyaluronic Acid-Mediated Motility (RHAMM) in Endometrial Cancer is Associated With Adverse Histologic Parameters and Tumor Progression.

Endometrial cancer is one of the most common gynecologic malignancies worldwide. Only 2 agents have been approved by Food and Drug Administration for endometrial cancer since 1971. There is a need to identify molecular targets to treat advanced endometrial cancer. The receptor for hyaluronic acid-mediated motility (RHAMM) is upregulated in various types of cancer. Here, we aimed to determine the clinical significance of RHAMM expression in endometrial cancer. Two hundred twenty-five cases of endometrial cancer, including serous and endometrioid types, and 8 cases of normal endometrium were used for studying RHAMM protein levels. The Cancer Genome Atlas database was also queried for RHAMM mRNA expression in endometrial cancer. Increased expression of RHAMM protein was seen in endometrial cancer compared with no or weak expression in normal endometrium. RHAMM expression positively correlated with tumor grade. RHAMM expression was significantly increased in endometrial serous carcinomas, which are high-grade, aggressive types of endometrial cancer, compared with the relatively less aggressive endometrioid carcinomas. RHAMM expression also correlated with the presence of lymphovascular invasion. RHAMM mRNA expression correlated with decreased survival in The Cancer Genome Atlas cohort. Therefore, increased RHAMM expression in endometrial cancer is associated with high-grade tumors and is indicative of more aggressive behavior. These findings suggest RHAMM as a prognostic factor in endometrial cancer and as a potential therapeutic target in advanced endometrial cancer for future studies.

Functional impact of cancer patient-associated Bcl-xL mutations.

Bcl-xL, an antiapoptotic protein, is frequently overexpressed in cancer to promote survival of tumor cells. However, we have previously shown that Bcl-xL promotes migration, invasion, and metastasis independent of its antiapoptotic function in mitochondria. The pro-metastatic function of Bcl-xL may require its translocation into the nucleus. Besides overexpression, patient-associated mutations of Bcl-xL have been identified in large-scale cancer genomics projects. Understanding the functions of these mutations will guide the development of precision medicine. Here, we selected four patient-associated Bcl-xL mutations, R132W, N136K, R165W, and A201T, to investigate their impacts on antiapoptosis, migration, and nuclear translocation. We found that all four mutation proteins could be detected in both the nucleus and cytosol. Although all four mutations disrupted the antiapoptosis function, one of these mutants, N136K, significantly improved the ability to promote cell migration. These data suggest the importance of developing novel Bcl-xL inhibitors to ablate both antiapoptotic and pro-metastatic functions of Bcl-xL in cancer.

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.

Insights into the biology and treatment strategies of pancreatic neuroendocrine tumors.

Pancreatic neuroendocrine tumors (PNETs) are the second most common primary pancreatic neoplasms after pancreatic ductal adenocarcinoma. PNETs present with widely various clinical manifestation and unfavorable survival rate. The recent advances in next generation sequencing have significantly increased our understanding of the molecular landscape of PNETs and help guide the development of targeted therapies. This review intends to outline a holistic picture of the tumors by discussing current understanding of clinical presentations, up-to-date treatment strategies, novel mouse models, and molecular biology of PNETs. Furthermore, we will provide insight into the future development of more effective targeted therapies that are necessary to manage PNETs.

(-)-Oleocanthal and (-)-oleocanthal-rich olive oils induce lysosomal membrane permeabilization in cancer cells.

(-)-Oleocanthal (oleocanthal) is a phenolic compound found in varying concentrations in extra virgin olive oil oleocanthal has been shown to be active physiologically, benefiting several diseased states by conferring anti-inflammatory and neuroprotective benefits. Recently, we and other groups have demonstrated its specific and selective toxicity toward cancer cells; however, the mechanism leading to cancer cell death is still disputed. The current study demonstrates that oleocanthal, as well as naturally oleocanthal-rich extra virgin olive oils, induced damage to cancer cells' lysosomes leading to cellular toxicity in vitro and in vivo. Lysosomal membrane permeabilization following oleocanthal treatment in various cell lines was assayed via three complementary methods. Additionally, we found oleocanthal treatment reduced tumor burden and extended lifespan of mice engineered to develop pancreatic neuroendocrine tumors. Finally, following-up on numerous correlative studies demonstrating consumption of olive oil reduces cancer incidence and morbidity, we observed that extra virgin olive oils naturally rich in oleocanthal sharply reduced cancer cell viability and induced lysosomal membrane permeabilization while oleocanthal-poor oils did not. Our results are especially encouraging since tumor cells often have larger and more numerous lysosomes, making them especially vulnerable to lysosomotropic agents such as oleocanthal.

Alleles of Insm1 determine whether RIP1-Tag2 mice produce insulinomas or nonfunctioning pancreatic neuroendocrine tumors.

The two most common types of pancreatic neuroendocrine tumors (PanNETs) are insulinomas and nonfunctioning PanNETs (NF-PanNETs). Insulinomas are small, rarely metastatic tumors that secrete high amounts of insulin, and nonfunctioning PanNETs are larger tumors that are frequently metastatic but that do not secrete hormones. Insulinomas are modeled by the highly studied RIP1-Tag2 (RT2) transgenic mice when bred into a C57Bl/6 (B6) genetic background (also known as RT2 B6 mice). But there has been a need for an animal model of nonfunctioning PanNETs, which in the clinic are a more common and severe disease. Here we show that when bred into a hybrid AB6F1 genetic background, RT2 mice make nonfunctioning PanNETs. Compared to insulinomas produced by RT2 B6 mice, the tumors produced by RT2 AB6F1 mice were larger and more metastatic, and the animals did not suffer from hypoglycemia or hyperinsulinemia. Genetic crosses revealed that a locus in mouse chromosome 2qG1 was linked to liver metastasis and to lack of insulin production. This locus was tightly linked to the gene encoding Insm1, a beta cell transcription factor that was highly expressed in human insulinomas but unexpressed in other types of PanNETs due to promoter hypermethylation. Insm1-deficient human cell lines expressed stem cell markers, were more invasive in vitro, and metastasized at higher rates in vivo when compared to isogenic Insm1-expressing cell lines. These data demonstrate that expression of Insm1 can determine whether a PanNET is a localized insulinoma or a metastatic nonfunctioning tumor.

Tissue-specific induced DNA methyltransferase 1 (Dnmt1) in endocrine pancreas by RCAS-TVA-based somatic gene transfer system promotes ß-cell proliferation.

We reported that inactivation of menin (the protein product of MEN1) increases activity of Dnmt1 and mediates DNA hypermethylation in the development of multiple endocrine neoplasia type 1 (MEN1) syndrome. We have developed a RCAS-TVA-based somatic gene transfer system that enables tissue-specific delivery of Dnmt1 to individual ß-cells of the pancreas in a RIP-TVA mouse model. In the present study, we mediated Dnmt1 expression in islet ß-cells in RIP-TVA mice by utilizing the RCAS-TVA system to test if the upregulation of Dnmt1 can promote ß-cell proliferation. In vitro, we demonstrated that upregulation of Dnmt1 increased ß-cell proliferation. In vivo, our results showed that the levels of serum insulin were increased in the RIP-TVA mice with RCASBP-Dnmt1 infection compared with wild-type control mice with RCASBP-Dnmt1 infection. Furthermore, we confirmed that mRNA and protein expression of Dnmt1 as well as Dnmt1 enzyme activity were upregulated in the RIP-TVA mice with RCASBP-Dnmt1 infection compared with wild-type control mice with RCASBP-Dnmt1 infection. Finally, we demonstrated that upregulation of Dnmt1 resulted in hyperplasia through ß-cell proliferation. We conclude that the upregulation of Dnmt1 promotes islet ß-cell proliferation and targeting Dnmt1 may be a promising therapy for patients suffering from pancreatic neuroendocrine tumors.