Phenotype Of Adenocarcinoma Research Articles

Silencing microRNA-330-5p increases MMP1 expression and promotes an invasive phenotype in oesophageal adenocarcinoma

BackgroundMany patients diagnosed with oesophageal adenocarcinoma (OAC) present with advanced disease and approximately half present with metastatic disease. Patients with localised disease, who are managed with curative intent, frequently undergo neoadjuvant chemoradiotherapy. Unfortunately, ~ 70% of patients have little or no response to chemoradiotherapy. We previously identified miR-330-5p as being the most significantly downregulated microRNA in the pre-treatment OAC tumours of non-responders to treatment, but that loss of miR-330-5p had a limited impact on sensitivity to chemotherapy and radiation in vitro. Here, we further examined the impact of miR-330-5p loss on OAC biology.MethodsmiR-330-5p was suppressed in OE33 OAC cells following stable transfection of a vector-driven anti-sense RNA. Whole transcriptome digital RNA-Seq was employed to identify miR-330-5p regulated genes, and qPCR was used for validation. Protein expression was assessed by protein array, Western blotting and zymography. Invasive potential was measured using a transwell assay system. Tumour xenograft growth profile studies were performed in immunocompromised CD1 mice.ResultsIn OE33 cells, suppression of miR-330-5p significantly altered expression of 42 genes, and several secreted proteases. MMP1 gene expression and protein secretion was significantly enhanced with miR-330-5p suppression. This corresponded to enhanced collagen invasion in vitro. In vivo, OE33-derived tumour xenografts with miR-330-5p suppression grew faster than controls.ConclusionsLoss of miR-330-5p expression in OAC tumours may influence tumour cell invasive capacity, tumour growth and therapeutic sensitivity via alterations to the tumour microenvironment.

Abstract 2272: Exploratory analysis of T cell repertoire dynamics upon systemic treatment with the oncolytic virus pelareorep in combination with pembrolizumab and chemotherapy in patients with advanced pancreatic adenocarcinoma

Abstract Background: Pelareorep is an immuno-oncolytic virus that induces an inflamed tumor phenotype in metastatic adenocarcinoma of the pancreas (MAP). Systemically delivered pelareorep in combination with chemotherapy achieves 1 & 2 year-survival rates of 46% & 24% in MAP pts, respectively. Analysis of tumor tissue from patients (pts) treated with pelareorep, chemotherapy and anti-PD-L1 have shown reovirus RNA and protein replication, T-cell infiltration, and upregulation of PD-L1, highlighting that effective T-cell recognition of tumor antigens may be critical to success for this combination therapy. Thus, we hypothesized that pelareorep in combination with chemo and pembrolizumab in pts with MAP would alter the peripheral T-cell repertoire, creating new T-cell clones via the release of novel neoantigens in addition to expanding existing T-cell clones. Methods: A phase 1b study enrolled 11 MAP pts who progressed after first-line treatment. Pts received pelareorep (4.5e10 TCID 50 IV, D1 & D2), plus pembrolizumab (2mg/kg IV, D8) plus either 1) 5-FU (LV (200 mg/m2 /5-FU 200 mg /m2 IV bolus, 5-FU 1200mg/m2 continuous IV infusion D1) or 2) gemcitabine (1000 mg/m2 IV, D1), or 3) irinotecan (125 mg/m2 IV, D1) q3w, until disease progression/unacceptable toxicity. DNA from peripheral blood mononuclear cells from nine patients at cycle 1 day 1 (C1D1) & C2D1 (approx. 3 weeks later) was analyzed using the immunoSEQ® Assay (Adaptive Biotechnologies, Seattle) sequencing the T-cell receptor beta chain region to interrogate changes in the T-cell repertoire. Results: The median Morisita index between C2D1 and C1D1 was 0.83 with 3 samples below 0.6, indicative of significant peripheral repertoire turnover. The median number of expanded clones equated to 45.7 per 100,000 cumulative templates; normal variation over 4 weeks is ~ 5-10 expanded clones. Strikingly, most (median: 86%) peripheral clonal expansion occurred in clones below the limit of detection at C1D1. Cox regression analysis revealed that high peripheral clonality correlates with progression free survival at C1D1 (p=0.01, HR=0.053). Moreover, high clonality correlates with overall survival at both C1D1 (p=0.013, HR=0.124) and C2D1 (p=0.010, HR=0.079). Conclusions: High levels of peripheral T-cell repertoire turnover occur between C1D1 and C2D1. Repertoire turnover is accompanied by significant clonal expansion, mostly by expansion of new clones (i.e. undetected in C1D1). Higher peripheral clonality is associated with better progression free survival at C1D1, and overall survival at C1D1 and C2D1. This research highlights the potential utility of T-cell clonality as a predictive and prognostic biomarker to pelareorep therapy and warrants further clinical investigation. Citation Format: Grey A. Wilkinson, Devalingam Mahalingam, Sukeshi Patel Arora, Paul A. Fields, Patrick Raber, Karol Cheetham, Matt Coffey. Exploratory analysis of T cell repertoire dynamics upon systemic treatment with the oncolytic virus pelareorep in combination with pembrolizumab and chemotherapy in patients with advanced pancreatic adenocarcinoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 2272.

Abstract LB-034: Myc instructs and maintains pancreatic adenocarcinoma phenotype

Abstract Pancreatic ductal adenocarcinoma (PDAC) sporadically develops from pre-cancerous pancreatic intraepithelial neoplasia (PanIN). It has a very poor prognosis and lacks effective therapies. Expression of the pleiotropic transcription factor Myc is deregulated in the majority of human cancers, including PDAC. Using a rapidly and reversibly switchable genetic mouse model to toggle Myc activity, we show that Myc drives immediate transition of indolent KRasG12D-induced PanINs into aggressive pancreatic adenocarcinomas that share all the signature features of spontaneous human PDAC. Myc induces a distinct set of instructive signals, some hitherto unknown, that together orchestrate coordinated changes in the multiple, distinct stromal and inflammatory cell types that generate the characteristic PDAC stroma. We also demonstrate that continued Myc activity is required for PDAC maintenance - Myc deactivation immediately triggers meticulous disassembly of PDAC, including its attendant stroma. Thus, both the formation and deconstruction of the complex PDAC phenotype can be mediated by a single, reversible molecular switch. We have also identified a previously unknown Myc-dependent GAS6-AXL pathway that is required for stromal changes characteristic of PDAC. Using the same pre-clinical mouse PDAC model, we show that TP-0903, a specific inhibitor of AXL kinase, profoundly blocks Myc-induced influx of neutrophils into PanINs and B220+ lymphoid cells into the PanIN periphery. This significantly retards Myc-driven stellate cell activation and proliferation and impedes PDAC growth. Hence, the GAS6-AXL-mediated signaling pathway is a promising therapeutic target for pancreatic ductal adenocarcinoma. TP-0903 is currently in a Phase I/Ib clinical study in in patients with solid tumors, including PDAC. Citation Format: Nicole M. Sodir, Roderik M. Kortlever, Valentin J.A. Barthet, Luca Pellegrinet, Tania Campos, Steven Kupczak, Laura Soucek, Mark J. Arends, Trevor D. Littlewood, Gerard I. Evan. Myc instructs and maintains pancreatic adenocarcinoma phenotype [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr LB-034.

Morphological characteristics of mucinous adenocarcinoma of the colon and its embryogenetic premises

This review reveals the anatomical, physiological and embryogenetic features of the proximal colon to explain the reasons for such frequent localization of mucinous phenotype of adenocarcinoma (MAC) here. It was shown that later differentiation in embryogenesis causes relative insufficiency of the proximal part as a structure of the digestive and immune systems. Physiologically lower lymphoid tissue (GALT) density here leads to the formation of a certain composition of the intestinal microbiota, which is different from that in the distal part, which is a significant link in the etiopathogenesis of proximal colon cancer. It is confirmed also by different composition of biofilms on the surface of epithelial tumors in the right and left halves of the colon, as well as differences in the molecular mechanisms of carcinogenesis, depending on the location of the cancer. Common for proximal part genetic changes, called as CIMP-phenotype, microsatellite instability and BRAF proto-oncogene mutation lead to excessive secretion of specific mucin fractions (mainly MUC2 and MUC5AC), the imbalance of its composition and the formation of MAC. An earlier age of onset, frequent association with hereditary non-polypous colorectal carcinoma, the predominance of MUC2 and MUC5AC fractions, similar to the embryonic period, as well as a higher level of cancer-embryonic antigen in patients with MAC indicate the influence of anatomical, physiological and embryogenetic features of the proximal colon on carcinogenesis long before its formation. Thus, a detailed understanding of MAC carcinogenesis is necessary for an adequate assessment of its effective prevention in time, as well as dealing with it as with specific nosological unit requiring specific treatment principles.

METASTASIS AROUND DENTAL IMPLANTS MASCARAING AS PERI-IMPLANTITIS - A WOLF IN SHEEP'S CLOTHING

Background Placement of dental implants is a common procedure with a high success rate. A minority of the cases fail, however, a manifestation termed peri-implantitis, which is considered clinically obvious, hence often not biopsied. Case report In this report, a case of metastatic lung adenocarcinoma mimicking peri-implantitis adjacent two dental implants is presented. The lesion occurred eight months after the surgical implantation procedure and by that time, the underlying malignancy was unknown to the patient. During the surgical procedure itself’, and even in the follow up meetings, there was no apparent clinical or radiographic sign of a metastatic disease, but during an eight months follow up session, the soft tissue around the implants was firmly swelled and mildly erythematous, and the x ray imaging revealed a nonspecific ill-defined radiolucency of the alveolar bone. The tissue was excised, adjacent bone was curated and the tissue was submitted to histopathological analysis. The pathological picture presented a malignant tumor composed of epithelial islands embedded within a fibrous stroma. The epithelial islands presented atypical features and an immunohistochemical phenotype of lung adenocarcinoma that included positive pan keratin, thyroid transcription factor-1 and napsin-A and negative thyroglobulin and prostatic specific antigen. Concurrently, the patient was diagnosed with a lung adenocarcinoma and started systemic treatment. Conclusion This case of metastatic disease masquerading as peri-implantitis reflects the importance of submitting any tissue to histopathological assessment.

Transcriptomic profiling reveals three molecular phenotypes of adenocarcinoma at the gastroesophageal junction.

Cancers occurring at the gastroesophageal junction (GEJ) are classified as predominantly esophageal or gastric, which is often difficult to decipher. We hypothesized that the transcriptomic profile might reveal molecular subgroups which could help to define the tumor origin and behavior beyond anatomical location. The gene expression profiles of 107 treatment‐naïve, intestinal type, gastroesophageal adenocarcinomas were assessed by the Illumina‐HTv4.0 beadchip. Differential gene expression (limma), unsupervised subgroup assignment (mclust) and pathway analysis (gage) were undertaken in R statistical computing and results were related to demographic and clinical parameters. Unsupervised assignment of the gene expression profiles revealed three distinct molecular subgroups, which were not associated with anatomical location, tumor stage or grade (p > 0.05). Group 1 was enriched for pathways involved in cell turnover, Group 2 was enriched for metabolic processes and Group 3 for immune‐response pathways. Patients in group 1 showed the worst overall survival (p = 0.019). Key genes for the three subtypes were confirmed by immunohistochemistry. The newly defined intrinsic subtypes were analyzed in four independent datasets of gastric and esophageal adenocarcinomas with transcriptomic data available (RNAseq data: OCCAMS cohort, n = 158; gene expression arrays: Belfast, n = 63; Singapore, n = 191; Asian Cancer Research Group, n = 300). The subgroups were represented in the independent cohorts and pooled analysis confirmed the prognostic effect of the new subtypes. In conclusion, adenocarcinomas at the GEJ comprise three distinct molecular phenotypes which do not reflect anatomical location but rather inform our understanding of the key pathways expressed.

Epithelial-mesenchymal Transition Phenotypes in Vertebral Metastases of Lung Cancer.

Vertebral metastases of non-small cell lung cancer (NSCLC) are frequently diagnosed in the metastatic setting and are commonly identified in the thoracic vertebrae in patients. Treatment of NSCLC bone metastases, which are often multiple, is palliative, and the median survival times are 3 to 6 months. We have characterized spontaneous vertebral metastases in a brain metastases model of NSCLC and correlated these findings with epithelial-mesenchymal transition (EMT). Brain metastases were established in athymic nude mice following intracardiac injection of brain-seeking adenocarcinoma NSCLC cells. Thirty-nine percent of mice (14/36) developed spontaneous vertebral metastases, spinal cord compression, and hind-limb paralysis. Vertebral metastases consisted of an adenocarcinoma phenotype with neoplastic epithelial cells arranged in cords or acini and a mesenchymal phenotype with spindloid neoplastic cells arranged in bundles and streams. Quantitative and qualitative immunohistochemical and immunofluorescence assays demonstrated an increase in vimentin expression compared to cytokeratin expression in vertebral metastases. A correlation with EMT was supported by an increase in CD44 in vertebral metastases and parenchymal metastases. These data demonstrate a translational lung cancer metastasis model with spontaneous vertebral metastasis. The mesenchymal and epithelial phenotype of these spontaneous metastases coupled with EMT provide a conduit to improve drug delivery and overall patient survival.

Mo1005 – Heartburn and the American College of Gastroenterology Criteria for Barrett’s Screening are Limited in Detecting De Novo and Non-Intestinal Metaplasia Phenotype of Esophageal Adenocarcinoma

Mo1005 – Heartburn and the American College of Gastroenterology Criteria for Barrett’s Screening are Limited in Detecting De Novo and Non-Intestinal Metaplasia Phenotype of Esophageal Adenocarcinoma

Strawberry tree honey as a new potential functional food. Part 2: Strawberry tree honey increases ROS generation by suppressing Nrf2-ARE and NF-кB signaling pathways and decreases metabolic phenotypes and metastatic activity in colon cancer cells

Abstract The aim of the present study was to evaluate the effects of Strawberry tree honey (STH) on oxidative stress, metabolic phenotype, migration, invasion and epithelial-mesenchymal transition in adenocarcinoma (HCT-116) and metastatic (LoVo) colon cancer cells as well as in human dermal fibroblasts (HDF). Significant oxidative stress was observed through the increase of intracellular ROS generation, lipid and protein damage and reduction of antioxidant enzyme activities in colon cancer cells; in HDF these effects were limited or none. The expression of NF-кB, p-IкBα, Nrf2 was suppressed after STH treatment in colon cancer cells. All the parameters of mitochondrial respiration and glycolysis were reduced after STH treatment in cancer cells, while they were unchanged in HDF. Wound-closure percentages and the expression of MMP-2, MMP-9, N-cadherin, β-catenin decreased, while those of E-cadherin increased after STH treatment in colon cancer cells. Thus, STH can be used for its potential in cancer prevention.

ALDH1 expression correlates with an epithelial-like phenotype and favorable prognosis in lung adenocarcinoma: a study based on immunohistochemistry and mRNA expression data.

Cancer stem cells (CSC) and epithelial-mesenchymal transition (EMT) pathways are crucial for cancer progression. However, synergistic interactions between CSC and EMT are not clear in non-small cell lung cancer (NSCLC). The objective of this study was to investigate CSC markers such as CD44, NANOG, and ALDH1 expression and its correlation with EMT markers in NSCLC patients. Its association with survival was also determined. CD44, NANOG, and ALDH1 protein expression was evaluated in 267 resected NSCLC and its correlation with e-cadherin, β-catenin, p120 catenin, vimentin, SNAIL, and TWIST expressions was determined based on immunohistochemical and mRNA expression data from The Cancer Genome Atlas (TCGA) database. Survival analyses also were performed based on immunohistochemistry and mRNA expression data from Gene Expression Omnibus dataset. ALDH1 expression in lung adenocarcinoma was positively correlated with the epithelial-like phenotype, low vimentin and low TWIST in immunohistochemical and mRNA expression data. NANOG and ALDH1 expressions measured by immunohistochemical and mRNA expression profiling data of adenocarcinomas were associated with a favorable prognosis. ALDH1 was an independent favorable prognostic marker for overall survival or recurrence-free survival in adenocarcinoma (P = 0.026 and P = 0.033, respectively). The epithelial-like phenotype expressing P120-catenin and beta-catenin was associated with a favorable prognosis; however, the TWIST-expressing mesenchymal-like phenotype was correlated with an unfavorable prognosis. NANOG and ALDH1 protein or mRNA expression showed improved prognosis in adenocarcinoma alone. ALDH1 expression correlated with an epithelial-like phenotype.

Migrated T lymphocytes into malignant pleural effusions: an indicator of good prognosis in lung adenocarcinoma patients

The presence of leukocyte subpopulations in malignant pleural effusions (MPEs) can have a different impact on tumor cell proliferation and vascular leakiness, their analysis can help to understand the metastatic microenvironment. We analyzed the relationship between the leukocyte subpopulation counts per ml of pleural fluid and the tumor cell count, molecular phenotype of lung adenocarcinoma (LAC), time from cancer diagnosis and previous oncologic therapy. We also evaluated the leukocyte composition of MPEs as a biomarker of prognosis. We determined CD4+ T, CD8+ T and CD20+ B cells, monocytes and neutrophils per ml in pleural effusions of 22 LAC and 10 heart failure (HF) patients by flow cytometry. Tumor cells were identified by morphology and CD326 expression. IFNγ, IL-10 and IL-17, and chemokines were determined by ELISAs and migratory response to pleural fluids by transwell assays. MPEs from LAC patients had more CD8+ T lymphocytes and a tendency to more CD4+ T and CD20+ B lymphocytes than HF-related fluids. However, no correlation was found between lymphocytes and tumor cells. In those MPEs which were detected >1 month from LAC diagnosis, there was a negative correlation between pleural tumor cells and CD8+ T lymphocytes. CXCL10 was responsible for the attraction of CD20+ B, CD4+ T and CD8+ T lymphocytes in malignant fluids. Concentrations of IL-17 were higher in MPEs than in HF-related effusions. Survival after MPE diagnosis correlated positively with CD4+ T and CD8+ T lymphocytes, but negatively with neutrophils and IL-17 levels. In conclusion, lymphocyte enrichment in MPEs from LAC patients is mostly due to local migration and increases patient survival.

Super-enhancers: novel target for pancreatic ductal adenocarcinoma.

Super-enhancers (SEs) are unique areas of the genome which drive high-level of transcription and play a pivotal role in the cell physiology. Previous studies have established several important genes in cancer as SE-driven oncogenes. It is likely that oncogenes may hack the resident tissue regenerative program and interfere with SE-driven repair networks, leading to the specific pancreatic ductal adenocarcinoma (PDAC) phenotype. Here, we used ChIP-Seq to identify the presence of SE in PDAC cell lines. Differential H3K27AC marks were identified at enhancer regions of genes including c-MYC, MED1, OCT-4, NANOG, and SOX2 that can act as SE in non-cancerous, cancerous and metastatic PDAC cell lines. GZ17-6.02 affects acetylation of the genes, reduces transcription of major transcription factors, sonic hedgehog pathway proteins, and stem cell markers. In accordance with the decrease in Oct-4 expression, ChIP-Seq revealed a significant decrease in the occupancy of OCT-4 in the entire genome after GZ17-6.02 treatment suggesting the possible inhibitory effect of GZ17-6.02 on PDAC. Hence, SE genes are associated with PDAC and targeting their regulation with GZ17-6.02 offers a novel approach for treatment.

CME Exam 2: Identification of Prognostic Phenotypes of Esophageal Adenocarcinoma in 2 Independent Cohorts

CME Exam 2: Identification of Prognostic Phenotypes of Esophageal Adenocarcinoma in 2 Independent Cohorts

Pyruvate Treatment Restores the Effectiveness of Chemotherapeutic Agents in Human Colon Adenocarcinoma and Pleural Mesothelioma Cells.

Emerging evidence supports the idea that a dysfunction in cell metabolism could sustain a resistant phenotype in cancer cells. As the success of chemotherapeutic agents is often questioned by the occurrence of multidrug resistance (MDR), a multiple cross-resistance towards different anti-cancer drugs represent a major obstacle to cancer treatment. The present study has clarified the involvement of the carbon metabolites in a more aggressive tumor colon adenocarcinoma phenotype and in a chemoresistant mesothelioma, and the role of pyruvate treatment in the reversion of the potentially related resistance. For the first time, we have shown that human colon adenocarcinoma cells (HT29) and its chemoresistant counterpart (HT29-dx) displayed different carbon metabolism: HT29-dx cells had a higher glucose consumption compared to HT29 cells, whereas human malignant mesothelioma (HMM) cells showed a lower glucose consumption compared to HT29 cells, accompanied by a lower pyruvate production and, consequently, a higher production of lactate. When treated with pyruvate, both HT29-dx and HMM cells exhibited a re-established accumulation of doxorubicin and a lower survival ability, a decreased activity of multidrug resistance protein 1 (MRP1) and a restored mitochondrial respiratory chain function, improving the effectiveness of the chemotherapeutic agents in these resistant cancer cells.

Identification of Prognostic Phenotypes of Esophageal Adenocarcinoma in 2 Independent Cohorts

Most patients with esophageal adenocarcinoma (EAC) present with de novo tumors. Although this could be due to inadequate screening strategies, the precise reason for this observation is not clear. We compared survival of patients with prevalent EAC with and without synchronous Barrett esophagus (BE) with intestinal metaplasia (IM) at the time of EAC diagnosis. Clinical data were studied using Cox proportional hazards regression to evaluate the effect of synchronous BE-IM on EAC survival independent of age, sex, TNM stage, and tumor location. Weanalyzed data from a cohort of patients with EAC from the Mayo Clinic (n=411; 203 with BE and IM) and a multicenter cohort from the United Kingdom (n=1417; 638 with BE and IM). In the Mayo cohort, BE with IM had a reduced risk of death compared to patients without BE and IM (hazard ratio [HR] 0.44; 95% CI, 0.34-0.57; P<.001). In a multivariable analysis, BE with IM was associated with longersurvival independent of patient age or sex, tumor stage or location, and BE length (adjusted HR, 0.66; 95% CI, 0.5-0.88; P=.005). In the United Kingdom cohort, patients BE and IM had a reduced risk of death compared with those without (HR, 0.59; 95% CI, 0.5-0.69; P<.001), with continued significance in multivariable analysis that included patient age and sex and tumor stage and tumor location (adjusted HR, 0.77; 95% CI, 0.64-0.93; P=.006). Two types of EAC can be characterized based on the presence or absence of BE. These findings could increase our understanding the etiology of EAC, and be used in management and prognosis ofpatients.

Overexpression of KIF20A confers malignant phenotype of lung adenocarcinoma by promoting cell proliferation and inhibiting apoptosis

Increasing studies showed that kinesin family member 20A (KIF20A) was overexpessed in several types of cancer, and its overexpression correlated with the oncogenesis and prognosis of cancers. However, little is known about the role of KIF20A in lung adenocarcinoma (LUAD). In this study, we employed the bioinformatics analysis to identify the upregulation of KIF20A in LUAD, then verified the results in human tumor specimens and LUAD cell lines. Compared with normal lung tissues, a ubiquitous upregulation of KIF20A was observed in LUAD tissues by immunohistochemistry (IHC) as well as TCGA analysis. Higher expression of KIF20A was significantly associated with more advanced clinicopathological features and shorter overall survival (OS). Moreover, multivariate Cox regression analysis revealed that KIF20A was an independent prognostic factor for OS. The expression of KIF20A was significantly elevated in LUAD cell lines. After silencing KIF20A, lung cancer cell cycle arrested in G1 phase and apoptosis increased. The same results were observed in vivo. Thus, our study demonstrated that KIF20A might confer malignant phenotype to LUAD by regulating cell proliferation and apoptosis, providing a new potential biomarker for clinical treatment of LUAD.

Abstract 3178: Up-regulation of GLI1 expression in drug resistant rhabdomyosarcoma (RMS) and Ewing sarcoma (EWS) cell lines

Abstract Outcome for pediatric patients with recurrent sarcomas remains poor. Therefore, understanding mechanisms that lead to drug resistance and ways to overcome drug resistance therapeutically are essential. Recent reports have suggested a role for the Hedgehog signaling pathway and its downstream mediator GLI1 in acquisition of a multidrug resistance phenotype in esophageal adenocarcinoma, glioblastoma, and myeloid leukemia. Since GLI1 is expressed in the majority of RMS and EWS cases, we hypothesized that GLI1 up-regulation occurs as cells become drug resistant in vitro and in vivo. We established vincristine (VCR) resistant RMS cell lines (Embryonal RMS: RD, Ruch-2; Alveolar RMS: Rh30, Rh41) by serial exposure to increasing concentrations of VCR. Drug-resistance was defined as an IC50 of ≥30 fold of the baseline that was established using parental cells. In addition, we obtained EWS cell lines, representing untreated cases (CHLA-9 and TC-32) and cases following recurrence (CHLA-258 and TC-71). We used an 86-gene cancer drug resistance PCR array (Qiagen), to characterize gene expression differences between the parental vs. VCR-resistant RMS cells lines and between the untreated and recurrent EWS cell lines. GLI1 expression was significantly increased (p≤0.05) in VCR-resistant alveolar RMS cell lines (Rh30 [2.3 fold], and Rh41 [10.3 fold]) compared with parental cells. Expression of 2 additional genes (MDR1 [13,307 fold in Rh30 and 1,755 fold in Rh41 cells] and MVP [2.4 fold in Rh30 and 8.8 fold in Rh41]) was also significantly up-regulated in these cells. Gel mobility shifts showed interaction of GLI1 with the MVP promoter, and cotransfection assays showed that GLI1 up-regulates reporter gene expression through the MVP promoter. GLI1 expression was significantly up-regulated in Ruch-2 embryonal RMS cells (2 fold). We also showed higher GLI1 expression in recurrent EWS cell lines (TC-71 [3.4 fold] and CHLA-258 [4.8 fold]) compared with untreated CHLA-9 cells. Expression of 1 additional gene (androgen receptor [11.1 fold in CHLA-258 cells and 274.6 fold in TC-71 cells]) was also significantly up-regulated in CHLA-258 and TC-71 cells. Treatment of VCR-resistant UKF Rhb-1 RMS cells with VCR together with either GANT61 or Cpd#33 (GLI1 inhibitors) significantly decreased cell viability by MTT assay at doses that did not reduce cell viability individually. Our results suggest that GLI1 up-regulation contributes to development of drug-resistance in RMS and EWS and that GLI1-inhibitors may reduce multidrug resistance. Citation Format: Joon Won Yoon, Iris Smith, Sumeed Manzoor, Marilyn Lamm, Philip Iannaccone, David Walterhouse. Up-regulation of GLI1 expression in drug resistant rhabdomyosarcoma (RMS) and Ewing sarcoma (EWS) cell lines [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 3178.

SALL4 suppresses reactive oxygen species in pancreatic ductal adenocarcinoma phenotype via FoxM1/Prx III axis

Pancreatic ductal adenocarcinoma (PDAC) is a major malignant phenotype in pancreatic cancer, which is one of the most death causes by cancer in the world. PDAC developed from pancreatic intra-epithelial neoplasms (PanINs) and poorly diagnosed at early stages. Beside of high drug resistance, metastasis is the great concern during pancreatic cancer treatment. SALL4 expression is inherent in the upregulations of endothelial mesenchymal transition (EMT) genes and therefore promoting cancer metastasis. Furthermore, some of evidences indicated reactive oxygen species (ROS) is also influent to metastasis and self-antioxidant capacity seems a gold standard for successful metastasis rate. In this study, we have found the role Spalt like protein 4 (SALL4) to PDAC proliferation, mobility and its regulation to mitochondrial ROS via FoxM1/Prx III axis. It is possible that SALL4 mainly induces endothelial-mesenchymal transition (EMT) phenotype and favors ROS loss to facilitate metastasis efficiency in PDAC cells. Therefore, SALL4 might be a promising marker for PDAC treatment and targeting SALL4 would benefit anti-proliferative and anti-metastasis therapies.

KrasG12D-LOH promotes malignant biological behavior and energy metabolism of pancreatic ductal adenocarcinoma cells through the mTOR signaling pathway.

Oncogenic Kras with loss of heterozygosity (LOH) is frequently detected in various tumours. However, the exact function and mechanism by which KrasG12D-LOH operates remain unclear. Therefore, the current study investigated the effect of KrasG12D-LOH on the malignant phenotype of pancreatic ductal adenocarcinoma (PDAC) cells. Our investigation revealed that KrasG12D-LOH is associated with increased proliferation, invasion and reduced apoptosis in PDAC cells. The results also exhibited enhanced glycolytic phenotype of KrasG12D-LOH PDAC cells. Hyperactive mTOR plays a significant role in the initiation and maintenance of tumors. To investigate the correlation between KrasG12D-LOH and mTOR, the mTOR signaling pathway was detected by western blot analysis. We found that KrasG12D-LOH up-regulated Akt, AMPK, REDD1 and mTOR in PDAC cells. In summary, our results demonstrated that KrasG12D-LOH promotes oncogenic Kras-induced PDAC by regulating energy metabolism and mTOR signaling pathway. These data may provide novel therapeutic perspectives for PDAC.

P2.02-060 Prognostic Significance of EDIL3 Expression and Correlation with Mesenchymal Phenotype and Microvessel Density in Lung Adenocarcinoma

P2.02-060 Prognostic Significance of EDIL3 Expression and Correlation with Mesenchymal Phenotype and Microvessel Density in Lung Adenocarcinoma