Effect of altering fibroblast integrin associated protein expression on the growth and protein expression of pancreas cancer cells.

2013 ◽  
Vol 31 (4_suppl) ◽  
pp. 251-251
Author(s):  
Jill E. Shea ◽  
Kelly C. Hewitt ◽  
Courtney L. Scaife
Keyword(s):  
Protein Expression ◽  
Cancer Cells ◽  
Patient Outcomes ◽  
Fluorescent Protein ◽  
Ductal Adenocarcinoma ◽  
Associated Proteins ◽  
Stromal Response ◽  
Growth Promoting ◽  
Gapdh Protein ◽  
Transwell System

251 Background: The lack of success of current treatments for pancreatic ductal adenocarcinoma (PDA) may be due in part to the presence of the dense surrounding stromal response and the interactions between the stroma and cancer cells. We have shown that integrin associated proteins, in particular PINCH, are expressed to a higher degree in the stroma adjacent to the tumor cells and PINCH expression is positively correlated with poorer PDA patient outcomes. We hypothesize that decreasing PINCH protein expression in the tumor associated stroma will decrease the growth and expression of growth promoting proteins in PDA cells. Methods: Red fluorescent protein (RFP) stably expressing MiaPaCa-2 cells were co-cultured with WI38 fibroblasts, or WI38 fibroblasts with shRNA knockdown of PINCH protein (PINCH KD). Cell growth of the PDA cells alone and upon exposure to WI38 and PINCH KD fibroblasts was determined using RFP at 48 hours. PINCH, Akt, and pAKt protein expression in cultured PDA cells, exposed to media, WI38, or WI38 PINCH KD cells via a transwell system, were determined by western blot with values normalized to GAPDH protein expression at 48 hours. Results: MiaPaCa-2 cells grown in co-culture with WI38 cells had a 70%±5% increase in RFP, while those grown in co-culture with WI38 PINCH KD cells had a 40%±3% increase relative to MiaPaCa2 cells grown alone. In terms of relative PINCH protein expression there was an increase in MiaPaCa-2 (20%±5%) cells grown in co-culture with WI38 cells compared to alone but not when grown with PINCH KD (6%±3%). MiaPaCa-2 cells had a greater increase in AKT (47%±6% compared with 19%±4%) and pAKT (36%±5 and 22%±4%) in the cells grown with WI38 cells compared to PDA cells grown with WI38 PINCH KD. Conclusions: Reductions in fibroblast PINCH protein expression are associated with reductions in the growth of adjacent PDA cells, as well as the expression of growth enhancing proteins (PINCH, AKT, and pAKT). Since greater PINCH expression within PDA stromal cells is associated with poorer patient outcomes, understanding the mechanisms associated with this tumor-stromal interaction may provide intervention opportunities.

Frequent Detection and Immunophenotyping of Prostate-Derived Cell Clusters in the Peripheral Blood of Prostate Cancer Patients

2004 ◽  
Vol 19 (2) ◽  
pp. 93-99 ◽  
Author(s):  
H. Schmidt ◽  
G. De Angelis ◽  
O. Bettendorf ◽  
E. Eltze ◽  
A. Semjonow ◽  
...  
Keyword(s):  
Prostate Cancer ◽  
Protein Expression ◽  
Cancer Cells ◽  
Cancer Patients ◽  
Peripheral Blood ◽  
Preoperative Staging ◽  
Specific Antigen ◽  
Cell Clusters ◽  
Cancer Aggressiveness ◽  
Associated Proteins

Background Recent scientific studies have failed to determine parameters for the assessment of prostate cancer aggressiveness. The present study deals with the detection of blood-borne cancer cells based on polymerase chain reaction (PCR) and cell enrichment methods. The contradictory results reported in the literature have called into question the clinical usefulness of this diagnostic method in the preoperative staging of clinically localized prostate cancer. Methods We established a combined method of density gradient centrifugation and immunomagnetic separation using epithelium-specific antibodies, i.e. cytokeratins, to isolate prostate-derived circulating cells from the peripheral blood of patients with prostate cancer. Isolated cells were characterized by DNA staining and immunocytochemistry using antibodies for the detection of prostate-specific antigen (PSA), proliferation-associated proteins (MIB-1, H1 and H3) and apoptosis-associated proteins (M30, c-FasR). Results We applied these methods to 68 prostate cancer patients and were able to isolate cell clusters in 98%. Immunophenotypic and morphological characterization of PSA-positive prostate-derived cell clusters found in the peripheral blood of prostate cancer patients showed two main populations: 1) in 35% of the investigated prostate cancer patients we detected rounded cell aggregates of probable cancer cells expressing proliferation-associated proteins and lacking apoptosis-associated protein expression; 2) in all cases there was a high frequency of circulating dysmorphic cell clusters positive for apoptosis-associated protein expression. Conclusion Our results demonstrate the existence of at least two different types of blood-borne prostate-derived circulating cell clusters. Of these, only the less frequent, round, small cell clusters harbor features that are probably necessary for the cells to survive for metastatic spread.

Galectin-3 is modulated in pancreatic cancer cells under hypoxia and nutrient deprivation

2020 ◽  
Vol 401 (10) ◽  
pp. 1153-1165 ◽  
Author(s):  
Antônio F. da Silva Filho ◽  
Lucas B. Tavares ◽  
Maira G. R. Pitta ◽  
Eduardo I. C. Beltrão ◽  
Moacyr J. B. M. Rêgo
Keyword(s):  
Pancreatic Cancer ◽  
Cell Death ◽  
Mrna Expression ◽  
Cancer Cells ◽  
Ductal Adenocarcinoma ◽  
Reverse Transcription Pcr ◽  
Pancreatic Cancer Cells ◽  
Through Flow ◽  
Galectin 3

AbstractPancreatic ductal adenocarcinoma is one of the most aggressive tumors with a microenvironment marked by hypoxia and starvation. Galectin-3 has been evaluated in solid tumors and seems to present both pro/anti-tumor effects. So, this study aims to characterize the expression of Galectin-3 from pancreatic tumor cells and analyze its influence for cell survive and motility in mimetic microenvironment. For this, cell cycle and cell death were accessed through flow cytometry. Characterization of inside and outside Galectin-3 was performed through Real-Time Quantitative Reverse Transcription PCR (qRT-PCR), immunofluorescence, Western blot, and ELISA. Consequences of Galectin-3 extracellular inhibition were investigated using cell death and scratch assays. PANC-1 showed increased Galectin-3 mRNA expression when cultivated in hypoxia for 24 and 48 h. After 24 h in simultaneously hypoxic/deprived incubation, PANC-1 shows increased Galectin-3 protein and secreted levels. For Mia PaCa-2, cultivation in deprivation was determinant for the increasing in Galectin-3 mRNA expression. When cultivated in simultaneously hypoxic/deprived condition, Mia PaCa-2 also presented increasing for the Galectin-3 secreted levels. Treatment of PANC-1 cells with lactose increased the death rate when cells were incubated simultaneously hypoxic/deprived condition. Therefore, it is possible to conclude that the microenvironmental conditions modulate the Galectin-3 expression on the transcriptional and translational levels for pancreatic cancer cells.

scholarly journals The Prognostic Impact of HER2 Genetic and Protein Expression in Pancreatic Carcinoma—HER2 Protein and Gene in Pancreatic Cancer

Diagnostics ◽  
2021 ◽  
Vol 11 (4) ◽  
pp. 653
Author(s):  
Song-Hee Han ◽  
Ki Hyun Ryu ◽  
Ah-Young Kwon
Keyword(s):  
Protein Expression ◽  
Genetic Heterogeneity ◽  
Growth Factor Receptor ◽  
Ductal Adenocarcinoma ◽  
Prognostic Impact ◽  
Her2 Expression ◽  
Her2 Protein ◽  
Epidermal Growth ◽  
Her2 Heterogeneity

Pancreatic ductal adenocarcinoma (PDAC) is a lethal and clinically heterogeneous disease with a limited benefit from human epidermal growth factor receptor 2 (HER2)-targeted therapy. Recently, some studies have addressed the antitumoral effect of novel anti-HER2 drugs in HER2 low-expressing tumors. However, there have been few studies on the significance of low HER2 expression and genetic heterogeneity in PDAC. Using immunohistochemistry and dual-color silver-enhanced in situ hybridization based on the Trastuzumab for a gastric cancer scoring scheme, we evaluated HER2 protein expression, gene amplification, and genetic heterogeneity in three groups (HER2-neg, HER2-low, HER2-pos) of 55 patients. Among the 55 cases, 41.8% (23/55) showed HER2 expression of any intensity. HER2 amplification independent of HER2 expression was 25.5% (14/55). Patients in both these groups had a shorter overall survival than did patients in the HER2-neg group. HER2 genetic heterogeneity was identified in 37 (70.9%) of the 55 cases, mainly in HER2-neg and HER2-low groups. HER2 genetic heterogeneity significantly correlated with worse survival in the HER2-low and HER2-neg groups of PDAC. These findings support the hypothesis that low-level HER2 expression and heterogeneity have significant clinical implications in PDAC. HER2 heterogeneity might indicate the best strategies of combination therapies to prevent the development of subdominant clones with resistance potential.

scholarly journals Inhibition of mutant KRAS-driven overexpression of ARF6 and MYC by an eIF4A inhibitor drug improves the effects of anti-PD-1 immunotherapy for pancreatic cancer

2021 ◽  
Vol 19 (1) ◽  
Author(s):  
Ari Hashimoto ◽  
Haruka Handa ◽  
Soichiro Hata ◽  
Akio Tsutaho ◽  
Takao Yoshida ◽  
...  
Keyword(s):  
Cancer Cells ◽  
Mrna Translation ◽  
Immune Checkpoint Blockade ◽  
Small Gtpase ◽  
Initiation Factor ◽  
Ductal Adenocarcinoma ◽  
Cancer Type ◽  
Intact Cells ◽  
Cell Functions

AbstractMany clinical trials are being conducted to clarify effective combinations of various drugs for immune checkpoint blockade (ICB) therapy. However, although extensive studies from multiple aspects have been conducted regarding treatments for pancreatic ductal adenocarcinoma (PDAC), there are still no effective ICB-based therapies or biomarkers for this cancer type. A series of our studies have identified that the small GTPase ARF6 and its downstream effector AMAP1 (also called ASAP1/DDEF1) are often overexpressed in different cancers, including PDAC, and closely correlate with poor patient survival. Mechanistically, the ARF6-AMAP1 pathway drives cancer cell invasion and immune evasion, via upregulating β1-integrins and PD-L1, and downregulating E-cadherin, upon ARF6 activation by external ligands. Moreover, the ARF6-AMAP1 pathway enhances the fibrosis caused by PDAC, which is another barrier for ICB therapies. KRAS mutations are prevalent in PDACs. We have shown previously that oncogenic KRAS mutations are the major cause of the aberrant overexpression of ARF6 and AMAP1, in which KRAS signaling enhances eukaryotic initiation factor 4A (eIF4A)-dependent ARF6 mRNA translation and eIF4E-dependent AMAP1 mRNA translation. MYC overexpression is also a key pathway in driving cancer malignancy. MYC mRNA is also known to be under the control of eIF4A, and the eIF4A inhibitor silvestrol suppresses MYC and ARF6 expression. Using a KPC mouse model of human PDAC (LSL-Kras(G12D/+); LSL-Trp53(R172H/+)); Pdx-1-Cre), we here demonstrate that inhibition of the ARF6-AMAP1 pathway by shRNAs in cancer cells results in therapeutic synergy with an anti-PD-1 antibody in vivo; and furthermore, that silvestrol improves the efficacy of anti-PD-1 therapy, whereas silvestrol on its own promotes tumor growth in vivo. ARF6 and MYC are both essential for normal cell functions. We demonstrate that silvestrol substantially mitigates the overexpression of ARF6 and MYC in KRAS-mutated cells, whereas the suppression is moderate in KRAS-intact cells. We propose that targeting eIF4A, as well as mutant KRAS, provides novel methods to improve the efficacy of anti-PD-1 and associated ICB therapies against PDACs, in which ARF6 and AMAP1 overexpression, as well as KRAS mutations of cancer cells are biomarkers to identify patients with drug-susceptible disease. The same may be applicable to other cancers with KRAS mutations.

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