Role of nitric oxide in pancreatic cancer cells exhibiting the invasive phenotype

Abstract

Pancreatic cancer is a highly metastatic tumor with an extremely low 5-year survival rate. Lack of efficient diagnostics and dearth of effective therapeutics that can target the cancer as well as the microenvironment niche are the reasons for limited success in treatment and management of this disease. Cell invasion through extracellular matrix (ECM) involves the complex regulation of adhesion to and detachment from ECM and its understanding is critical to metastatic potential of pancreatic cancer. To understand the characteristics of these cancer cells and their ability to metastasize, we compared human pancreatic cancer cell line, PANC-1 and its invading phenotype (INV) collected from transwell inserts. The invasive cell type, INV, exhibited higher resistance to Carbon-ion radiation compared to whole cultured (normally dish-cultured) PANC-1 (WCC), and had more efficient in vitro spheroid formation capability. Invasiveness of INV was hampered by nitric oxide synthase (NOS) inhibitors, suggesting that nitric oxide (NO) plays a cardinal role in PANC-1 invasion. In addition, in vitro studies indicated that a MEK-ERK-dependent, JAK independent mechanism through which NOS/NO modulate PANC-1 invasiveness. Suspended INV showed enhanced NO production as well as induction of several pro-metastatic, and stemness-related genes. NOS inhibitor, l -NAME, reduced the expression of these pro-metastatic or stemness-related genes, and dampened spheroid formation ability, suggesting that NO can potentially influence pancreatic cancer aggressiveness. Furthermore, xenograft studies with INV and WCC in NSG mouse model revealed a greater ability of INV compared to WCC, to metastasize to the liver and l -NAME diminished the metastatic lesions in mice injected with INV. Overall, data suggest that NO is a key player associated with resistance to radiation and metastasis of pancreatic cancer; and inhibition of NOS demonstrates therapeutic potential as observed in the animal model by specifically targeting the metastatic cells that harbor stem-like features and are potentially responsible for relapse.

Metabolic characterization of aggressive breast cancer cells exhibiting invasive phenotype: Impact of non-cytotoxic doses of 2-DG on diminishing invasiveness

“…Interestingly, SNAI1, the transcription factor that represses Ecadherin expression (the marker of EMT induction), also enhances gene expression patterns that promote glucose uptake and glycolysis (56). Although we have not determined the expression of E-cadherin in SUM149 INV, it is possible, given the higher invasiveness and higher glycolytic rate, that SUM149 INV was also in the mesenchymal state, similar to our previous report about INV established from human pancreatic cell line, PANC-1, that showed higher ability than WCC, to invade and metastasize in mice, exhibited reduced E-cadherin expression with induction of pro-metastatic genes (29).…”

Metabolic characterization of aggressive breast cancer cells exhibiting invasive phenotype: Impact of non-cytotoxic doses of 2-DG on diminishing invasiveness

“…Interestingly, SNAI1, the transcription factor that represses Ecadherin expression (the marker of EMT induction), also enhances gene expression patterns that promote glucose uptake and glycolysis (57). Although we have not determined the expression of E-cadherin in SUM149 INV, it is possible, given the higher invasiveness and higher glycolytic rate, that SUM149 INV was also in the mesenchymal state, similar to our previous report about INV established from human pancreatic cell line, PANC-1, that showed higher ability than WCC, to invade and metastasize in mice, exhibited reduced E-cadherin expression with induction of pro-metastatic genes (30).…”

Metabolic characterization of aggressive breast cancer cells exhibiting invasive phenotype: impact of non-cytotoxic doses of 2-DG on diminishing invasiveness

“…To prepare the invaded cells of SUM149 (INV), transwell invasion assays were performed as described previously [28,29]. For the re-invasion assay, WCC and INV were both cultured for 1, 4, 7, 12, or 19 days, and the transwell invasion assay was performed repeatedly with these cells as described previously [29].…”

“…Spheroid invasion assay was performed in ultra-low attachment multiple 96 well plates (Sigma-Aldrich) [28,29]. Cells (5 × 10^3) were plated in each well and incubated in 37°C CO 2 incubator.…”