2-2--(hydroxynitrosohydrazono)bis-ethanamine and Carcinoma--Squamous-Cell

It is not understood why some head and neck squamous cell carcinomas, despite having identical morphology, demonstrate different tumor aggressiveness, including radioresistance. High levels of the free radical nitric oxide (NO) and increased expression of the NO-producing enzyme nitric oxide synthase (NOS) have been implicated in tumor progression. We previously adapted three human tongue cancer cell lines to high NO (HNO) levels by gradually exposing them to increasing concentrations of an NO donor; the HNO cells grew faster than their corresponding untreated ("parent") cells, despite being morphologically identical. Herein we initially characterize the HNO cells and compare the biological properties of the HNO and parent cells. HNO/parent cell line pairs were analyzed for cell cycle distribution, DNA damage, X-ray and ultraviolet radiation response, and expression of key cellular enzymes, including NOS, p53, glutathione S-transferase-pi (GST-pi), apurinic/apyrimidinic endonuclease-1 (APE1), and checkpoint kinases (Chk1, Chk2). While some of these properties were cell line-specific, the HNO cells typically exhibited properties associated with a more aggressive behavior profile than the parent cells (greater S-phase percentage, radioresistance, and elevated expression of GST-pi/APE1/Chk1/Chk2). To correlate these findings with conditions in primary tumors, we examined the NOS, GST-pi, and APE1 expression in human tongue squamous cell carcinomas. A majority of the clinical samples exhibited elevated expression levels of these enzymes. Together, the results herein suggest cancer cells exposed to HNO levels can develop resistance to free radicals by upregulating protective mechanisms, such as GST-pi and APE1. These upregulated defense mechanisms may contribute to their aggressive expression profile.

Topics: Adaptation, Physiological; Blotting, Western; Carcinoma, Squamous Cell; Cell Proliferation; Cell Survival; Checkpoint Kinase 1; Checkpoint Kinase 2; DNA-(Apurinic or Apyrimidinic Site) Lyase; Glutathione S-Transferase pi; Humans; Hydrogen Peroxide; Immunoenzyme Techniques; Nitric Oxide; Nitric Oxide Donors; Nitric Oxide Synthase Type I; Nitroso Compounds; Oxidants; Protein Kinases; Protein Serine-Threonine Kinases; Tongue Neoplasms; Tumor Cells, Cultured; Tumor Suppressor Protein p53; Ultraviolet Rays; X-Rays

The free radical nitric oxide (NO) is over-expressed in many tumors, including head and neck squamous cell carcinomas (HNSCC); however, the role NO plays in tumor pathophysiology is still not well understood. We, herein, report the development of an in vitro model system which can be used to probe the role of NO in the carcinogenesis of HNSCC. Five HNSCC cell lines were adapted to a high NO (HNO) environment by gradually introducing increasing concentrations of DETA-NONOate, a nitrogen-based NO donor, to cell media. The adaptation process was carried out until a sufficiently high enough donor concentration was reached which enabled the HNO cells to survive and grow, but which was lethal to the original, unadapted ("parent") cells. The adapted HNO cells exhibited analogous morphology to the parent cells, but grew better than their corresponding parent cells in normal media, on soft agar, and in the presence of hydrogen peroxide, an oxygen-based free radical donor. These results indicate that the HNO cell lines are unique and possess biologically different properties than the parent cell lines from which they originated. The HNO/parent cell lines developed herein may be used as a model system to better understand the role NO plays in HNSCC carcinogenesis.

Topics: Adaptation, Physiological; Carcinoma, Squamous Cell; Cell Proliferation; Cell Survival; Head and Neck Neoplasms; Humans; Hydrogen Peroxide; Models, Biological; Nitric Oxide; Nitric Oxide Donors; Nitroso Compounds; Oxidants; Tumor Cells, Cultured