Nes such as c-myc, and cKit and mutations in tumor suppressor genes p53 and Rb are also Rosiglitazone characteristically found in HNSCC [4,11-13]. Lastly, it has been reported that Notch1, anLiebertz et al. Head Neck Oncology 2010, 2:5 http://www.headandneckoncology.org/content/2/1/Page 3 ofembryonically-associated receptor for inhibition of differentiation, may play a role in the oncogenesis of multiple types of cancers including leukemias, lung, melanoma, breast, and neurological tumors [14] but its role in HNSCC has not been studied extensively to date. In this report, we now describe the establishment and characterization of a unique HNSCC cell line designated USC-HN1. The cell line was derived from an invasive primary right superior alveolar ridge squamous cell carcinoma in a nonsmoking patient (Stage IVa, T4aN0M0, based on the American Joint Commission on Cancer Staging, 6th Ed.) and has been found to recapitulate the phenotype of the original tumor biopsy. Heterotransplantation and cytogenetic studies demonstrate its oncogenic derivation and monoclonality, respectively. This cell line has been made available for others in the scientific community through the American Tissue-type Cell Collection (ATCC, http://www.atcc.org) and represents an important model for further studies of HNSCC.MethodsCell Lines and Cellsrepeatedly pipetted with a 2 ml glass pipette for further dissociation. After centrifugation, the cells and residual small fragments were resuspended in freezing medium (RPMI-1640 medium with 30 FCS, 1 Penicillin/ Streptomycin, and 10 DMSO), aliquoted into four-six 1.8 ml cryotubes (Nunc, Denmark), and placed in liquid nitrogen for long-term storage. Cells and fragments that adhered to the T-75 flasks were grown in complete medium for 2 weeks before being trypsinized and passaged weekly to new flasks. When the malignant cells were seen to grow, the normal fibroblastic cells were removed by differential trypsinization to enrich the malignant cell population. Finally, malignant cells were cloned in petri dishes using cloning rings to isolate a pure population of HNSCC cells to establish the cell line. The doubling time was determined by cell count measurements at 24 hr intervals for one week from cells in culture after trypsinization. After establishment of the cell line, interval screening for Mycoplasma was performed using the MycoAlert Mycoplasma Detection Kit (Lonza, Rockland, ME).Heterotransplantation in Nude miceHeLa, HUT102, Raji, FaDu, and SW579 cell lines were obtained from the ATCC. All cell lines were maintained in complete medium in humidified incubators at 37 with 5 CO2. IRB PubMed ID:https://www.ncbi.nlm.nih.gov/pubmed/16989806 approval from the USC Keck School of Medicine (HS-09-00048) has been obtained for the collection and use of HNSCC tumor biopsies. Tumor biopsies were surgically resected and placed into 50 mL collection tubes containing approximately 30 mL of RPMI-1640 medium with 20 FCS, 1 Antibiotic-Antimycotic Solution (Mediatech, Inc., Manassas, VA), 10 ug/ml Ciprofloxacin-HCl (Mediatech, Inc.), and 10 ug/ ml Gentamicin Sulfate (Irvine Scientific, Santa Ana, CA). The tubes were immediately put on ice and transported to the laboratory for tissue dissociation.Establishment of Cell Line USC-HNSix-week-old female Nude mice were purchased from Harlan Sprague Dawley (Indianapolis, IN). Institutional Animal Care and Use Committee-approved protocols and institutional guidelines for the proper humane care and use of animals in research were followed. Mice (n = 5) were.