Research article

Searching for molecular markers in head and neck squamous cell carcinomas (HNSCC) by statistical and bioinformatic analysis of larynx-derived SAGE libraries

Nelson JF Silveira^1* , Leonardo Varuzza^2* , Ariane Machado-Lima³ , Marcelo S Lauretto² , Daniel G Pinheiro⁴ , Rodrigo V Rodrigues^5,6 , Patrícia Severino⁷ , Francisco G Nobrega⁸ , Head and Neck Genome Project GENCAPO⁹ , Wilson A Silva Jr⁴ , Carlos A de B Pereira² and Eloiza H Tajara^5,6

¹  Instituto de Pesquisa e Desenvolvimento, Universidade do Vale do Paraíba, UNIVAP, São José dos Campos, SP, Brazil

²  Instituto de Matemática e Estatística, USP, São Paulo, SP, Brazil

³  BIOINFO-USP Núcleo de Pesquisas em Bioinformática, USP, SP, Brazil

⁴  Departamento de Genética, Faculdade de Medicina de Ribeirão Preto-USP, Centro de Terapia Celular, Centro Regional de Hemoterapia, SP, Brazil

⁵  Departamento de Biologia Molecular, Faculdade de Medicina de São José do Rio Preto, FAMERP, São José do Rio Preto, SP, Brazil

⁶  Departamento de Genética e Biologia Evolutiva, Instituto de Biociências, USP, São Paulo, SP, Brazil

⁷  Instituto de Ensino e Pesquisa Albert Einstein, São Paulo, SP, Brazil

⁸  Departamento de Biociências e Diagnóstico Bucal, Faculdade de Odontologia, UNESP, São José dos Campos, SP, Brazil

⁹  Complete authors list and addresses is presented in the Appendix

author email corresponding author email* Contributed equally

BMC Medical Genomics 2008, 1:56doi:10.1186/1755-8794-1-56

Published:	11 November 2008

Abstract

Background

Head and neck squamous cell carcinoma (HNSCC) is one of the most common malignancies in humans. The average 5-year survival rate is one of the lowest among aggressive cancers, showing no significant improvement in recent years. When detected early, HNSCC has a good prognosis, but most patients present metastatic disease at the time of diagnosis, which significantly reduces survival rate. Despite extensive research, no molecular markers are currently available for diagnostic or prognostic purposes.

Methods

Aiming to identify differentially-expressed genes involved in laryngeal squamous cell carcinoma (LSCC) development and progression, we generated individual Serial Analysis of Gene Expression (SAGE) libraries from a metastatic and non-metastatic larynx carcinoma, as well as from a normal larynx mucosa sample. Approximately 54,000 unique tags were sequenced in three libraries.

Results

Statistical data analysis identified a subset of 1,216 differentially expressed tags between tumor and normal libraries, and 894 differentially expressed tags between metastatic and non-metastatic carcinomas. Three genes displaying differential regulation, one down-regulated (KRT31) and two up-regulated (BST2, MFAP2), as well as one with a non-significant differential expression pattern (GNA15) in our SAGE data were selected for real-time polymerase chain reaction (PCR) in a set of HNSCC samples. Consistent with our statistical analysis, quantitative PCR confirmed the upregulation of BST2 and MFAP2 and the downregulation of KRT31 when samples of HNSCC were compared to tumor-free surgical margins. As expected, GNA15 presented a non-significant differential expression pattern when tumor samples were compared to normal tissues.

Conclusion

To the best of our knowledge, this is the first study reporting SAGE data in head and neck squamous cell tumors. Statistical analysis was effective in identifying differentially expressed genes reportedly involved in cancer development. The differential expression of a subset of genes was confirmed in additional larynx carcinoma samples and in carcinomas from a distinct head and neck subsite. This result suggests the existence of potential common biomarkers for prognosis and targeted-therapy development in this heterogeneous type of tumor.