Biomedical subjects

Nicole E Benoit

Publications and source records attributed to Nicole E Benoit.

4 recordsLinked to original sources

PubMed2005

Colorimetric approach to high-throughput mutation analysis.

High-throughput genomic mutation screening for primary tumors has characteristically been expensive, labor-intensive, and inadequate to detect low levels of mutation in a background of wild-type signal. We present a new, combined PCR and colorimetric approach that is inexpensive, simple, and can detect the presence of 1% mutation in a background of wild-type. We compared manual dideoxy sequencing of p53 for eight lung cancer samples to a novel assay combining a primer extension step and an enzymatic colorimetric step in a 96-well plate with covalently attached oligonucleotide sequences. For every sample, we were able to detect the presence or absence of the specific mutation with a statistically significant difference between the sample optical density (OD) and the background OD, with a sensitivity and specificity of 100%. This assay is straightforward, accurate, inexpensive, and allows for rapid, high-throughput analysis of samples, making it ideal for genomic mutation or polymorphism screening studies in both clinical and research settings.

Biological Assay↗

PubMed2004

Epstein-Barr virus in head and neck cancer assessed by quantitative polymerase chain reaction.

OBJECTIVES/HYPOTHESIS: Epstein-Barr virus (EBV) has classically been associated with nasopharyngeal carcinoma and Burkitt's lymphoma. Recently, multiple studies have been published linking EBV with oral squamous cell carcinoma and, to a lesser extent, hypopharyngeal and laryngeal tumors. Using a sensitive method of detection, the authors sought to analyze the presence and quantity of EBV DNA in a large cohort of head and neck cancers. STUDY DESIGN: : Retrospective cohort study. METHODS: Three hundred head and neck cancer samples exclusive of nasopharyngeal carcinoma were examined for the presence of EBV using quantitative polymerase chain reaction. Eighty-four tumor samples from the larynx, 30 from the hypopharynx, 73 from the oropharynx, and 113 from the oral cavity were analyzed for EBV quantity, which was expressed as the number of viral copies per cell genome. Representative samples, which contained the highest EBV DNA levels, were examined using in situ hybridization. Results were correlated with tumor grade and site and tobacco and alcohol exposure. RESULTS: Three of 300 (1%) tumor samples were overtly positive for EBV DNA (defined as >0.1 copies of viral DNA/cell genome). Five of 300 (2%) tumor samples showed low levels (defined as >0.01 and <0.1 copies of viral DNA/cell genome), and 68 of 300 tumor samples (23%) showed trace levels (defined as < 0.01 copies of viral DNA/cell genome) of EBV DNA. No correlation was found between EBV positivity and tobacco exposure, alcohol exposure, or tumor grade. CONCLUSION: In the overwhelming majority of head and neck cancers in this North American cohort, EBV did not appear to contribute to growth of a dominant clonal population with integrated EBV genome and was unlikely to be a genetic etiological agent in tumor development. The low quantities of EBV detected in a minority of head and neck cancers may be related to the presence of EBV genome in rare lymphoid or epithelial cells adjacent to the primary head and neck cancer.

Alcohol Drinking↗

PubMed2003

Epigenetic inactivation of RASSF1A in head and neck cancer.

PURPOSE: RASSF1A, a recently identified candidate tumor suppressor gene, was found to be inactivated in lung cancer and other tumor types. We sought to understand the role of RASSF1A in head and neck cancer. EXPERIMENTAL DESIGN: We analyzed the status of RASSF1A and presence of high-risk human papilloma virus (HPV) in head and neck cancer squamous cell carcinoma (HNSCC) cell lines and primary tumors. We used methylation-specific PCR to detect promoter hypermethylation and direct sequence analysis to detect point mutations in primary tumors and cell lines. 5-aza-2-deoxycytidine was used to demethylate the RASSF1A promoter in cell lines. RESULTS: Promoter methylation of RASSF1A was detected in 42.9% (3 of 7) cell lines and 15% (7 of 46) primary tumors but not in the normal control DNA. Direct sequence analysis revealed a point mutation in a cell line and another in a primary HNSCC. After treatment with 5-aza-2-deoxycytidine, re-expression and demethylation of RASSF1A gene were detected in cell lines with promoter hypermethylation. HPV DNA was detected in 34.7% (16 of 46) primary HNSCC. We found a significant inverse correlation between RASSF1A promoter methylation and HPV infection (P = 0.038). CONCLUSIONS: Our results suggest that RASSF1A is inactivated in a subset of HNSCC primary tumors. Moreover, an inverse correlation between RASSF1A and HPV supports a biological mechanism in which both RASSF1A promoter methylation and HPV infection abrogate the same pathway in tumorigenesis.

Azacitidine↗

PubMed2003

A transcriptional progression model for head and neck cancer.

PURPOSE: A genetic progression model for head and neck squamous cell carcinoma (HNSC) has been established and implies the presence of transcriptional dysregulation as a consequence of accumulation of genetic alterations. Although expression array data have been provided for HNSC, the timing of transcriptional dysregulation in the progression from normal mucosa to dyplastic epithelium to invasive HNSC has not been described. Here, we describe a transcriptional progression model of HNSC. EXPERIMENTAL DESIGN: Expression arrays representing >12,000 genes and expressed sequence tags were used to examine malignant lesions (M), premalignant lesions (PM), distant, histopathologically normal mucosa from patients with premalignant or malignant lesions (MN), and normal mucosa from the upper aerodigestive tract of patients with noncancer diagnoses (N). Significance analysis of microarrays, hierarchical clustering, and principal components analysis was used to identify genes with differential expression patterns. RESULTS: Using a false discovery rate of <5% for significance analysis of microarray, the M group revealed 965 up-regulated and 1106 down-regulated genes relative to the N group. The PM group demonstrated 108 up-regulated and 226 down-regulated genes relative to the N group, whereas the M group demonstrated only 5 up-regulated and 13 down-regulated genes relative to the PM group. Both hierarchical cluster analysis and principal components analysis revealed a consistent separation between the N, PM, and M groups, with a closer association between the PM and M groups. To provide independent validation of the microarray data, quantitative reverse transcription-PCR was performed for a significantly up-regulated gene, integrin alpha 6, correlating well with microarray data (linear regression analysis, P < 0.0001). CONCLUSIONS: Similarly to the genetic progression model of HNSC, this transcriptional model shows that the majority of alterations occurs before the development of malignancy and identifies key targets of transcriptional dysregulation during progression from a normal to a premalignant state and from a premalignant to a malignant state.

Carcinoma↗