Head and neck squamous cell carcinoma (HNSCC) is the sixth commonest cancer worldwide with an increasing incidence in developing countries. Despite numerous advances in surgery, radiotherapy and chemotherapy over the past few decades the overall survival rate for patients with HNSCC has changed little. Currently, the management of HNSCC patients is based on the assessment of a variety of clinical and pathological parameters. However, in many instances, these factors fail to accurately predict the clinical behaviour of an individual patients tumour. HNSCC therefore, is a tumour entity that would benefit from a greater insight into the chromosomal alterations underlying the disease. Knowledge of such alterations would be expected to provide many benefits to the HNSCC clinician in terms of diagnostic and prognostic markers and may eventually identify novel molecular targets for therapeutic intervention. This thesis was aimed at characterising the chromosomal abnormalities involved in the tumourigenesis of HNSCC, principally using the powerful molecular cytogenetic technique of comparative genomic hybridisation (CGH), and the clinical applications of such data. Firstly, the technique was optimised and initially applied to specimens of primary HNSCC and surrounding uninvolved mucosa from 19 patients in order to investigate the phenomenon of 'field cancerization'. Specimens of primary HNSCC and histologically normal mucosa taken from 1cm and 5cm distant to the primary site were analysed from each patient in order to characterise the chromosomal abnormalities associated with malignant tissue and attempt to identify aberrations underlying the `field change'. CGH of the primary tumour specimens revealed numerous chromosomal aberrations with a relatively consistent pattern. Frequent deletions of DNA were identified on chromosome 3p, 4p, 8p, 9p, 11 q, 13q and 18q and frequent gains on chromosomes 2q, 3q, 5p, 7q, 8q, 9q and 11q. The histologically normal mucosa did not show chromosomal abnormalities within the cells analysed. Therefore, if molecular abnormalities were present in the mucosa surrounding a primary HNSCC they would be below the resolution of CGH, such as subtle point mutations, or only present in a minority of cells. In order to investigate the genetic relationship between primary HNSCC and lymph node metastases, matched pairs of primary and metastatic tumours were obtained from 18 patients and analysed by CGH. Whilst the overall frequency of genetic alterations was similar between primary and metastatic tumours, a surprising degree of discordance was identified between each individual's matched pair of tumours. At least one common aberration was identified in all cases studied, however the percentage of aberrations detected in the lymph node metastases that were shared with the primary tumour varied greatly, ranging from 100% - 8.3%. Several chromosomal regions were found to be altered at similar frequencies in both the primary and metastatic tumours. Most interestingly, regions of the genome found to be altered at a higher frequency in the population of metastatic HNSCC included deletion of 4p15.3-pter and 17q22-qter and gain of 6gcen-q15 and 13q21-22. In addition, both gains and deletions of material from chromosome 22 were found at a higher frequency in the metastatic tumours. These chromosomal regions may contain genes important in the process of metastasis in HNSCC. The level of discordance identified between matched pairs of tumours also suggests that a linear progression model may not satisfactorily explain the progression to metastases in all HNSCC. This thesis also addressed the important clinical question of resistance to radiotherapy demonstrated by a significant fraction of laryngeal tumours. No markers that reliable predict the response of an individual tumour to radiotherapy are currently available. The expression of a panel of markers involved in DNA damage recognition, cell cycle arrest, DNA repair and apoptosis were evaluated in 23 glottic laryngeal tumours (8 radio-resistant and 13 radio-sensitive). Of these, the expression of bcl-2, an anti-apoptotic marker, was specifically associated with the resistant phenotype. This statistically significant association provides preliminary evidence for the dysregulation of apoptosis as a mechanism by which resistant tumours can evade radiotherapy induced tumour regression. Overall, CGH analysis of primary HNSCC identified a relatively consistent pattern of DNA alterations with several distinct regions of DNA deletion and gain identified. Frequent deletions of DNA were identified on chromosomes 1p, 2q, 3p, 4p, 4q, 5q, 7q, 8p, 9q, 10q, 11p, 11q, 13q, 17p, 18q, 19 and 21 and frequent gains of DNA on chromosomes 1q, 2q, 3q, 4q, 5p, 6q, 7p, 7q, 8q, llq, 12p, 13q, 18p and 18q. Chromosome 3 was the most frequent site of both deletions and gains. Follow up data was obtained for all patients analysed by CGH and Kaplan-Meier survival analysis demonstrated a significant correlation between gain of DNA on 3q25-27 and reduced overall survival. This finding highlights the necessity for further, high resolution, characterisation of this region in order that the specific genetic marker can be identified. This thesis demonstrates that molecular analysis of tumours is able to offer new, and valuable information for the understanding of HNSCC carcinogenesis and that these data can be used to compliment existing methodology. Further work is required to isolate specific genes and to understand their interactions.
| Identifer | oai:union.ndltd.org:bl.uk/oai:ethos.bl.uk:271986 |
| Date | January 2002 |
| Creators | Ashman, James Nicholas Edmund |
| Contributors | Greenman, John |
| Publisher | University of Hull |
| Source Sets | Ethos UK |
| Detected Language | English |
| Type | Electronic Thesis or Dissertation |
| Source | http://hydra.hull.ac.uk/resources/hull:6912 |
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