Thein, Angela Thanda Aung; (1997) Molecular genetic analysis of cervical cancer using fluorescence in situ hybridization and comparative genomic hybridization. Doctoral thesis (Ph.D), UCL (University College London).

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Abstract

The aim of this research was to investigate common cytogenetic changes in material from cervical tumours, using modern molecular techniques, in order to throw some light on the role of such changes in the initiation and development of cervical cancer, and to identify the potential locations of genes predisposing to this type of malignancy. Cervical tumours nearly all have complex karyotypes, and more precise cytogenetic information is required to establish whether specific rearrangements occur, and if they are related to the type of HPV infection found. The karyotypes of five recently established cervical cancer cell lines, three from squamous cell carcinomas (two HPV 16 +ve and one HPV 18 +ve), one from an adenocarcinoma (HPV -ve), and one from an adeno-squamous carcinoma (HPV 16 +ve), have been analysed using fluorescence in situ hybridization (FISH), with 23 chromosome specific paints, YACs and cosmids as probes, in addition to conventional G banding. Conventional G banding was used on metaphases from all 5 lines to obtain background karyotype information. A Fluorescence In Situ Hybridization (FISH) technique was applied to metaphases, using 23 different chromosome-specific paint probes to clarify the chromosome rearrangements. These techniques identified the origin of markers and revealed some common chromosome rearrangements. The highest number of breakpoints occurred in chromosome 1. Breaks at 1q10/11 were seen in three different cell lines (DE3, JE6 and XH1) and a break at 1q21 occurred in all three squamous carcinoma lines. Isochromosomes i(1q) were found in two of the lines (DE3 & JE6). Double minutes seen initially in some cells from one squamous line, SM7, were shown also to originate from chromosome 1. Chromosome 3 rearrangements were seen in all five cell lines, involving the short arm in the adenocarcinoma and the adeno-squamous carcinoma cell lines, and the long arm in all three squamous carcinoma cell lines. Yeast Artificial Chromosomes (YACs) and Cosmids, cloned from chromosome 3, were used to map the precise breakpoints on this chromosome and all of them were found to be in different regions. An i(3q) was found in two lines out of five. Small metacentrics involving chromosome 5 were a del(5q) in one line, and a t(X;5) in another, rather than the i(5p) observed by other investigators. The region 6q21 was involved in three cases, and chromosome 9 was rearranged in four. An i(8q) was found in the three squamous carcinoma cell lines. Structural changes of 11q were found in two cases, and a marker 11 representing amplification in the 11q14-22 region was duplicated in the adeno-squamous line. The Comparative Genomic Hybridzation (CGH) technique was applied to three of the cell lines, the squamous line, SM7, the adenocarcinoma line, JE6, and the adeno-squamous line, XH1. All these lines were part of the previous investigation, so it was possible to test the sensitivity of CGH on cell lines where all the major cytogenetic changes had been catalogued in detail. For the near-tetraploid squamous carcinoma cell line, SM7, the hybridization and CGH profiles reflected clearly the numbers of each chromosome present, and the breakpoints of unbalanced aberrations. For the hyper-diploid adenocarcinoma line, JE6, most of the chromosomes present as 2 copies had a profile of 1, but several did show lower profiles which did not represent losses. Most extra chromosomes and regions of change could be clearly seen and, as expected, the reciprocal translocation was not detectable. Over representation of all or part of 3q and 8q in material from four squamous tumours by both techniques shows that these are consistent genetic changes which could provide useful markers for invasive squamous carcinoma of the cervix. (Abstract shortened by ProQuest.)

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