American Association of Cancer Research
1 to 5 April, 2000
San Francisco, California


3D BREAST CANCER TISSUE ANALYSIS REVEALS HIGH CELL-TO-CELL HETEROGENEITY IN THE COPY NUMBERS OF SPECIFIC DNA SEQUENCES

S. J. Lockett*, K. Chin#, C. Ortiz de Solorzano*, W. Chou*, A. Jones*, D. Knowles*, J. Gray#.
*Lawrence Berkeley National Laboratory, CA 94720;
#University of California, San Francisco, CA 94143.

Genetic instability, which is believed to lead to phenotypic diversity and the greatly increased chance of cancer cells evolving, was measured in breast cancer. Technology based on fluorescence in situ hybridization in thick (~ 30um) tissue, confocal microscopy and 3D image analysis to accurately enumerate the copy number of specific DNA sequences in the individual nuclei within intact tissue was used.  Histologically normal tissue had two copies of both the centromeric region of chromosome 1 (1c) and the 20q13 locus in each nucleus.  Both an invasive cancer specimen and a ductal carcinoma in situ (DCIS) specimen reported subsets of nuclei where the copy numbers of the two loci were increased, correlated with each other and correlated with nuclear volume.  These results suggest a mechanism of uniform genome amplification leading to polyploid cells in these specimens. These polyploid cells appeared as clumps, which were mixed at a microscopic scale with clumps of cells with two copies of 20q13.  Furthermore, the DCIS specimen had another subset of cells with 1 copy of 1c.  Another invasive cancer specimen was normal at the 20q13 locus , but contained equal numbers of cells with 3 and 2 copies of 1c.  Spatial statistical analysis indicated clumping of the 3 copy cells rather than random mixing of the two components.  For a typical hyperplasia specimen, a third of the nuclei had 1 copy of 1c and these cells appeared to be randomly mixed with the remaining nuclei with 2 copies of 1c.  These results indicate that breast tumors are highly heterogeneous genetically, that genetic changes exist in early pre-cancerous stages and that polyploidization is associated with cancer cells.  Supported by CA58207, CA67412, Vysis Inc. and Zeiss Inc.