Breast cancer proliferation measured on cytological samples: a study by flow cytometry of S-phase fractions and BrdU incorporation

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Abstract
Cell kinetics have been shown to be an important predictor of clinical evolution of operated breast cancer. We established a method for the estimation of the proliferative activity of tumour cells obtained by fine needle sampling without aspiration (FNS), using simultaneously S-phase fractions (SPF) measured on DNA histograms and 5-bromodeoxyuridine (BrdU) labelling index (BLI) measured by flow cytometry. Biparametric BrdU/DNA flow cytometry could be performed in 122 of 189 (65%) consecutive patients. The mean BLI of the cytologically malignant FNS (118) was of 3.0 and the median of 2.2%. One hundred and forty-eight DNA histograms (78%) were suitable for SPF analysis, of which 141 presented malignant cells, showing a mean of 4.5 and a median of 3.5%, comparable to BLIs. These results were obtained from fluorescence peak area histograms with doublet discrimination and background subtraction allowing the measurements of SPFs as low as 0.4%. An excellent correlation was thus observed between BLIs and SPFs, for the 94 cases for which both results were available (r = 0.85). Infrequent discordances (9%) were noted with SPFs considerably higher than BLIs. Seven patients had three consecutive FNS of their tumour at weekly intervals before treatment. Some variability in the proportions of multiple subpopulations of tumour cells was observed on the DNA histograms. In contrast, proliferation indices (SPF or BLI) were reproducible, suggesting homogeneous growth rates. We conclude that an estimation of the proliferative activity of breast tumours at any stage of the disease is possible routinely by SPF and/or BLI analysis of FNS. At least one quantitative proliferation index could be obtained for 91% of patients.