Potential prognostic indicators for the radiotherapy of cervical carcinoma
Biopsies from 117 patients with proven cervical carcinoma were taken under anaesthesia immediately prior to radiotherapy. The Courtenay Mills soft agar clonogenic assay was used to determine the colony forming efficiency (CFE) of tumour cells from disaggregated tumours and the survival of these cells following radiation. Validation of the assay was carried out by demonstrating linearity of colony formation and production of radiation survival curves. The mean CFE was 0.18±0.49% (±lsd) with a range of 0.003 - 4.28% (based on total viable nucleated cell counts) for 84 (72%) specimens. No significant association was demonstrated between in vitro growth and either clinical stage (r=0.02) or tumour differentiation (r=0.08). A wide range of values (0.13-0.97) for surviving fraction at 2Gy (SF2) was obtained with a mean value 0.44 (sd=0.19) for 77 tumours. There were statistically significant differences between the individual tumours (p=<. 001). Heterogeneity in intrinsic radiosensitivity was not demonstrated (p=0.3) when multiple biopsies were processed independently from 18 tumours. From analysis of variance of the SF2 results it appears that the surviving fraction below 0.4 and those above 0.7 which show significant differences in radiosensitivity between pairs of tumours (p=OO5). Differential cell counts were made on cytospin preparations of tumour cell suspensions. There was no correlation between either CFE or SF2 (r = -0.05, r=0.15, respectively) and the degree of lymphocyte (r = 0.12) or macrophage (r = 0.001) infiltration. Ki67 staining of 29 specimens gave a mean proportion of positively stained nuclei of 20.5% (sd=23). The Ki67 index was not correlated with tumour stage (r=0.58), differentiation (r=0.02), CFE (r=0.04) or SF2 (r="0.07). Vascularity was assessed on paraffin sections of 87 tumours. The mean intercapillary distance (ICD) was 240μm (sd=37pm) and mean proportion of vessels was 2.68% (sd=1.64%). Clinical data from 35 patients with 2 year minimum follow-up revealed no significant difference between the mean ICD or mean proportion of vessels for the group of patients which had died or recurred and the group which remained disease free using t-tests (t=0.74, p=0.47, DF=32; t= 0.6, p=0.55, DF=29 respectively).