Independent prognostic factors in T2/T3 transitional cell bladder tumours with special reference to histoquantitative methods.

A cohort of 233 T2/T3 transitional cell carcinomas were followed up for over 10 years. Five nuclear factors, two mitotic indices, DNA ploidy and S-phase fraction (SPF) were related to progression and survival of TCCs during that time period. SPF predicted pelvic lymph node involvement at diagnosis (P = 0.064). Progression in T-category was related to T-category (P = 0.035), DNA ploidy (P = 0.0180), papillary status (P = 0.0021), mitotic activity index (MAI) (P = 0.0011), volume corrected mitotic index (M/V index) (P = 0.0017), WHO grade (P = 0.0003) and S-phase fraction (P = 0.0002). Progression in N and M-categories was related to the same variables. Independent predictors of progression in T-category were SPF (P = 0.0161) and WHO grade (P = 0.0236), whereas progression in M-category was independently related to MAI (P = 0.0012) and T-category (P = 0.0004). The SPF (P < 0.0001), M/V index (P < 0.0001), MAI (P < 0.0001), WHO grade (P < 0.0001) and papillary status (P < 0.0001) were the most important predictors of survival in univariate analysis. In a multivariate analysis SPF and M/V index (P < 0.0001) were the best predictors of survival followed by papillary status and T-category. The results show that the proliferation rate of T2/T3 TCCs as determined by flow cytometric SPF or M/V index are equally powerful predictors. They are clearly better than nuclear morphometry, DNA ploidy or WHO grading as prognostic factors.     

Independent clinical, histological and quantitative prognostic factors in transitional-cell bladder tumours, with special reference to mitotic frequency.

A cohort of 537 transitional-cell bladder cancers (TCC) was followed up for a mean of 9 years. Clinical stage, WHO grade, papillary status, 6 nuclear factors and volume-corrected mitotic index (M/V index) were related to progression and survival. Classic and quantitative prognostic factors were significantly interrelated (p less than 0.001). In Ta-Tl tumours M/V index predicted progression independently (p less than 0.001) and in the entire cohort progression was related independently to the M/V index (p = 0.0001) and to the WHO grade (p = 0.0022). In survival analysis, clinical stage (p less than 0.0001), M/V index (p less than 0.0001), WHO grade (p less than 0.0001), papillary status (p less than 0.0001) and nuclear factors (p less than 0.0001) were significant predictors. In papillary tumours, clinical stage (p less than 0.0001), M/V index (p less than 0.0001), WHO grade (p less than 0.0001) and nuclear factors (p = 0.0001-0.0133) were related to survival. In a multivariate analysis T-category (p less than 0.001), WHO grade (p less than 0.001), M/V index (p = 0.002) and papillary status (p = 0.034) predicted survival independently in the entire cohort whereas in papillary tumours T-category (p less than 0.001) and M/V index (p less than 0.001) were independent predictors. If tumours with pelvic lymph-node metastases or distant metastases at diagnosis were excluded from the analysis, T-category (p less than 0.001), M/V index (p less than 0.001) and WHO grade (p less than 0.001) were independent predictors. In papillary tumours T-category (p less than 0.001), M/V index (p less than 0.001) and WHO grade (p = 0.048) predicted survival. The results emphasize the importance of mitotic activity as a most important histological prognostic factor in TCC, second only to clinical stage. In Ta-TI tumours quantitative mitotic frequency analysis includes all the available independent prognostic information. Accordingly, TCC can be graded by mitotic frequency analysis in place of subjective grading systems.     

Over-expression of p53 nuclear oncoprotein in transitional-cell bladder cancer and its prognostic value

Two hundred and twelve archival bladder-cancer biopsy specimens were analyzed immunohistochemically to detect over-expression of p53 protein. The results of immunohistochemical analysis were correlated to established histological and quantitative prognostic factors and survival of patients during a mean follow-up period of more than 10 years. Twentynine percent of tumours were positive for p53 protein, and over-expression was associated with high histological grade, non-papillary growth architecture, dense inflammatory cell reaction, DNA aneuploidy, high S-phase fraction, high mitotic frequency and high SD of nuclear area. Progression in T, N and M categories was significantly related to over-expression of p53 protein. In univariate survival analysis, over-expression of p53 predicted poor outcome in the entire cohort, in papillary tumours and in muscle-invasive tumours but not in superficial tumours. In a multivariate survival analysis, over-expression of p53 oncoprotein had no independent prognostic value over clinical stage and mitotic index. The results confirm that p53 is involved in the growth regulation of bladder cancer and is certainly a subject for detailed analysis of specific mutations.     

Classic prognostic factors, flow cytometric data, nuclear morphometric variables and mitotic indexes as predictors in transitional cell bladder cancer

One hundred and eighty-seven patients with a transitional cell bladder cancer were followed by retrospectively for a mean of 9.5 years. Clinical stage (p less than 0.0001), histological grade (p less than 0.0001), papillary (p less than 0.0001), SPF (p less than 0.0001), M/V index (p less than 0.0001), MAI (p less than 0.0001), DNA index (p = 0.0001) SDNA (p = 0.0004), NA10 (p = 0.0023), NA (p = 0.0044) and G2% (p = 0.0158) were significantly correlated with survival in univariate analysis. In Cox's analysis T-category, papillarity and MAI had independent prognostic value and in combination they predicted bladder cancer-related survival significantly (X2 = 117.5, p less than 0.0001). When histological parameters only were analysed, WHO grade, SPF and papillarity were independent predictors and their combined prognostic significance was high (X2 = 76.6, p less than 0.0001). In papillary tumours SPF (p v 0.0005), MAI (p = 0.0009), DNA index (p = 0.0010) and M/V index (p = 0.0021) predicted survival significantly in addition to stage (P less than 0.0001) and grade (p = 0.0003) in univariate analysis. In Cox's analysis T-category, MAI and M/V index were independent predictors and their combined prognostic value was high (X2 = 54.1, p less than 0.0001). In Ta-T1 and in T2-T3 tumours, WHO grade, papillarity, SPF, mitotic indexes and DI were significant predictors in univariate analysis. In Cox's analysis of Ta-T1 tumours papillarity had independent prognostic value, whereas in papillary Ta-T1 tumours mitotic activity included all prognostic information. In T2-T3 tumours WHO grade and SPF were independent predictors. In conclusion, in papillary bladder tumours T-category is the most important predictor of survival followed by parameters reflecting proliferative activity of cancer cells.     

DNA index, S-phase fraction, histological grade and prognosis in breast cancer

DNA index and S-phase fraction (SPF) were measured by flow cytometry on paraffin embedded tissue from 140 primary breast tumours. The results of DNA analysis were compared with the size, degree of axillary node involvement, histological grade and steroid receptor content of the tumours, as well as with the patients' subsequent clinical course. Forty-four (31.4%) of the 140 tumours were diploid. S-phase fraction was evaluable for 134 (95.7%). The median SPF of the whole population was 7.1%, with diploid tumours having a significantly lower median SPF (3.2%) than aneuploid (10.1%, P less than 0.001). Both aneuploidy (P = 0.002) and high SPF (P less than 0.001) were strongly associated with high histological grade. There was no significant association between either DNA ploidy or SPF and tumour size, nodal status or steroid receptor content. An SPF below the median was strongly associated with better relapse-free survival (P = 0.008), overall survival (P = 0.004) and survival after relapse (P less than 0.001). Ploidy did not correlate significantly with clinical course. Multivariate analysis using the Cox model suggested that, while SPF gave prognostic information independent of tumour size or nodal status, this independent significance was lost when histological grade was included in the analysis.     

Nucleolar organiser regions (AgNORs) as predictors in transitional cell bladder cancer.

The predictive value of silver stained nucleolar organiser regions (AgNORs) was assessed in 229 patients with transitional cell bladder cancer followed up for over 10 years. The AgNORs were enumerated in pretreatment biopsy specimens. The AgNORs were related to clinical stage (T) (P = 0.0111), papillarity (P less than 0.0001), WHO grade (P less than 0.0001), DNA ploidy (P = 0.0010) and S-phase fraction (P less than 0.0001). Tumours presenting with pelvic lymph node involvement (P = 0.0085) or metastasis (P = 0.0780) at the time of diagnosis had more AgNORs than tumours confined to the bladder wall. Progression in T-, N- and M-categories (P = 0.0010-0.0030) was related to AgNORs and consequently they predicted bladder cancer related survival (P = 0.0005). The diploid tumours could be regrouped according to survival by AgNORs (P = 0.0001). In papillary tumours AgNORs predicted progression (P = 0.0110) and survival (P = 0.0038). In Ta-T1 tumours AgNORs predicted progression (P = 0.11) and survival (P = 0.0751) and also in T2-T3 tumours AgNORs contributed to survival significantly (P = 0.0039). The AgNORs subdivided WHO grade III tumours according to their ability to progress during the follow-up time (P = 0.0711). In a multivariate analysis AgNORs predicted progression independently in Ta-T1 category (P = 0.0165). AgNORs predicted recurrence free period like SPF (P = 0.0010). In conclusion, AgNORs are inferior to classic prognostic factors or DNA flow cytometric variables in muscle invasive bladder cancers whereas they have independent predictive value in superficial cancers.     

Ag-nor protein area in transitional cell bladder-cancer as measured by image-analysis

Nucleolar organizer regions were visualised in 50 transitional cell carcinomas by using the silver staining technique. The area occupied by the individual Ag-NOR dots and other structural features related to Ag-NORs were measured by image analyzer (IBAS 2) at a total magnification of x9500. The mean Ag-NOR area ranged between 0.37-0.53 mum2 and the mean Ag-NOR area was 0.47 mum2. The Ag-NOR area or other structural features were not related to clinical stage, WHO grade, papillary status, DNA ploidy, S-phase fraction, volume corrected mitotic index or Ag-NOR count. The Ag-NOR area had no prognostic value in terms of progression or survival. The results indicate that the area of individual Ag-NOR dots is relatively constant in transitional cell bladder cancer and the enumeration of the Ag-NORs has prognostic value. In case the Ag-NORs are packed together, the Ag-NOR area reflects the number of individual Ag-NOR dots and may have prognostic value on that basis.     

Prognostic value of DNA flow cytometry in stomach cancer: a 5-year prospective study

The role of DNA flow cytometry in the prediction of prognosis for patients with stomach cancer remains to be defined. Thus we studied prospectively the role of DNA flow cytometry as a prognosis indicator in stomach cancer patients in a high-incidence area. Between November 1990 and December 1992, primary stomach cancer tissues were obtained from the surgical specimens from 217 patients (148 male, 69 female). DNA flow cytometric analyses of DNA ploidy and S-phase fraction were performed and the results were correlated with patient survival. The median age of the patients was 55 years (range 24-78). Aneuploid cell population was found in 114 of 217 samples (53%). Tumour S-phase fraction was obtained in 96 of 103 diploid tumours (93%) and 61 of 114 aneuploid tumours (54%). After median follow-up of 66.1 months, the patients with tumours with an S-phase fraction over 17% had significantly worse survival rates than patients with tumours with S-phase fractions of lower than 8% or 8-17% (45% vs 59% and 63% of patients surviving, P = 0.007). Tumour ploidy status did not correlate with patient survival. Multivariate analyses showed that the TNM stage remained the most important prognostic indicator. The tumour S-phase fraction was also an independent prognostic indicator (relative risk 2.300, 95% CI, 1.252-4.223). Tumour S-phase fraction obtained by DNA flow cytometry is an independent prognostic indicator for the survival of the patients with stomach cancer.     

Cell proliferation of transitional cell bladder tumours determined by PCNA/cyclin immunostaining and its prognostic value.

Cell proliferation of transitional cell bladder cancer (TCC) was determined by PCNA (proliferating cell nuclear antigen)/cyclin immunostaining in 178 TCCs and the results were related to established prognostic factors, progression and survival during a mean follow-up period of 10 years. The fraction of PCNA/cyclin positive nuclei was related to T-category (P = 0.008), papillary status, WHO grade, DNA ploidy, S phase fraction, M/V index (volume corrected mitotic index) and AgNORs (silver stained nucleolar organiser regions) (for all P less than 0.001). TCCs presenting with pelvic lymph node metastasis at diagnosis had a significantly higher growth fraction than the tumours confined to the bladder wall (P less than 0.001). The fraction of PCNA/cyclin positive nuclei predicted progression in T-, N- and M-categories (P less than 0.001). In Ta-T1 tumours high fraction of PCNA/cyclin positive nuclei predicted metastasis (P = 0.019). In survival analysis the fraction of PCNA/cyclin positive nuclei predicted survival in the entire cohort (P less than 0.001) and in Ta-T1 tumours (P = 0.0005). In a multivariate survival analysis the fraction of PCNA/cyclin positive nuclei showed independent predictive value in the entire cohort (P = 0.046), in papillary tumours (P = 0.006) and in Ta-T1 tumours (P = 0.015). The results show that the growth fraction as determined by PCNA/cyclin immunostaining is a significant prognostic variable in TCC.     

DNA ploidy, S-phase fraction and mitotic indices as prognostic predictors of female breast cancer.

DNA ploidy, S-phase fraction (SPF), mitotic index (MI), volume corrected mitotic index (M/V index) and standard prognostic factors were related to disease outcome in a series of 363 women with breast cancer followed-up for over 10 years in our clinic. DNA ploidy and SPF were significantly related to histological type, tumour grade and mitotic indices (p < 0.001). In univariate survival analysis, pN status (p < 0.0001), tumour diameter (p < 0.0001), MI (p = 0.001), M/V index (p = 0.0003) and SPF (p = 0.015) predicted survival. In pN(-) tumours. MI (p = 0.059) was related to survival. In pN(+) tumours, tumour diameter (p = 0.0004), M/V index (p = 0.023) and SPF (p = 0.045) predicted survival. In multivariate survival analysis, tumour diameter (p < 0.001). M/V index (p < 0.007), pN status (p = 0.014) and patient age (p = 0.09) were independently related to survival. In pN(-) tumours, tumour diameter independently predicted survival (p = 0.033). In pN(+) tumours, tumour diameter (p < 0.001), M/V index (p = 0.006) and the year of treatment (p = 0.08) were independent predictors. The results show that tumour diameter, pN status and proliferative activity of cancer cells are important prognostic factors in breast cancer. Of the proliferation indices, M/V index and SPF are equally powerful predictors, and the use of M/V index is advocated due to simplicity of the assessment.     

Cytometric and histopathologic features of tumors detected in a randomized mammography screening program: Correlation and relative prognostic influence

Summary Cytometric determination of S-phase fraction and ploidy type was performed on 430 tumors detected within a randomized trial of mammographic screening. The results were compared to several histopathologic features. A high S-phase fraction was estimated in tumors with a high grade of malignancy and other histopathologic findings related to rapid tumor progression, including lack of tubule formation, a high mitotic index, marked nuclear pleomorphism, multifocal cancer growth, tumor emboli in lymphatic and blood vessels, tumor necrosis, and inflammatory reaction. DNA aneuploidy was correlated with a high malignancy grade, frequent mitoses, a high degree of nuclear pleomorphism, vascular invasion, necrosis, and the presence of noninvasive ductal carcinoma. Both cytometric variables were inversely related to the degree of elastosis. Positive nodes, large tumor size, DNA aneuploidy, a high S-phase fraction, high grade of malignancy, lack of tubule formation, as well as high mitotic index and pleomorphism, presence of multifocal cancer, and vascular invasion, predicted a significantly shorter distant recurrence-free interval after a median follow-up time of 46.6 months. Elastosis and the presence of estrogen and progesterone receptors indicated favorable prognosis. In the multivariate analysis, only lymph node status, tumor size S-phase fraction, and multifocal growth pattern had independent prognostic value.     

Ploidy aberrations and agar cloning ability of malignant cells from human bladder carcinoma

Nine patients with urinary bladder tumours were examined by cystoscopy. Cold cup biopsies were obtained from the tumours and histological sections made. Mapping of the bladder mucosa was performed by selectively aspirating cells from the tumours and from different areas of the bladder. Cellular DNA content was measured by flow cytometry. Additionally, tumour cells were grown in soft agar. Plating efficiency seemed to relate neither to histological grade in the original tumour, to ploidy state, nor to the proliferative fraction of cells in the tumours. Flow cytometry and morphology combined seemed to be more reliable indicators for biological properties of the tumours.     

DNA ploidy and S-phase fraction as prognostic factors in patients with uveal melanomas.

In 96 patients with uveal malignant melanomas the tumours were investigated by DNA flow cytometry. Thirty-eight per cent of the melanomas were aneuploid. By univariate analysis significant correlations with survival were found for histological type, tumour size, DNA ploidy, evidence of ''blind eye'' and S-phase fraction. By multivariate analysis, significant prognostic variables were found to be histological type (P = 0.0008), tumour size (P < 0.0001) and DNA ploidy (P = 0.0038). Evidence of ''blind eye'' was not significantly correlated with survival after adjustments for the other variables mentioned above. The S-phase fraction could be estimated in all 60 diploid tumours and in 12 of 36 aneuploid melanomas. By univariate analysis this variable was found to be a significant prognostic factor, but did not remain so after adjustment for ploidy, histological type and tumor size. We further conclude that patients with small DNA diploid uveal melanomas of spindle cell type have a rather favourable prognosis.     

Flow cytometry and interactive image cytometry in endometrial carcinoma. A comparative and prognostic study

Comparative DNA measurements were performed in 139 women with endometrial carcinoma using flow cytometry (FCM) and interactive image cytometry (ICM). Ploidy level and the percentage of S-phase cells were determined by FCM and ploidy level and the percentage of cells with a DNA content exceeding 2.5c, 3c, 4c and 5c, respectively, were calculated by ICM. The aim was to compare ploidy level obtained by the two methods and to evaluate the prognostic value of all the above-mentioned parameters. Recurrence or residual disease after completing treatment were used as end-points. Follow-up time was 18-48 months. An agreement was obtained in 85% of the cases as regards ploidy level, but in 15% the tumors were regarded as near-diploid by one method and as grossly aneuploid by the other. Both ploidy level (both methods) and S-phase rate (FCM) were correlated with histopathologic grade (p less than 0.001 and p less than 0.05, respectively). In univariate analysis, ploidy level (obtained either by FCM or ICM) correlated with recurrence rate, with a more favourable prognosis for near-diploid cases. When using multivariate models (Cox analysis) including clinical variables, ploidy level by FCM (but not ICM) was still significant as regards prognosis. In the multivariate analysis, S-phase fraction (as measured by FCM) also yielded independent prognostic information. In a separate analysis the proportion of cells with a DNA content greater than 5c gave independent prognostic information besides that of the S-phase fraction and ploidy level. We conclude that measurements of the percentage of cells exceeding 5c give prognostic information beyond the information obtained from flow cytometric determination of ploidy level and S-phase fraction.     

Heterogeneity of colorectal adenocarcinomas evaluated by flow cytometry and histopathology

Flow cytometry and histopathology were utilised in evaluating 50 primary and 16 metastatic colorectal carcinomas to determine the influence of heterogeneity and proportion of dying cells on pathological assessments. A new procedure was developed for staining unfixed whole cells with acridine orange and ethidium bromide to quantify DNA and RNA content and number of dead and dying cells. Attempts were made to reduce interobserver variation in histological assessment and to determine whether flow cytometry could refine current grading and staging procedures. Interobserver variation in grading was not improved by estimating proportions of differing grades in multiple samples from individual tumours. Considerable heterogeneity was observed within tumours although this was less apparent when defining ploidy status than histological grade. No consistent differences were observed between superficial and deep parts of tumours or between primary and secondary tumours by either method of analysis. The proportion of dead and dying cells varied widely between tumours but there was no correlation with tumour grade or stage. Non-diploid tumours were not of more advanced stage or poorer histological grade than diploid tumours. Since ploidy status may be an important prognostic factor, analysis of colorectal carcinomas by flow cytometry could be of greater value than conventional grading and staging procedures.     

DNA ploidy and S-phase in primary malignant melanoma as prognostic factors for stage III disease.

In 82 patients with stage III malignant melanoma, the primary tumours were investigated by DNA flow cytometry. The tumours were classified as DNA diploid (n = 36), tetraploid (n = 11) and aneuploid (n = 35). By univariate analysis a significant correlation with post-recurrence survival was found for time to first metastasis, DNA-ploidy and S-phase fraction. By multivariate analysis, significant prognostic variables were found to be the time to first metastasis (P = 0.006), and ploidy (P = 0.011). Patients with diploid melanomas and a long recurrence-free interval had a median post-recurrence survival time of 45 months compared to 18 months in patients with DNA aneuploid tumours and an early recurrence. The S-phase could be estimated in 47 primary melanomas and was found to be a significant prognostic variable (P = 0.017). The median survival was 45 months for patients with melanomas with a S-phase fraction below 5%, and 19 months for melanomas with S-phase above 10%. The prognostic value of the S-phase remained significant even after adjustment for recurrence-free interval and DNA ploidy.     

The prognostic significance of DNA flow cytometry in breast cancer: results from 881 patients treated in a single centre.

In this single-centre study of 881 patients, S-phase fraction (SPF) was shown to be a significant prognostic marker in terms of overall survival (OS), relapse-free survival (RFS) and survival after relapse (SAR). Further, SPF had independent prognostic significance when considering a range of other clinicopathological variables, namely tumour grade and stage, nodal status, patient age, tumour size, menstrual status and treatment details. For OS and RFS, SPF was the second strongest predictor of the clinical course of the disease after nodal status, and for SAR it was the strongest prognostic marker. SPF correlated positively with histological grade but was the stronger predictor of survival. The distribution of SPF values was markedly different for the two ploidy classes of tumour, with DNA aneuploid tumours having a significantly higher average SPF. However, SPF retained its independent prognostic ability when DNA diploid and aneuploid tumours were analysed separately, DNA ploidy itself also proved to be an independent prognostic marker but the survival difference between the two ploidy classes was much less than that seen for different levels of SPF. Tumours with several DNA aneuploid populations (multiploid tumours) tended to have a worse prognosis than other aneuploid tumours but this trend did not reach statistical significance. In this and other studies from this centre, SPF has proved to be a robust predictor of clinical outcome in carcinoma of the breast.     

Predicting outcome for patients with node negative breast cancer: a comparative study of the value of flow cytometry and cell image analysis for determination of DNA ploidy.

This study was aimed at determining whether tumour DNA content measured by cell image analysis could provide additional prognostic information when compared to that provided by flow cytometry. Sections cut from paraffin blocks of tumours from 101 patients with node negative breast cancer were analysed by both methods and the results related to other prognostic variables and to patient relapse and overall survival. DNA ploidy measured by flow cytometry classified 46 tumours as diploid and 55 as aneuploid, whereas by cell image analysis 30 were diploid and 71 aneuploid (P less than 0.002). There were 20 tumours with discrepancies between the two methods; 18 of these were tumours with only one peak in flow analysis, but determined to be aneuploid with image analysis. DNA content as measured by both methods was significant for predicting relapse and survival by log-rank test, as were tumour histological grade, c-erbB-2 expression and tumour size. Multivariate analysis showed DNA ploidy measured by flow cytometry to be the only variable of independent significance (P less than 0.02) for both relapse and overall survival. Compared with cell image analysis, flow cytometry demonstrated a significantly higher proportion of diploid tumours, which may be related to differences in the internal standards applied to each method. We suggest that cell image analysis techniques can provide more sensitive information on the DNA content of tumour cells by direct measurement of nuclear DNA density of both normal lymphocytes and tumour cells in the same section. However, although image analysis appears to be more sensitive than flow cytometry in detecting DNA aneuploidy, the image technique appears to lack the specificity of flow cytometry in correlation with clinical outcome.     

Correlation of DNA ploidy and S-phase fraction with chemotherapeutic response and survival in a randomized study of disseminated malignant melanoma

DNA ploidy and S-phase fraction were measured by flow cytometry in the tumour tissue of 87 patients with disseminated malignant melanoma, who had been classified either as responders or with progressive disease in a study of the effects of 2 chemotherapeutic regimens. The patients had been randomized to receive treatment with dacarbazine (DTIC) and vindesine (Eldesine) with or without addition of cisplatin (Platinol). Tumour tissue was obtained from both the primary tumours and the last histologically verified metastases, but in some cases only the primary tumours or the last metastases could be evaluated. There was a significantly higher mean S-phase value in melanoma metastases from patients with complete or partial responses compared with patients with progressive disease. Neither the S-phase fraction of the primary tumour, nor the DNA ploidy of the primary tumour or of the last histologically verified metastases taken before inclusion into the study were associated with therapeutic response. In the multivariate analysis, both the anatomical location of the metastases and the S-phase fraction measured on the last metastases remained significant prognostic factors of response. In the univariate survival analysis, there was an association between high S-phase fractions of the metastases and longer survival. In the multivariate survival analysis, the S-phase fraction, the number of involved metastatic sites and the treatment response were independent predictive factors. We conclude that, in disseminated melanoma treated with chemotherapy, a high S-phase fraction measured in the last histologically verified metastases is associated with a higher response rate and a longer survival. Our results clearly support the role of S-phase measurement as a potential tool for selecting patients for treatment. © 1996 Wiley-Liss, Inc.