Value of nuclear DNA ploidy patterns in patients with prostate cancer after radical prostatectomy.

Flow cytometric analysis of DNA ploidy was performed on prostatic adenocarcinoma specimens from 80 patients. In all these patients a radical retropubic prostatectomy had been performed. The nuclei for DNA ploidy determination were extracted from paraffin-embedded material of whole sections of the prostate from patients treated by radical prostatectomy between 1980 and 1985. DNA ploidy was a strong prognostic indicator independent of tumor grade and tumor stage. DNA ploidy offered additional information on both tumor stage and tumor grade. In stage C disease the likelihood of progression-free survival was 89.5% in diploid tumors and 27.8% in aneuploid tumors after 9 years. In tetraploid tumors all patients progressed after 9 years. The computed survival rates in stage C disease showed that patients with diploid tumors did significantly better than those with aneuploid or tetraploid tumor patterns. These data indicate therefore that DNA ploidy patterns determined by flow cytometric analysis provide important additional prognostic information on prostatic adenocarcinoma treated by radical prostatectomy.     

Prognostic significance of flow cytometric DNA analysis in colorectal cancer

Significance of flow cytometric DNA analysis for assessing malignant potential and survival of colorectal cancer was investigated using paraffin-embedded materials from 144 patients with primary colorectal cancer who had been treated from 1971 to 1985. Forty-four percent of colorectal cancer were composed of diploid and 56 percent were aneuploid. DNA indices (DI) of aneuploid tumors showed a bimodal distribution. There was no significant correlation between ploidy pattern and clinicopathological factors. While, DI level showed significantly higher in poorly differentiated adenocarcinomas and in clinicopathological stage III and V tumors. Overall survival in the patients with aneuploid tumor was significantly worse than that in those with diploid tumor (p less than 0.001). Survival rate was poorer in the patients with aneuploid tumor than in those with diploid tumor, who were stratified according to categories of curable resection, stage, histological type, negative peritoneal or hepatic involvement and negative node metastases. However, there was no significant relation between DI and survival among the patients with aneuploid tumor. From these results, it was concluded that the nuclear DNA content of colorectal cancer may represent biological malignant potential of the disease, and that the DNA ploidy pattern may be an important prognostic indicator, being independent of clinicopathological factors.     

Flow cytometric DNA analysis of stage D2 prostatic carcinoma.

Flow cytometric DNA analysis was performed on 34 samples of prostatic carcinoma patients with stage D2 disease to study the relationship between DNA ploidy pattern, Gleason sum score, and prognosis. Nuclei were extracted from paraffin-embedded needle biopsy specimens. The DNA ploidy pattern was diploid in 53% of the tumors, tetraploid in 38%, and aneuploid in 9%. A significant correlation between DNA ploidy pattern, Gleason sum score, and prognosis was not proved. It was considered that DNA ploidy pattern and Gleason sum score of limited primary lesion did not allow a prognosis of the patients with stage D2 prostatic carcinoma to be made.     

Flow cytometric DNA analysis in colorectal cancer

Flow cytometric DNA analysis for assessing malignant potential of colorectal carcinoma was investigated by paraffin-embedded materials. Preservation time of paraffin blocks and formalin fixation time of surgical specimens within 14 days do not influence the nuclear DNA content. There was seen a good correlation between the DNA contents of paraffin-embedded and fresh materials obtained from the same surgical specimens. Using deparaffinized tumor specimens, the nuclear DNA content was measured by flow cytometry in 144 patients with primary colorectal cancer, who had been treated from 1971 to 1985. Forty-four percent of colorectal cancer were diploid and 56% were aneuploid. There was no significant correlation between ploidy pattern and clinicopathological factors. However, the patients with aneuploid tumor had a significantly worse survival than those with diploid tumor (Generalized Wilcoxon test, p less than 0.001). The patients with aneuploid seemed to have an unfavorable survival than those with diploid in the same stage, and had a significantly worse survival in each group of negative nodes, P0 and H0. It is concluded, therefore, that the nuclear DNA content of colorectal cancer may be an important prognostic factor, being independent of pathological stage.     

Prognostic significance of nuclear deoxyribonucleic acid ploidy patterns in resected hepatic metastases from colorectal carcinoma

Nuclear deoxyribonucleic acid (DNA) ploidy studies of paraffin-embedded archival tumor specimen blocks were performed by flow cytometry on extracted nuclei from 101 surgically resected hepatic metastases from colorectal cancer. In 28 patients, the corresponding primary carcinoma of the metastases was also studied. Tumor clinicopathology and clinical course of the patients were reviewed. Preparation of paraffin-embedded tissue specimens was performed by the technique of Hedley et al. and stained with propidium iodide according to the method of Vindelov et al. Eighty-eight of 101 metastatic tumors and 26 of 28 primary tumors yielded evaluable DNA histograms. Twenty-six metastases showed a DNA diploid pattern, 25 showed a significantly increased 4C peak (DNA tetraploid/polyploid), and 37 had a DNA aneuploid peak. Ploidy pattern was constant between primary and metastases in 84.6% of tumors. No significant relationship between host and tumor characteristics and ploidy pattern was found except for a correlation between grade 3 metastases and DNA aneuploid. Survival of patients with DNA aneuploid metastases was significantly less than that of patients with DNA diploid metastases (p = 0.03). However, among DNA nondiploid metastases, survival was significantly less for low DNA index metastases (less than or equal to 1.5) than for high DNA index (greater than 1.5) metastases (p less than 0.05). Flow cytometric DNA ploidy measurements may have prognostic value for patients with resected hepatic metastases from colorectal carcinoma.     

Nuclear DNA content in breast carcinoma with special reference to the parameters of malignant potentiality

Nuclear DNA ploidy pattern was examined in 22 breast carcinomas with a tumor mass of up to 2.2 cm and compared with the clinical and histological parameters of the malignant potentiality of breast carcinomas. The 22 carcinomas were divided into 11 aneuploid and 11 near-diploid carcinomas. It was found that the histological type, histological grade, nuclear grade, mitotic index and lymphatic invasion were either only slightly, or not correlated at all with the DNA ploidy pattern. Nevertheless, lymph node metastasis and negative estrogen receptor status was more frequently seen in the patients with aneuploid carcinoma than in those with near-diploid carcinoma.     

Nuclear DNA content in breast carcinoma with special reference to the parameters of malignant potentiality

Nuclear DNA ploidy pattern was examined in 22 breast carcinomas with a tumor mass of up to 2.2 cm and compared with the clinical and histological parameters of the malignant potentiality of breast carcinomas. The 22 carcinomas were divided into 11 aneuploid and 11 near-diploid carcinomas. It was found that the histological type, histological grade, nuclear grade, mitotic index and lymphatic invasion were either only slightly, or not correlated at all with the DNA ploidy pattern. Nevertheless, lymph node metastasis and negative estrogen receptor status was more frequently seen in the patients with aneuploid carcinoma than in those with near-diploid carcinoma.     

Prognostic significance of cellular DNA ploidy level of the breast cancer]

This paper reports that cellular DNA ploidy level of paraffin-embedded histological material from 47 patients with stage II breast cancer was measured by flow cytometry. Meanwhile, the influence of cellular DNA level on disease-free survival (DFS) of postoperative patients for the breast cancer was analyzed. The results showed eighteen (38.3%) of tumors examined were DNA diploid, and the remainder were DNA aneuploid. Patients with diploid cancer had a significantly better disease-free survival compared with the patients with aneuploid cancer (P less than 0.02). Our results suggested that cellular DNA ploidy level is a good prognostic factor and of important significance in predicting prognosis of patients with the breast cancer.     

Squamous cell carcinoma of the renal pelvis: nuclear deoxyribonucleic acid ploidy studied by flow cytometry

From 1944 to 1987, 28 patients with squamous cell carcinoma of the upper urinary tract were treated and also had tumor specimens that were fully evaluable by flow cytometric nuclear deoxyribonucleic acid ploidy analysis: 22 had squamous cell carcinoma of the intrarenal collecting system, 4 had tumors of the ureter, and 2 had tumors of the renal pelvis and ureter. Eight patients (29%) had deoxyribonucleic acid diploid, 11 (39%) tetraploid and 9 (32%) aneuploid ploidy patterns. Ploidy pattern significantly correlated with histological grade and tumor stage. Almost all tumors were histologically of high grade; among the patients with high grade tumors ploidy analysis separated fair and poor prognosis groups. Pathological stage was the dominant clinical variable. A total of 14 patients (50%) had advanced stage disease and all died within 12 months of diagnosis. Nearly all of these patients showed abnormal ploidy patterns and ploidy analysis was not useful prognostically for this group. In contrast, all 3 patients with squamous cell carcinoma of the renal pelvis who were long-term survivors had deoxyribonucleic acid diploid tumors. However, there is no clear statistical evidence from this study that ploidy analysis provides important prognostic information independent of stage and grade for patients with squamous cell carcinoma of the renal pelvis.     

Flow cytometric analysis of DNA ploidy, cell cycle and cytomorphometry in sarcomatoid renal cell carcinoma.

In 5 patients with sarcomatoid renal cell carcinoma, the nuclear DNA content and size were determined by flow cytometry (FCM), and the prognostic value of DNA ploidy, the percentage of S-phase cells (SPF), and the ratio of modal nuclear size with clinicopathologic behavior was analyzed. Age and clinical stage have been shown to have a strong correlation with prognosis. Older patients with a high stage of cancer had poor outcome with a shorter survival time. In all 60% of the tumors were aneuploid. Tumor invasiveness was related to DNA ploidy. With increasing stage, the overall incidence of aneuploid rises. One alive patient had diploid DNA while 75% of the patients who died of sarcomatoid renal cell carcinomas had aneuploid DNA. Diploid sarcomatoid renal cell carcinomas show significantly lower SPF than aneuploid tumors. There was no significant association between the modal nuclear size and the invasiveness of tumors or survival time. This study suggests that FCM analysis of tumor DNA content and cell cycle could be regarded as an additional prognostic determinant of sarcomatoid renal cell carcinoma.     

Transitional cell carcinoma of the renal pelvis and ureter: prognostic relevance of nuclear deoxyribonucleic acid ploidy studied by slide cytometry: an 8-year survival time study.

In 72 patients with urothelial carcinoma of the renal pelvis or ureter the ploidy, deoxyribonucleic acid (DNA) heterogeneity and counts of cell cycle phases in the tumor were analyzed by means of single cell DNA cytophotometry with the intention of finding new prognostic factors in addition to those already known (stage and grade). Followup ranged from 1 to 8 years. The results of the DNA analyses were related to the tumor categories, histopathological grading of the tumors and clinical course. Malignancy grade 1 tumors showed DNA frequency peaks in the diploid range, while tumors assessed as malignancy grade 2 showed heterogeneous DNA distribution patterns. Malignancy grade 3 tumors exhibited 71% aneuploid and 29% tetraploid DNA values. The proliferation rate of the tumor cells was statistically significantly higher in malignancy grades 2 and 3 than in malignancy grade 1. The prognosis for grade 1 tumors is good, whereas it is unfavorable in the case of grade 3 tumors. For these 2 groups (patients with grades 1 and 3 tumors) DNA ploidy affords no additional prognostic information. Grade 2 tumors, on the other hand, are heterogeneous in respect to DNA ploidy although they exhibit the same histomorphological degree of differentiation. These tumors can be subclassified as aneuploid (biologically aggressive) and diploid or tetraploid (biologically less aggressive) tumors. There was also a positive correlation between T category and DNA ploidy. The cell lines were aneuploid in 38% of the patients with stage T1 tumors, 56% with stage T2 tumors and almost 85% with stage T3, N+ tumors. A significant correlation was found between the results of DNA cytophotometry and the clinical course of the disease. Patients with diploid tumor cell nuclei had no metastases and no local tumor progression for up to 8 years, whereas patients with aneuploid tumor cell nuclei suffered metastasis and local tumor progression within 24 to 36 months. The patients died of the tumor 36 months after primary diagnosis on the average. The determination of DNA ploidy, tumor heterogeneity and tumor cell proliferation by means of DNA cytophotometry affords valuable clues as to prognosis.     

Prostate cancer in men less than 45 years old: influence of stage, grade and therapy

We analyzed patients with histologically proved adenocarcinoma of the prostate to determine the natural history of prostatic cancer in men less than 45 years old. The mean age of the patients was 41 years (range 36 to 44 years). At presentation 5 patients were asymptomatic, 5 had voiding symptoms, 3 had bone pain, 3 had hematuria and 1 had testicular pain. Followup in these patients ranged from 19 to 270 months, with a mean of 111 months. Six patients with clinical stage B disease at diagnosis underwent radical retropubic prostatectomy. These patients enjoy projected 10 and 15-year survival rates of 100 and 82 per cent, respectively. Four patients with stage C disease died of prostatic cancer, although 1 survived for 204 months. Of 4 patients with stage D disease 3 died within 13 months, while 1 still is alive at 48 months. No patient with a Gleason tumor score of 8 to 10 survived more than 13 months. Patient age at presentation appears to be less important than clinical stage, histological grade or treatment modality in the prediction of the course of prostatic cancer. Young men with localized disease at presentation should be treated aggressively and they should have survival rates comparable to actuarial expectancy.     

Flow cytometric assessment of deoxyribonucleic acid content in renal adenocarcinoma: does ploidy status enhance prognostic stratification over stage alone?

Flow cytometry was used to analyze deparaffinized primary renal cell carcinoma specimens from 106 patients to evaluate deoxyribonucleic acid ploidy as a predictor of disease progression and survival. Of these specimens 62 (58%) demonstrated aneuploid stem lines: 30 (48%) of these were tetraploid aneuploid while 32 were nontetraploid aneuploid. Two or more specimens were analyzed from a single primary tumor in 17 patients and heterogeneity of ploidy status was observed in 5 (30%). Specimens of the primary tumor, and regional and/or distant metastases from 11 patients were analyzed; 5 (45%) demonstrated discordance between the ploidy of the primary and the metastatic site. A significant correlation was noted between the presence of aneuploid stem lines and high stage disease (p equals 0.004) but there was no significant correlation between ploidy status and tumor grade. Although there was a significant difference (p equals 0.037) in the incidence of disease progression in patients with diploid tumors (13%) versus those with aneuploid tumors (35%) in the total population, and Kaplan-Meier disease-specific survival curves demonstrated a survival advantage for patients with diploid tumors in the total population, no clear survival advantage was demonstrated for evaluable patients with diploid tumors when controlled for tumor, nodes and metastases stage. In conclusion, the heterogeneity of ploidy status in primary renal cell carcinoma, the high incidence of disease progression in patients with diploid primary tumors and the lack of a clearly demonstrable stage-independent impact of ploidy on prognosis currently would not support widespread clinical application of ploidy status of the primary tumor in the management of individual patients with renal cell carcinoma.     

The prognostic value of deoxyribonucleic acid flow cytometric analysis in stage D2 prostatic carcinoma

This study was designed to compare the prognostic potential of tumor grade and ploidy status in patients with stage D2 prostate cancer. Two outcome groups were selected on the basis of survival after orchiectomy: a bad outcome group consisting of 66 patients who died of the disease within 12 months and a good outcome group comprising 37 patients who survived beyond 5 years. Tumors were classified histologically as well (17%), moderately (17%) or poorly (66%) differentiated. Tumor grade was a significant predictor of outcome, with 76% of poorly differentiated tumors in the bad outcome group and 65% of well differentiated tumors in the good outcome group (p less than 0.005). Deoxyribonucleic acid (DNA) ploidy analysis was performed on formalin fixed, paraffin embedded samples of the primary tumor to yield 97 final tracings that were classified using set criteria for DNA ploidy status. Over-all, 54% of the tumors were nondiploid (33% aneuploid and 21% tetraploid) and the remaining 46% were diploid. DNA ploidy status was a significant indicator of outcome (p less than 0.001), with 64% of diploid tumors in the good outcome group and 88% of the nondiploid tumors in the poor outcome group. Tetraploid tumors behaved no differently from other nondiploid tumors. We conclude that DNA ploidy status and tumor grading are significant independent predictors of outcome after orchiectomy and when combined yield important additional prognostic information.     

Nuclear DNA analysis: the relevance of ploidy, DNA heterogeneity and phases of the cell cycle in 329 patients with prostatic carcinoma. A study on a follow-up of eight years

The ploidy, DNA heterogeneity and the phases of the cell cycle of the tumor were analyzed, by means of single-cell DNA cytophotometry, in 329 patients with locally advanced prostatic carcinoma to find out and establish prognostic factors apart from those already known (stage, grade). Follow-up periods ranged from 1 to 9 years. 253 (76.8%) of the 329 patients had carcinoma stage T3 Nx M0, and 76 of them (23.1%) had carcinoma stage T3/T4 N2-4 M1. 11.8% of the patients showed a cytological grade of malignancy I, while 64.3% had grade II carcinoma and 23.8% had grade III carcinoma. Single-cell DNA cytophotometry demonstrated aneuploidy rates of up to 71% and diploidy rates of up to 23.8% for the higher grades of malignancy, i.e. grades II and III, whereas the diploidy rate established for grade I was 68% and the respective aneuploidy rate was 21%. These differences are significant (p less than 0.001). There was a significant correlation between the results of DNA cytophotometry and the clinical course of the disease. Only 3 (3.7%) of the patients with diploid tumor cell nuclei developed metastases and local tumor progression within 8 years, whereas patients with aneuploid tumor cell nuclei showed metastases and local tumor progression within 8-22 months. These patients died of carcinoma after an average 18 months following primary diagnosis.     

Flow cytometric analysis in colorectal cancer with hepatic metastases and its relationship to metastatic characteristics and prognosis

Nuclear DNA ploidy studies were performed by flow cytometry on extracted nuclei from 65 heptic metastases from colorectal cancer. In 25 patients, both primary and metastatic lesions were available for analysis. Primary carcinomas were DNA diploid pattern in 48.1%, DNA aneuploid in 51.9%. Of 31 hepatic metastases, 11 (35.5%) metastases showed a DNA diploid pattern, and 25 (64.5%) showed a DNA aneuploid pattern. Ploidy pattern was constant between primary and metastases in 80% of tumors. No significant relationship between metastatic characteristics and DNA ploidy pattern was found. The DNA aneuploid cancers had a relatively poorer prognosis in patients with unresectable hepatic metastasis. In resected hepatic metastases from colorectal cancer, rate of hepatic recurrence with DNA diploid pattern was lower than that with DNA aneuploid pattern. Survival of patients with DNA diploid metastases (71% alive at 5 years) was significantly better than that of patients with DNA aneuploid metastases (21% alive at 5 years) (p less than 0.05). These results demonstrated that flow cytometric DNA ploidy measurements may have prognostic value for patients with hepatic metastases from colorectal cancer.     

The prognostic role of the DNA ploidy pattern in colorectal cancer analysis using paraffin-embedded tissue by an improved method

To assess the prognostic value of DNA ploidy in colorectal cancer, compared with the histopathological findings, paraffin-embedded surgical specimens from 330 patients who underwent resection for primary adenocarcinoma were studied using a new modified method of flow cytometry. Of these specimens, 141 were DNA diploid and 189, DNA aneuploid, among which there were 3 DNA hypodiploid lesions. Of the ten variables studied in curative resection, DNA ploidy ranked fourth in prognostic significance according to the linear trend by the ₂ test, after nodal status, grade of cellular differentiation, and degree of invasive growth, if the DNA ploidy pattern was classified into three categories. Conversely, DNA ploidy was the sixth most significant factor if DNA hypodiploidy was included in the DNA aneuploidy. The Cox multivariate analysis showed that DNA ploidy was one of the five significant factors independently determining prognosis; however, if adjustment for the modified Dukes' stage was made by the Mantel-Haenszel test, the survival difference between the diploid and aneuploid groups did not reach a statistically significant level. Thus, we conclude that from a practical point of view, DNA ploidy is not an essential factor which must be combined with histopathological variables for a better prediction of patient outcome.     

DNA ploidy pattern and tumour spread in gastric cancer

The DNA ploidy pattern of gastric cancer was studied in 58 patients to investigate the heterogeneity between primary tumour and metastases. In both primary tumours and lymph node metastases, diploid patterns accounted for 33 per cent, whereas all liver metastases were aneuploid. The percentage of polyploid cells was higher in the liver metastases than in primary tumours and lymph node metastases. When the heterogeneity of DNA ploidy pattern between primary tumour and metastasis was evaluated, diploid tumours had a significantly lower rate of lymph node metastasis heterogeneity than aneuploid tumours. When the DNA ploidy pattern and survival were evaluated, the patients who had a diploid pattern in both primary tumour and metastasis had a significantly higher survival rate than the patients who had an aneuploid pattern in the primary tumour and metastasis (57 per cent versus 26 per cent at 5 years). These data suggest that cell heterogeneity is a common phenomenon in gastric cancer, and this may be important in the evolution of the disease. Furthermore, the role of the DNA ploidy pattern as a prognostic factor is emphasized.     

Primary adenocarcinoma of the bladder: favorable prognostic significance of deoxyribonucleic acid diploidy measured by flow cytometry

Flow cytometric nuclear deoxyribonucleic acid ploidy analysis was done successfully on 38 specimens of primary bladder adenocarcinoma treated between 1954 and 1985. Of the specimens 10 (26%) were deoxyribonucleic acid diploid, 8 (21%) were tetraploid and 20 (53%) were aneuploid. Distribution of ploidy patterns between the 14 histological low grade and 24 high grade tumors was similar. Of 38 tumors 35 (92%) showed muscle invasion. One tumor arose in a previously exstrophied bladder, 10 were of urachal origin and 27 arose in an anatomically normal bladder. Of the urachal origin tumors 80% were deoxyribonucleic acid aneuploid. At 5 and 10 years after diagnosis 80 and 70%, respectively, of the patients with diploid tumors were free of disease. By contrast, at 5 and 10 years after treatment only 20 and 12%, respectively, of the patients with nondiploid tumors have not had disease progression (p less than 0.001 log-rank test). None of the 6 patients with diploid, high grade, high stage, muscle invasive tumors had subsequent progression. In contrast, 16 of 17 patients (94%) with high grade, high stage, nondiploid tumors had either local or distant tumor recurrence (p less than 0.0005). Nuclear deoxyribonucleic acid ploidy pattern appears to be the most significant prognostic information currently available to stratify expected prognosis for patients with muscle invasive adenocarcinoma of the bladder. This test probably should be a standard tool in the clinical management of patients with this rare bladder malignancy.     

Prognostic factors for disease progression in patients with renal cell carcinoma

IntroductionMore than 40% of patients with renal cell carcinoma present with disease progression after surgery. The objective of the current study was to identify a clinically useful set of prognostic factors that would correlate significantly with the capacity of progression.Material and methodsThe authors studied 252 patients with renal cell carcinoma who underwent radical nephrectomy. Followup ranged from 12-246 months (median 36 months). Several morphologic parameters of the tumors were considered. DNA content was analyzed by flow cytometry and tumor size was determined from the surgical specimen. A Cox proportional hazards regression model was used to identify significant independent prognostic factors for disease progression.ResultsA total of 224 out of 252 were available for suitable histograms. Of the 224 patients, 95 (42,4%) were aneuploid tumors, 106 (47,2%) were organ-confined renal cell carcinoma and 87 (39,74%) presented disease progression. At 5 and 10 years of followup, disease free survival was found to be 66,31% and 62,23%, respectively. Univariate analysis revealed that DNA ploidy, Furhman grade and stage (TNM) had a statistically significant predictive value for disease progression. Survival univariate analysis found a worse probability of survival for aneuploid tumors, grade III-IV tumors, non organ-confined tumors and conventional and undiferentiated tumors. Using multivariate survival analyses, Furhman grade, stage (TNM) and DNA ploidy were the only independent prognostic factors. So, the probability of death for aneuploid tumor was 1,7 times higher than for diploid tumors.ConclusionsStage, DNA content and Furhman grade were the only significant independent predictors of disease progression. Tumoral size and histological type did not provide more additional information.