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.     

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.     

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.     

Formal hepatic resection of colorectal liver metastases. Ploidy and prognosis.

Fifty consecutive patients who underwent 52 formal hepatic resections (excluding isolated wedge resections) for metastatic colorectal cancer were analyzed to determine whether DNA content was of prognostic significance. The Dukes' stages of the colorectal primaries were: A (10%), B (20%), C (40%), D (28%), and unknown in 2%. Four patients whose liver metastases were discovered at the time of resection of the primary bowel cancer underwent concomitant liver resection, and the remaining patients underwent delayed resections. The hepatic resections performed were right lobectomy (50%), extended right lobectomy (19%), left lobectomy (13%), left lateral segmentectomy (6%), left lobectomy and right wedge (6%), extended left lobectomy (4%), and right lobectomy and left wedge (2%). The overall morbidity rate was 29%. The in-hospital mortality rate was 9%. As of November 1991, 36 patients have recurred. The 5-year actuarial survival was 28%. Flow cytometry could be performed on 37 archival specimens, 15 of which were found to be diploid whereas 22 were aneuploid. All metastases from Dukes A colorectal primaries demonstrated a diploid DNA content. In addition, there was no difference in actuarial survival between diploid and aneuploid tumors. These data suggest that in selected patients, formal hepatic resection of colorectal liver metastases can be performed with an acceptable morbidity rate, mortality rate, and survival, but ploidy of the resected tumor is not of prognostic significance.     

Relationship between DNA ploidy and survival in patients with primary breast cancer

The DNA ploidy of breast cancer tissue from paraffin blocks was measured by flow cytometry in 117 patients whose disease had been detected and treated with surgery between 1974 and 1976. Patients with aneuploid tumours had positive axillary nodes and distant metastases more often than those with diploid tumours. Aneuploid tumours were more common in postmenopausal than premenopausal women. The S-phase fraction (SPF) was significantly higher in aneuploid than in diploid tumours and positive axillary lymph nodes were found in 26 per cent of the patients who had a tumour with a SPF below the median (4.8 per cent) and in 48 per cent of those with tumours with SPF values above the median. At the primary clinical investigation 2 per cent of the patients with diploid tumours and 6 per cent of those with aneuploid tumours had distant metastases. During the follow-up, the proportion of patients with distant metastases increased to 42 and 72 per cent, respectively. With a follow-up of 11.5 years, the DNA aneuploidy of the tumour showed a significant association with decreased survival. Thirty-three per cent of patients with diploid and 65 per cent of patients with aneuploid tumours had died from breast cancer during the follow-up (P less than 0.001). All patients with hypertetraploid or multiploid tumours died from breast cancer. High SPF values were associated more closely with distant metastases or death during the follow-up than low SPF values. Our results suggest that DNA ploidy measured by flow cytometry from paraffin embedded tissue blocks of human breast cancer can be used to predict the aggressiveness of the tumour and the survival of the patients.     

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.     

DNA content and c-erbB-2 oncoprotein expression in hepatic metastases from colorectal carcinoma--in relation to clinicopathologic findings and prognosis

The nuclear DNA ploidy pattern and c-erbB-2 oncoprotein expression in primary and metastatic lesions were investigated using paraffin-embedded materials from 44 cases of colorectal carcinoma with hepatic metastases and 45 cases without hepatic metastases. The frequency of aneuploidy and positive staining of c-erbB-2 in primary tumor with hepatic metastases were significantly higher compared with those without hepatic metastases (p less than 0.05). There were significant correlations between diameter of metastases and DNA ploidy pattern of the metastases and between metachronous metastases, degree of metastases, vessel involvement and DNA ploidy pattern of the primary tumor. Positive staining of c-erbB-2 was detected more frequently with the advancement of depth of tumor invasion. There was no significant correlation between DNA ploidy pattern and c-erbB-2 expression. In the survival of patients whose primary tumor and hepatic metastases were resected, it was shown that DNA ploidy pattern of metastases was the most important independent prognostic factor. Expression of c-erbB-2 in primary tumor predicted the hepatic metastases.     

Flow cytometric DNA analysis in rectal carcinomas

The cellular DNA content was measured with flow cytometry from paraffin-embedded material in 329 patients and metastatic tumors of the liver from the rectum in 11 patients. The classification of the DNA ploidy pattern is as follows: A stem cell peak with a DNA index of 0.9-1.1 is defined as DNA diploid tumor and DNA aneuploid tumor is that with a DNA index greater than or equal to 1.1. There was a good correlation of DNA indices (r = 0.997) obtained from flesh and corresponding paraffin-embedded specimens. It is concluded that accurate determination of DNA index from paraffin-embedded materials is possible in the majority of cases. DNA ploidy of primary tumor cells correlated with clinicopathological findings such as lymphatic invasion, vascular invasion, lymph node metastasis and hepatic metastasis (p less than 0.01), but did not correlate with extramural carcinoma invasion. The cumulative survival rate (Kaplan-Meier) of curatively resected rectal carcinomas was worse in DNA aneuploid than in DNA diploid tumors (p less than 0.01). These observation showed that the determination of DNA ploidy in rectal carcinomas may prove to be of prognostic value.     

Metastatic mode and DNA ploidy in gastric carcinoma

We studied the amounts of nuclear DNA in gastric cancer metastases histologically and cytochemically by flow cytometry, which was performed retrospectively on paraffin-embedded specimens from 95 patients. At surgery, all cases of aneuploid cancer were positive for lymph node metastases. Liver metastases were frequently seen in aneuploid cancer (63%, P<0.01), while lung metastases were the most common in diploid cancer (50%, P<0.05). The incidence of peritoneal metastasis was high in undifferentiated diploid cancer (72%, P<0.01). Local lymph node recurrence after surgery was more common in aneuploid than in diploid cancer (P<0.01). The incidence of bone and distant lymph node metastasis was found to be strongly dependant on tissue differentiation. The DNA ploidy pattern is thus considered to be closely linked to lymph node, liver, and lung metastases in gastric cancer.     

Tumor DNA content in resectable, primary colorectal carcinoma.

Tumor DNA content was measured in patients with colorectal carcinoma in order to determine whether tumor ploidy was a prognostic indicator independent of standard clinical and pathologic characteristics. One hundred forty-seven patients were analyzed who had their primary resectable colorectal carcinomas resected with curative intent from 1974 to 1981. Aneuploid colorectal cancers (i.e., tumors with abnormal DNA content) tended to be less well-differentiated, to invade the serosa or extend beyond, and to have lymph node metastases rather than diploid tumors (i.e., tumors with normal DNA content). A significantly increased rate of recurrent disease was demonstrated in patients with aneuploid tumors as opposed to those with diploid tumors (46.7% vs. 4.8%, respectively [p less than 0.001]). In addition, patients with aneuploid tumors exhibited a significantly decreased disease-free and overall survival in comparison with patients with diploid colorectal carcinomas. A Cox regression analysis demonstrated that tumor DNA content was the single most important factor in predicting recurrence or death from colorectal carcinoma.     

"Benign" metastasizing giant cell tumor: evaluation of nuclear DNA patterns by flow cytometry

Nuclear deoxyribonucleic acid (DNA) ploidy was determined by flow cytometry for nine histologically benign giant cell tumors that developed systemic metastases and for eight tumors that did not metastasize. Specimens from the primary tumor, local recurrences, and pulmonary metastases were evaluated. No feature of the DNA ploidy pattern was identified to distinguish giant cell tumors that metastasized from those that did not. The mean percentage of diploid (G0/G1 peak, 2C) cells was 81% in the metastasizing group and 80% in the nonmetastasizing group. The DNA ploidy pattern of the primary tumors was not different from that of their metastases. No DNA aneuploid patterns were observed among the benign tumors.     

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.     

Flow cytometric analysis of DNA ploidy pattern in advanced gastric cancer and its relationship with prognosis

Cell nuclear DNA content was determined by flow cytometric analysis in 270 patients with advanced gastric cancer. Aneuploid DNA content was observed in 150 patients (55.6%). The DNA ploidy pattern was the third significantly prognostic factor behind peritoneal dissemination and liver metastasis in Cox regression multivariate analysis. About the relationship between DNA ploidy pattern and other prognostic factors, peritoneal dissemination and wall invasion ratio of aneuploid were significantly higher than those of diploid (p less than 0.01). Five-year survival rate of diploidy patients was significantly higher than that of aneuploidy patients. In stage I, five-year survival rate of patients with diploid tumor was 83.3% and that of patients with aneuploid tumor was 70.0%. In stage II, that of patients with diploid tumor was 81.3% and that of patients with aneuploid tumor was 66.7%. In stage III, that of patients with diploid tumor was 71.2% and that of patients with aneuploid tumor was 25.1%. In stage IV, that of patients with diploid tumor was 31.6% and that of patients with aneuploid tumor was 2.6%. Furthermore in the curative case, that of patients with diploid tumor was 77.2% and that of patients with aneuploid tumor was 48.2%. Aneuploid case has significantly worse prognosis in curative operation.     

Prognostic value of deoxyribonucleic acid content in metastatic renal cell carcinoma

The deoxyribonucleic acid content of tumor specimens from 23 patients with metastatic renal cell carcinoma was analyzed prospectively by flow cytometry and static cytophotometry. Of the primary tumors 10 (43 per cent) were homogeneously diploid or near diploid in 8 samples studied from each tumor and 13 (57 per cent) had an aneuploid deoxyribonucleic acid content in 1 to 8 samples. At the end of followup 9 of 10 patients with diploid or near diploid primary tumors were alive, compared to only 1 of 13 with aneuploid primary tumors. Patients with homogeneously diploid or near diploid tumors survived significantly longer compared to those with aneuploid tumor deoxyribonucleic acid content (p less than 0.001). After excision of solitary diploid or near diploid metastases 4 patients had no evidence of disease. In 3 of these patients the primary tumors were diploid or near diploid, whereas 1 had 1 aneuploid and 7 diploid or near diploid samples in the primary tumor. In 10 other patients 28 metastases revealed concordance in deoxyribonucleic acid content with the primary tumors. Our results indicate that deoxyribonucleic acid content might be a useful prognostic discriminator with implications for the clinical management of patients with metastatic renal cell carcinoma.     

The clinical usefulness of flow cytometric DNA analysis in prostatic cancer]

Flow cytometry was used to measure the DNA content in archival paraffin-embedded prostatic cancer specimens from 54 patients with known outcomes. The specimens were obtained by transurethral resection of the prostate. DNA ploidy as a predictor of prognosis was compared with histological grade and clinical stage. Although no significant correlation between histological grade or clinical stage and ploidy pattern was demonstrated, an increased percentage of DNA aneuploid tumors was seen in higher histological grade and in advanced clinical stage. The survival rate calculated by Kaplan-Meier analysis showed that DNA ploidy pattern was a more reliable indicator to predict survival probability than histological grade or clinical stage. All patients with a near diploid pattern (11 patients) survived more than 5 years, whereas all those with an aneuploid pattern (21 patients) died within 3.5 years. Of 22 patients with a tetraploid pattern, 15 died of tumor progression within 5 years. The remaining 7 patients with favorable outcome had a relatively lower proliferation index (less than 65) in DNA histogram and none of them suffered from stage D disease. In conclusion, the results from this retrospective study suggest that flow cytometric DNA analysis in prostatic cancer would be useful as a means of providing prognostic information.     

DNA ploidy and S phase fraction in human bladder cancer. Relation to survival and histological grade (WHO)

The nuclear DNA content of archival paraffin-embedded bladder cancer samples (70 patients) of WHO grades I-III has been measured by flow cytometry. The female/male ratio was 15/55. The mean follow-up time was 13 years (range 9.6-22.0 years). 37 of 70 (53%) patients had DNA index 1.0 (diploid DNA content), and the remaining 33 (47%) patients had an aneuploid tumor. There was no significant difference in the age (mean +/- SD) of the patients having a diploid (66 +/- 9 years) or an aneuploid tumor (68 +/- 11 years) at the time of diagnosis. 47 deaths occurred during the follow-up period; 24 (51%) of these were due to bladder cancer (12 diploid, 12 aneuploid tumors). No significant difference was found after radical treatment during the disease-free interval (mean +/- SD) between diploid (48 +/- 45 months) and aneuploid (35.5 +/- 35 months) groups of patients. Recurrences during the follow-up period were equally common among aneuploid and diploid tumors. A statistically significant relation between histological grade and survival could be demonstrated, but DNA ploidy and S phase fraction had little prognostic value in this respect. There was no statistically significant difference in survival between aneuploid (30%) and diploid (35%) groups of tumors during the follow-up period. The study suggests that flow cytometric determination of nuclear DNA ploidy from paraffin-embedded samples in bladder tumors does not add to the prognostic power of subjective histological grading.     

Malignancy in gastric carcinoma, with special reference to the DNA ploidy and proliferative activity

Analysis of DNA ploidy patterns was performed on 129 primary gastric cancers and the results correlated with histologic findings and in vivo bromodeoxyuridine (BrdU) labeling. Forty-nine cases were diploid (38%) and 80 cases were aneuploid. There was no correlation between DNA ploidy and histologic type. In aneuploid cancers, incidence of lymphatic invasion, lymph node metastasis and rate of advanced cases were significantly higher than those in diploid tumors. During the follow-up period of 5-10 years, 23 of the 40 patients (55%) with aneuploid tumors died of disease within 3-120 months. Only 13 of the 36 patients (36%) with diploid tumors died of disease. The BrdU labeling indices (BrdU LI) were from 2.8% to 26.7%, with a mean of 10.4%. There was no correlation between BrdU LI and histologic type or stage. The mean BrdU LI of early cancers was 8.1%, of advanced cancers, 11.9%. BrdU LI of cancers with lymphatic invasion or lymph node metastasis was higher than those without them. The mean BrdU LI of diploid cancer was 6.0%, of aneuploid cancers, 11.9%. There was a good correlation between BrdU labeling indices and DNA ploidy patterns. These results indicate that determination of DNA ploidy patterns and growth fractions by BrdU labeling may be useful in the conjecture of prognosis of the patient and in the selection of patients from various modalities.     

Prognostic significance of DNA ploidy in colorectal cancer: a prospective flow cytometric study

A prospective study of prognostic factors has been carried out in a group of 123 consecutive patients with colorectal cancer. The fate of all patients is known at 3 years after operation. Clinical and pathological data were recorded at the time of presentation and operation, and the patients have been subject to regular postoperative review. DNA ploidy status was determined by flow cytometry. In all, 39 (33 per cent) patients had DNA diploid tumours and 80 (67 per cent) patients had DNA aneuploid tumours. In four cases, tumour material was not obtained. The patients with DNA aneuploid tumours had a worse prognosis than those with DNA diploid tumours, but this was only seen in those patients classified as Dukes' B. In a Cox's regression analysis, the surgeon's assessment of operability was the strongest predictor of survival, followed by the pathological classification and the patient's age. After these factors had been considered, the DNA ploidy status conferred no independent survival value.     

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.     

Comparison between diploid and aneuploid hepatocellular carcinomas: a flow cytometric study.

Nuclear DNA content of hepatocellular carcinoma (HCC) was estimated by flow cytometry after hepatic resection in 91 patients during the past 5 years. There were 53 diploid and 38 aneuploid tumours. Clinicopathological features were compared retrospectively between the patients with diploid and those with aneuploid HCC. DNA ploidy did not show any correlation with age, sex, alcohol abuse, hepatitis B virus, serum alpha-fetoprotein level or underlying liver disease. Histopathologically, the incidence of HCC less than 2 cm in diameter tended to be higher in the diploid group but no difference was seen for large tumours (greater than 5 cm). The grade of tumour differentiation also tended to be higher in this group of small HCC. The ploidy pattern did not influence the rate of capsule or daughter nodule formation, or venous invasion. There were no significant differences in survival rate or in the incidence and time of intrahepatic tumour recurrence between the two groups. This study may indicate that nuclear DNA ploidy is not a particularly predictive factor for the surgical treatment of HCC.