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.     

The relationship of flow cytometric DNA analysis and clinicopathology in small-intestinal carcinoids

Paraffin-embedded archival tissue samples were used for nuclear deoxyribonucleic acid (DNA) content study by flow cytometry on 56 surgically resected, primary, small-intestinal carcinoid tumors. Sample preparation was carried out using the methods of Hedley and Vindelov. To reduce nuclear aggregation, a procedure of sonication was also performed. Nineteen (34%) cases were DNA diploid, 34 (61%) cases showed significantly increased 4C peak (DNA tetraploid), and only three (5%) cases were DNA aneuploid. Cell cycle phase analysis revealed that carcinoid tumors had significantly higher G2% than those of nontumor control tumors. However, there was no significant correlation between clinical parameters and both DNA ploidy pattern and cell cycle phase analysis. Although the difference in survival between patients with DNA nondiploid tumors and DNA diploid tumors was not significant, all of the patients with DNA aneuploid tumor had poor prognosis followed by death from carcinoid tumor.     

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 analysis of the DNA content in the urinary bladder cancers treated by radical cystectomy and pre-operative irradiation]

The DNA ploidy of bladder cancers treated by radical cystectomy following pre-operative irradiation was analyzed by flow cytometry using paraffin embedded samples. The DNA ploidy and its changes by irradiation were studied. We used flow cytometry in 30 patients with transitional cell carcinoma of the bladder who received pre-operative irradiation (40 Gy in 24 patients, 20 Gy in 5 patients and 60 Gy in one) with follow-up for at least 3 years. Total 140 paraffin embedded samples (4.6 samples per one patient) were available. The effects of therapy were related to the DNA patterns before irradiation and to the DNA ploidy changes after irradiation. 1. Eight DNA diploid tumors and twenty-two DNA aneuploid ones were detected before irradiation. Although diploid group didn't change its DNA ploidy after irradiation, of 22 aneuploid tumors 18 were changed to DNA diploid and 4 were not changed in their ploidy. 2. The tumor eradicating effect of irradiation was shown to be higher (p < 0.05) in the diploid group (5 of 8, 63%) than in the aneuploid group (5 of 22, 23%). 3. Overall survival rates were discussed in 3 groups (A, B and C), the group A was 10 of tumor free and 3 diploid tumors after irradiation, the group B was 13 of aneuploid tumors which changed to diploid ones and the group C was 4 of persistent aneuploid tumors. Each of 5 year survival rate was 100% (A), 58% (B) and 0% (C). Overall survival for C group was significantly shorter than for other groups (p < 0.01).(ABSTRACT TRUNCATED AT 250 WORDS)     

Deoxyribonucleic acid flow cytometry in invasive bladder carcinoma: a possible predictor for successful bladder preservation following transurethral surgery and chemotherapy-radiotherapy.

Tumor deoxyribonucleic acid (DNA) ploidy was evaluated as an objective parameter that may correlate better with the responsiveness of bladder cancer to chemotherapy plus radiotherapy than do clinical features or histopathological subtypes. A total of 40 patients with localized muscle-invading bladder cancer (clinical stages T2 to T4) underwent prospective treatment on a potential bladder preserving protocol. Tumors of 37 of the 40 patients were analyzed by DNA flow cytometry of multiple paraffin embedded specimens. Transurethral resection, neoadjuvant chemotherapy and 40 Gy. radiotherapy plus cisplatin were followed by urological reevaluation of the tumor (a complete response required a negative biopsy and a negative urine cytology study). A total of 7 noncomplete response patients underwent immediate radical cystectomy whereas the full bladder sparing treatment (radiotherapy to 64.80 Gy. plus cisplatin) was given to 23 complete response patients and 7 noncomplete response patients who were unsuited for surgery. Of the tumors 22 (59%) were purely aneuploid and 10 (27%) were purely diploid. Five tumors contained aneuploid and diploid patterns in different tumor specimens (partly diploid). Current status with a 30-month median followup in surviving patients includes an 82% overall survival rate in the aneuploid group versus 47% in the diploid/partly diploid group. Of all the patients 68% are free of invasive tumor: 82% in the aneuploid group versus 47% in the diploid/partly diploid group. By multivariate analysis pure aneuploidy was significantly (p = 0.05) correlated with freedom from invasive tumor in the bladder (either as persistence or as recurrence) and approached significance (p = 0.08) in correlation with overall patient survival. A longer observation time will be required to confirm this unexpectedly good outcome for patients with pure aneuploid tumors. We hypothesize that pretreatment DNA ploidy status may become a clinically useful prognostic factor in selecting patients for successful treatment with transurethral surgery and neoadjuvant chemotherapy plus radiotherapy.     

Flow cytometric analysis of the nuclear DNA content of hepatocellular carcinoma

The nuclear DNA content of 77 resected specimens from 65 cases of hepatocellular carcinoma (HCC) was measured by means of flow cytometry. The DNA index (DI) was calculated and the correlation between the DNA ploidy pattern and clinicopathological findings was studied. In the cases of HCC with a diameter of less than 5 cm, the 3-year survival rate of the aneuploid cases was 44.5 per cent, which was significantly lower than the 91.4 per cent of the diploid cases (p less than 0.001). Serum AFP levels were over 1000 ng/ml in 46.4 per cent of the aneuploid tumors and 18.5 per cent of the diploid tumors (p less than 0.05). The DI's were investigated in several sites of the same tumor and no difference was seen among the different sites in 16 out of 17 tumors. From 8 recurrent cases out of 12 who underwent a second resection, seven did not show any significant differences in DI from their primary tumor. On the other hand, four cases of second primary tumors showed different DI's to those of their first primary tumor. Intra-hepatic metastatic tumors exhibited the same DI's as their primary tumors. Thus, the nuclear DNA ploidy pattern may serve as a stable and valuable marker in predicting the malignant potential and prognosis of HCC.     

Flow cytometric DNA analysis of neuroblastoma: correlation with prognostic variables and survival of patients

The DNA ploidy of tumor tissues obtained from 41 patients with neuroblastoma, or ganglioneuroblastoma, which is the histologic variant of neuroblastoma, was determined by flow cytometry. The DNA ploidy was diploid in 7 tumors (5 neuroblastomas and 2 ganglioneuroblastomas) and aneuploid in 34 tumors (20 neuroblastomas and 14 ganglioneuroblastomas). The DNA ploidy of tumor cells did not correlate with the survival of patients and there was no correlation between the DNA ploidy of tumor cells and such prognostic variables, histology, primary site, staging of tumors and operative curability.     

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.     

Flow cytometric DNA analysis of neuroblastoma: Correlation with prognostic variables and survival of patients

The DNA ploidy of tumor tissues obtained from 41 patients with neuroblastoma, or ganglioneuroblastoma, which is the histologic variant of neuroblastoma, was determined by flow cytometry. The DNA ploidy was diploid in 7 tumors (5 neuroblastomas and 2 ganglioneuroblastomas) and aneuploid in 34 tumors (20 neuroblastomas and 14 ganglioneuroblastomas). The DNA ploidy of tumor cells did not correlate with the survival of patients and there was no correlation between the DNA ploidy of tumor cells and such prognostic variables, histology, primary site, staging of tumors and operative curability.     

Adrenocortical carcinoma: nuclear deoxyribonucleic acid ploidy studied by flow cytometry

Nuclear deoxyribonucleic acid (DNA) ploidy studies with use of paraffin-embedded specimens were performed by flow cytometry on 52 adrenocortical carcinomas. Specimens were prepared by the combined techniques of Hedley and Vindel?v. Clinical course was obtained by chart review and follow-up examination. Nine (17%) tumors had a normal (diploid) DNA pattern, 13 (25%) were DNA tetraploid, and 30 (58%) were DNA aneuploid. The DNA aneuploid group was subdivided: 18 tumors with one stemline and 12 tumors with two stemlines of abnormal DNA cells. For tumors that were resected for cure, the 5-year Kaplan-Meier disease-free survival rates of the five patients with DNA diploid tumors and of the six patients with DNA tetraploid tumors were 80% and 33%, respectively. For 21 patients of whom 12 had one-stemline and nine had two-stemline DNA aneuploid tumors, the survival was 67% and 0%, respectively. Following palliative resection, the 4-year survival rates of the four patients with DNA diploid, seven with DNA tetraploid, five (omitting one with short follow-up) with one-stemline DNA aneuploid, and three with two-stemline DNA aneuploid tumors were 0%, 0%, 0%, and 33%, respectively. Although adrenocortical carcinoma is in general markedly aggressive, the addition of nuclear DNA ploidy studies may help to identify certain groups of patients who have a relatively favorable prognosis.     

Treatment of hepatocellular carcinoma from the aspect of nuclear DNA analysis]

Flow cytometric DNA analysis was done on 302 resected hepatocellular carcinoma and 30 specimens by fine needle aspiration biopsy. Patients with DNA aneuploid tumors had a significant worse prognosis than those with DNA diploid tumors in subdivided groups by curability (p less than 0.01). The recurrence after hepatectomy occurred frequently in DNA aneuploid or non-curative resected group. In a cox multivariate analysis, nuclear DNA content provided significant prognostic value (p = 0.008) as did vascular invasion and intrahepatic metastasis. For the multiple tumors of hepatocellular carcinoma, it is helpful to get the DNA ploidy pattern to diagnose tumors with intrahepatic metastasis or multicentric origin. Nuclear DNA analysis by fine needle aspiration biopsy was useful to determine the treatment method of hepatocellular carcinoma.     

Evaluation of DNA ploidy combined with a cytometric proliferation index of imprints from core needle biopsies in prostate cancer.

OBJECTIVE: To evaluate if DNA ploidy analysis with a proliferation index (PI) derived from DNA cytometry of imprints from core needle biopsies predicts disease progression in patients with prostate cancer. METHODS: Touch imprints were done on a consecutive series of core needle biopsies taken by the same urologist from 240 patients with suspected prostate cancer, 137 (46%) of whom were found to have prostate cancer and included in the study. Scattered cells to the right of the image cytometry (ICM) ploidy-establishing peak, the S-phase fraction, and those in the G2M area of the ICM DNA histograms, were counted in percent of the total number of tumor cells, this value being designated the ICM PI. Based on previous results in archival fine needle aspiration material, the following classification was used: DNA group I, diploid tumor with a low PI; DNA group II, diploid tumor with an intermediate PI and tetraploid tumor with a low or intermediate PI, and DNA group III, diploid or tetraploid tumor with high PI and all tumors with an aneuploid pattern. RESULTS: Correlation was found to exist between DNA groups I-III and Gleason score (GS) (p < 0.0001), T-stage (p = 0.006), M-stage (p = 0.009) and disease progression (p < 0.0001). Among the 39 patients who had curative treatment and GS 5-7, the progression-free survival rate was 100% in DNA group I, as compared with only 38% in DNA group II and 55% in DNA group III within the follow-up period (p = 0.008). CONCLUSION: DNA ploidy combined with a PI derived from image cytometry of imprints from core needle biopsies yields additional prognostic information in patients with GS 5-7. Diploid tumors with a low PI (DNA group I) are associated with a low risk of disease progression.     

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.     

Heterogeneity of salivary gland tumors studied by flow cytometry

Intratumor DNA heterogeneity was investigated by flow cytometric analysis of multiple samples taken from different sites of 8 benign and 16 malignant primarily resected salivary gland tumors. All benign tumors had diploid DNA content. The overall incidence of DNA diploidy in 16 malignant cases examined was 50%. Intratumor differences in DNA ploidy were observed in four malignant tumors (25%); 2 of these 4 heterogenous tumors contained both aneuploid and diploid cell clones. The remaining 12 tumors showed a homogeneous DNA content in the different specimens; 8 were diploid, 3 aneuploid, and 1 was polypoid. The DNA nondiploid tumors were clinically more advanced than the DNA diploid ones (p < 0.01). The tumor proliferation rate (fraction of cells in S-phase) was higher in DNA nondiploid samples than in diploid ones (p < 0.01). The DNA nondiploid tumors seemed to recur more often than DNA diploid ones did. The data emphasize the usefulness of DNA measurements for the characterization of malignant salivary gland tumors but also the importance of adequate sampling in assessing their DNA ploidy.     

Analysis of flow cytometric nuclear DNA content of the thymoma and its relationship to malignant intensity

Determination was made of the nuclear DNA content of paraffin-embedded specimens of resected thymoma using flow cytometry in 39 patients. Relationships among nuclear DNA content, clinicopathological findings and prognosis were studied. The frequency of DNA aneuploid tumors was 30.8%, 6.7% in stage I (15 patients), 28.6% in stage II (7 patients), 44% in stage III (9 patients) and 62.5% in stage IV (8 patients) according to Masaoka's classification, indicating more in increase with advancing stage and significant (p less than 0.05) more increase invasive thymoma than in noninvasive thymoma. The 5 year and 10 year survival rates of DNA diploid tumors were each 94%, while DNA aneuploid tumors, 75% and 45%. Patients with DNA aneuploid tumors showed less favorable prognosis than those with DNA diploid tumors. Similar results were found in patients with invasive thymoma and in those on whom subtotal or partial resection were performed. The present results indicate DNA aneuploid tumors to have greater malignant intensity than DNA diploid tumors in resected thymoma. Flow cytometric nuclear DNA content analysis provides useful biological data and new indices for evaluating the malignancy of resected thymoma.     

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.     

"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.     

Molecular biological observations in some rare neoplasms: DNA ploidy and cell cycle data in liposarcoma and malignant fibrous histiocytoma

Introduction: The prognostic significance of tumor DNA ploidy and cell cycle analysis for long-term survival has been examined in 19 patients with liposarcoma or malignant fibrous histiocytoma. In many cases, different tumor areas of primary tumors and local recurrences have been analyzed to reveal intratumoral heterogeneity. Results: Among the primary tumors, there were eight aneuploid tumors, three of which showed diploid and aneuploid tumor regions. Correlations among DNA ploidy, grading, percentage of S-phase cells and infiltrative growth pattern of the tumors could be demonstrated. Poorly differentiated tumors (G3) showed aneuploidy in six of eight patients. Aneuploid tumors showed S-phase cells in 17.2% (range 3.2–38.1%), which was higher than the percentage of S-phase cells in diploid tumors (9.4%, range 2.1–27.4%). Aneuploid tumors showed a more infiltrative growth pattern (6 of 8 patients) than diploid tumors (6 of 11 patients). The median survival time of patients with diploid tumors was 86.5 months (8–144 months), compared with 40.9 months (11–54 months) for patients with aneuploid tumors. Conclusion: DNA ploidy and percentage of S-phase cells may be considered as prognostic factors. Received: 12 June 1998 Accepted: 14 September 1998     

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.     

Flow cytometric analysis of the nuclear DNA content of hepatocellular carcinoma

The nuclear DNA content of 77 resected specimens from 65 cases of hepatocellular carcinoma (HCC) was measured by means of flow cytometry. The DNA index (DI) was calculated and the correlation between the DNA ploidy pattern and clinicopathological findings was studied. In the cases of HCC with a diameter of less than 5 cm, the 3-year survival rate of the aneuploid cases was 44.5 per cent, which was significantly lower the 91.4 per cent of the diploid cases (p<0.001). Serum AFP levels were over 1000 ng/ml in 46.4 per cent of the aneuploid tumors and 18.5 per cent of the diploid tumors (p<0.05). The DI’s were investigated in several sites of the same tumor and no difference was seen among the different sites in 16 out of 17 tumors. From 8 recurrent cases out of 12 who underwent a second resection, seven did not show any significant differences in DI from their primary tumor. On the other hand, four cases of second primary tumors showed different DI’s to those of their first primary tumor. Intra-hepatic metastatic tumors exhibited the same DI’s as their primary tumors. Thus, the nuclear DNA ploidy pattern may serve as a stable and valuable marker in predicting the malignant potential and prognosis of HCC.