Diploid Cases Research Articles

Increased incidence of spontaneous apoptosis in the bone marrow of hyperdiploid childhood acute lymphoblastic leukemia

Hyperdiploidy of 51-65 chromosomes is associated with a good prognosis in childhood B-lineage acute lymphoblastic leukemia (ALL). Blasts from childhood ALL patients with a hyperdiploid karyotype have a tendency to apoptosis when cultured on stromal layers in vitro. In this study, we apply a novel method to investigate the relationship between apoptosis and hyperdiploidy in lymphoblasts of childhood ALL. The DNA content of individual ALL blasts in Feulgen-stained archival bone marrow smears can be determined by static cytometry. TUNEL (TdT-mediated dUTP-biotin nick end labeling) detects the DNA degradation associated with apoptosis. We performed TUNEL in situ sequential to DNA ploidy analysis in archival bone marrow smears from 12 patients with childhood ALL. Five patients were diploid and seven were hyperdiploid (51-65 chromosomes) by conventional cytogenetic analysis. In the five diploid cases, the percentage of TUNEL-positive blasts ranged from 1.0% to 1.3%; in the seven hyperdiploid cases, the percentage of TUNEL-positive blasts ranged from 3.6% to 9.0%. Comparing TUNEL and corresponding Feulgen images, we found that apoptotic blasts were predominantly of high DNA ploidy in both diploid and hyperdiploid cases. The mean DNA value of apoptotic blasts was larger than that of the total blast population in each case. The results demonstrate an increased incidence of spontaneous apoptosis in situ of hyperdiploid blasts in ALL bone marrow and indicate that this phenomenon is not restricted to in vitro cultures. The findings provide a possible rationale for the good prognosis associated with hyperdiploid childhood ALL.

Quantitative FISH analysis on interphase nuclei may improve diagnosis of DNA diploid breast cancers.

The detection of DNA aneuploid cells using flow cytometry is an indication for the presence of tumor cells, but when DNA diploid cells are found in 25-33% of the cases, the diagnostic and prognostic significance of DNA ploidy is more limited. We analyzed interphase nuclei after in situ hybridization and using image cytometry on 50 breast tumors with diploid DNA content to investigate whether early chromosome rearrangements were detectable and if their occurrence was clinically significant. Imbalances between the two arms of chromosome 1 were found in 55% of the cases and values ranged from 1.5-3.0. Comparison with histological data showed that Grade I tumors mainly have imbalances (67%) and that Grade III tumors were mainly without the imbalance (67%), whereas Grade II tumors were intermediate (50% imbalance). These data suggest that the diagnosis of DNA diploid cases may be improved by using interphase FISH. In addition, the data also indicates that early breast tumors may have different genetic origins, which is important in the comprehension of tumor malignancy in early stages, especially for preinvasive lesions.

Flow cytometric DNA analysis of benign hyperfunctioning parathyroid glands: significant difference in the S phase fraction and proliferative index between adenomas and hyperplasias

Flow cytometric DNA analysis was performed to measure the DNA content of benign parathyroid tumours in patients with primary hyperparathyroidism. DNA analysis of paraffin-embedded parathyroid samples was performed on 51 parathyroid glands from 29 patients after parathyroidectomy. Histopathology showed parathyroid adenoma in 25 cases and hyperplasia in four patients. DNA ploidy status, DNA index (DI), percentage of cells in S phase and proliferative index (PI) were determined. Normal cells from normal glands were all diploid. DNA cytometry showed 12 aneuploid and 13 diploid adenomas. There were 12 diploid and four aneuploid hyperplastic glands. Incidence of aneuploid DNA histograms did not show a statistically significant difference between adenomas and hyperplasias (P=0.216). Mean S phase fraction was 3.45% in adenomas and 1.53% in hyperplasias (P= 0.015). Mean PI was 6.48% in adenomas and 2.78% in hyperplastic parathyroid glands. This difference was statistically significant (P=0.006). Diploid cases had a mean PI of 4.78% and aneuploid glands a mean PI of 7.7% (P=0.08). Aneuploid DNA content did not reveal statistically significant correlation with age, gender, pre-operative Ca, alkaline phosphatase, i-PTH levels, and tumour size. The mean S phase fraction and PI were 2.25% and 4.78% in diploid glands, and 4.5% and 7.7% in aneuploid cases. Aneuploid DNA content may be present in benign parathyroid diseases, but not in normal parathyroid glands. Aneuploid DNA histograms and higher PI occur more often in adenomas compared with hyperplasias, but the nuclear DNA analysis is unable to make a distinction between adenomas and hyperplasias.

A semiautomated test for microsatellite instability and its significance for the prognosis of sporadic endometrial cancer in northern Norway.

Archival histologic material from 105 women (median age 62 years) treated for endometrial cancer was investigated for the replication error phenotype indicated by the observation of widespread microsatellite instability (MSI). Polymerase chain reaction (PCR) of DNA isolated from paraffin-embedded tissue was analyzed for MSI using six microsatellite loci with a fluorescent-based detection system. Flow cytometry and morphometric investigation were performed in the same material for each of the patients. Twenty percent (21 of 105) of screened endometrial cancers were found to have high MSI at two or more of the loci tested. The mean detection frequency per marker was highest in the dinucleotide repeat sequence, D2S123, and the mononucleotide repeat sequences amplified by Bat 25 and Bat 26. Death from endometrial cancer was not related to the occurrence of MSI (p=0.6). There was no significant association between MSI and FIGO stage (p=0.5), myometrial invasion depth (p=0.8), histological grade (p=0.3), or vessel invasion (p=0.5). There were, however, more MSI cases among the group of diploid cases compared with the aneuploid and tetraploid group. MSI is not a valuable prognosticator for survival of sporadic endometrial cancer, and diploid cases are significantly more often MSI positive than aneuploid and tetraploid cases.

Biopathological significance of single cell DNA aneuploidy measured by static cytometry in breast cancer

DNA ploidy image analysis in breast carcinomas has occasionally revealed cases with a diploid DNA content but with a single hypertetraploid element, the so-called single cell aneuploidy (SCA). To identify the biologic significance of this little known phenomenon, we selected 40 cases with SCA from a series of 599 consecutive operable breast carcinomas.The clinical, pathological and biological characteristics of SCA cases were compared with those of a control group of 40 pure diploid breast carcinomas. Hormonal receptor status, proliferative indexes (Ki-67) and p53 overexpression were determined immunohistochemically and quantitatively evaluated by image analysis. The overexpression of c-erbB-2 was determined semiquantitatively.SCA was observed in 6.6% of cases (40 of 599) and in 17% of otherwise diploid cases (40 of 236). Breast cancers with SCA occur in younger women (mean age 54.75 y vs 61.12 y, P<0.05), are smaller (mean diameter 20.00 vs 21.62 mm), less differentiated (percentage of G3 cases 13.2 vs 2.9), more frequently hormone independent (ER positivity 70.0 vs 77.5%; PgR positivity 57.5 vs 77.5%) and have a greater cell proliferative activity (mean S-phase: 4.6 vs 2.8, P<0.05), (percentage of Ki-67 immunostaining: 24.1 vs 19.7%). There is also more overexpression of c-erbB-2 and P53, particularly in pT1 cases in which the percentage of c-erbB-2 positive cells is 54 vs 32 (P value is not significant) and the percentage of p53 is 29 vs 4 (P<0.05).According to these results SCA may be a reliable marker of genetic instability and of greater biologic aggressiveness. Image cytometry of DNA content may be a cost effective means of identifying breast cancer patients with an increased risk of tumour recurrences despite otherwise favourable prognostic parameters.

DNA ploidy and S-phase fraction in carcinoma of the gallbladder related to histopathology, number of gallstones and survival.

Gallstones are a risk factor for the development of gallbladder cancer. We studied DNA ploidy and cell cycle composition by flow cytometry in archival specimens from 52 gall bladder carcinomas in relation to histopathological grade, tumour stage, gallstone number and survival. 69% of the gallbladder carcinomas showed aneuploidy. All tumours with single stones (N=11) were aneuploid while only 61% of tumours with multiple stones (N=41) were aneuploid (p=0.002). DNA aneuploidy was related to increase in T‐category (p=0.01), grade (p=0.02), and nuclear pleomorphism (p=0.0005). The distribution of DNA ploidy shifted from tetraploid in low stage towards triploid positions in high stage tumours (p=0.02) combined with higher S‐phase values in triploid tumours (p=0.05). S‐phase fraction increased during development from normal tissue to dysplasia, cancer in situ and cancer in diploid cases (p=0.0002), and further at the change from diploid to aneuploid (p=0.004). At a median cancer specific survival time of four months patients with diploid tumours had a better survival than those with aneuploid tumours (p=0.02). In multivariate analysis of the tumour characteristic, only T‐category and tumour grade were independent prognostic factors. The shift from diploid to aneuploid and the further shift of ploidy within aneuploid tumours are in agreement with the concept of a clonal development of gallbladder cancer. These changes are combined with a stepwise increase in the fraction of S‐phase cells. Low frequency of symptoms in single stone patients may be the reason for detection of malignancy at a late stage of tumour development.

Role of DNA flow cytometry and image cytometry on effusion fluid

The objective of the study was to assess the value of DNA flow cytometry (FCM) and image cytometry (ICM) as an adjunct to routine diagnostic cytology. In this prospective study, 100 consecutive effusion fluids were studied for routine cytology, DNA FCM, and in selected cases, ICM. One half of the centrifuged fluid sample was used for routine cytology and the remaining portion was used for DNA FCM. Nuclear area, nuclear diameter, nuclear perimeter, nuclear convex perimeter, nuclear roundess, and nuclear convex area were measured on at least 100 cells by ICM in cytologically malignant or DNA aneuploid cases along with control cases. Clinical follow-up was done in all cases. There were 22 cytologically malignant cases and 78 cytologically benign cases. Among the 22 cytologically malignant cases, there were 11 aneuploid and diploid cases each by DNA FCM. Out of 78 cytologically benign cases, six (7.7%) were aneuploid by DNA FCM. Smears of these cases showed predominantly reactive mesothelial cells, but the DNA histograms showed hypodiploid (one), hyperdiploid (three), tetraploid (one), and hypertetraploid (one) aneuploidy. Follow-up of these cases showed clinical or histologic features of malignancy except in one case of tetraploid aneuploidy, which did not show any features of malignancy and responded well to antitubercular therapy. Therefore, out of 27 malignant effusions, DNA FCM picked up 16 cases and routine cytology detected 22 cases. Sensitivity and specificity of DNA FCM were thus 59.25% and 98.63%, respectively. There was a statistically significant difference (Student's unpaired t-test, P < 0.05) between cytologically malignant cases and control benign cases in all the nuclear morphometric parameters except for nuclear roundness. There was, however, no statistically significant difference of nuclear morphometric parameters between cytologically benign vs. DNA aneuploid cases and control benign cases. DNA FCM is a useful adjunct for routine diagnostic cytology. Visual diagnostic cytology and morphometric digital microscopy miss some cases of malignancy which can be detected by DNA flow cytometry. Diagn. Cytopathol. 2000;22:81-85.

Relationship between Dysplasia, p53 Protein Accumulation, DNA Ploidy, and Glut1 Overexpression in Barrett Metaplasia

There is a need for molecular markers of malignant progression in Barrett metaplasia (BM). The aim of this study is to determine the relationship between dysplasia, p53 protein accumulation, DNA ploidy, and Glut1 in BM. Sections of esophageal biopsy specimens from 120 patients with BM were evaluated for dysplasia, p53 protein, and Glut1 expression by immunohistochemistry, and DNA ploidy by Feulgen stain and image analysis. In cases with diploid DNA histograms, the percentage cells in the G0G1 and G2M phases of the cell cycle were determined. Of 108 diploid cases 19 (28%) of 69 cases with G0G1 > or = 90% or G2M > or = 8.33% were p53-positive, in contrast to only 1 (3%) of 39 cases with lower G0G1 or G2M (P = 0.0008). Of 32 p53-positive cases 11 (32%) were aneuploid, in contrast to none (0%) of 88 p53-negative cases (P < 0.0001). Ten (91%) of 11 aneuploid cases were high-grade dysplasial adenocarcinoma (HGD/CA), compared with only 1 (1%) of 109 diploid cases (P < 0.0001). Five (45%) of 11 cases with HGD/CA were Glut1-positive, in contrast to none (0%) of 109 cases without HGD/CA (P < 0.0001). Our data strongly suggest that in BM, after oxidative DNA damage, as a result of gastroesophageal reflux, there is an increase in the percentage of cells in the G0G1 or G2M phases of the cell cycle to enable repair of damaged DNA; in some of these cases this is followed sequentially by p53 gene mutation and protein accumulation, DNA aneuploidy, HGD, and CA with or without Glut1 overexpression. These events can be detected in routinely processed biopsy samples.

S-phase fraction related to prognosis in localised prostate cancer. No specific significance of chromosome 7 gain or deletion of 7q31.1.

A flow-cytometric (FCM) and fluorescence in situ hybridization (FISH) study was performed in 153 patients with clinically localised prostate cancer (PC) to evaluate retrospectively the prognostic significance of DNA ploidy, S-phase fraction (SPF) and chromosome 7 copy number. Deletions in 7q31.1 were analysed in a subset of 26 tumours. The mean follow-up time was 6 years (range 4-16 years). Twelve cases of benign prostatic hyperplasia (BPH) were studied as a control. Chromosome 7 enumeration and deletion studies were conducted using the alpha-satellite D7Z1 probe and a cosmid probe specific for the marker D7S522 on 7q31.1. Higher SPF was associated with shorter overall survival and shorter time to local progression and metastasis. Near diploid (DNA index 1.05-1.20) cases had a lower frequency of metastases and lower Gleason scores than aneuploid cases. Increased absolute chromosome 7 copy number (centromere count) was associated with higher Gleason score, higher SPF and shorter local progression-free and prostate cancer survival. Absolute chromosome 7 copy number was concordant with FCM DNA ploidy in the majority (75%) of cases. Relative gain or loss of chromosome 7 (centromere counts compared to ploidy) was infrequent, and no correlation was found with clinical parameters. Deletions in 7q31.1 were infrequent. Our results indicate that in localised PC (i) SPF is a prognostic factor, (ii) absolute chromosome 7 copy number is concordant with the ploidy status of the tumour (relative gain or loss of chromosome 7 is infrequent and has no independent prognostic value) and (iii) the frequency of deletions in 7q31.1 is low and not correlated with clinical outcome.

Interphase cytogenetic and AgNOR analyses of hydatidiform moles.

AIM: To determine the potential value of interphase cytogenetic and argyrophilic nucleolar organiser region (AgNOR) analyses in the diagnosis and classification of hydatidiform moles. METHODS: Serial tissue sections from 37...

Cytogenetic Abnormalities of Primary Breast Carcinoma: Comparative Analysis of Fluorescence In Situ Hybridization (FISH) and Flow Cytometry

: Breast cancer is a heterogeneous disease that is frequently associated with genetic alterations. Nonrandom chromosomal changes are assumed to play a role in breast tumorigenesis and progression. These changes can be detected by standard cytogenetics, which is technologically cumbersome, or by flow cytometry, which only provides a general overview of genetic alterations by ploidy status. Alternatively, fluorescence in situ hybridization (FISH) is a recently developed technique that has the capability to detect chromosomal changes in individual nuclei with preserved morphology of breast cancer. Our study was designed to investigate the molecular cytogenetic basis of nonrandom chromosomal aberrations in breast carcinoma by FISH and compare this data with flow cytometry. Large paraffin sections and 5 μm sections of 57 cases of invasive ductal carcinoma and 5 normal control samples were subjected to ploidy analysis by standard flow cytometry and FISH. By flow, 27 (47.4%) cases were aneuploid; while 30 cases were diploid. FISH, performed using a panel of six α-satellite centromeric probes for chromosomes 1, 7, 8, 11, 17, and X, detected 37 (64.9%) aneuploid tumor, while 20 tumors were diploid. Vast intratumor heterogeneity (75%) occurred in 27 of the 37 aneuploid tumors. FISH also identified aneuploidy in 50% of cases that were interpreted as diploid by flow. The FISH and flow results were concordant in 15 diploid cases and 22 aneuploid cases, yielding an overall agreement of 64.9%. This agreement was significant above that expected by chance (kappa = 0.309, p = 0.007). Sensitivity and specificity of FISH for aneuploidy detected by flow were 81.5% and 50.0%, respectively. Discordant FISH aneuploid tumors were characterized by gain or loss of 1-3 chromosomes. Normal controls were consistently diploid by both flow and FISH. Our data suggest that FISH is more sensitive in detecting aneuploidy than flow cytometry and provides valuable genetic data of breast cancer cells.

DNA aneuploidy by flow cytometry is an independent prognostic factor in gastric cancer.

In the present study the prognostic value of both DNA ploidy and the proliferative activity of tomour cells were studied in a series of 76 consecutive patients suffering from gastric tumours. DNA ploidy and the proliferative index (as measured by the percentage of S-phase cells) were determined by flow cytometry using fresh tumour specimens.The presence of DNA aneuploid clones by flow cytometry was detected in 62% of the cases (mean DNA index of 1.63 ± 0.46; range 1.08–2.92), the mean proportion of S-phase cells being of 18.4 ± 11.5%. In comparison with diploid cases, aneuploid tumours showed a higher proliferative activity (cases with more than 15% S-phase cells: 18.4% versus 6.1%, p = 0.0001) as well as a higher incidence of node involvement (95% versus 68%, p = 0.001). By contrast, no significant differences were detected with respect to sex, age, histologic grade and type, clinical stage, tumour size and the incidence of extranodal involvement.Upon grouping the patients according to the proportion of S-phase cells no significant differences were observed for the clinical and biological parameters explored except for an association between a high percentage of S-phase cells and the presence of DNA aneuploidy (40% versus 96%, p = 0.0001). Regarding survival the presence of DNA aneuploidy was significantly associated with poor outcome as compared to the diploid cases (median of 15 versus 26 months, p = 0.005). By contrast, the proportion of S-phase cells did not predict patients’s outcome.Multivariate analysis of prognostic factors showed that the presence of DNA aneuploidy (p = 0.003) together with the histologic type (p = 0.03) and the existence of extranodal metastases (p = 0.05) were the best combination of prognostic factors for survival prediction.

Functional adrenal cortical tumors in childhood: a study of ploidy, p53-protein and nucleolar organizer regions (AgNORs) as prognostic markers.

Prognostic markers in pediatric adrenal cortical tumors are difficult to define. We determined the ploidy, immunostaining of p53-protein and number of nucleolar organizer regions (AgNORs) in 16 such tumors and related them to clinical outcome, tumor weight (TW) and histologic Weiss' criteria. Eleven females and 5 males aged 0.4 to 15.6 years were followed for 8.7 years; 10 presented Cushing's and 6 virilization syndrome. Diploid (n = 4, x TW = 269 g, range: 17-800 g) and near-diploid tumors (n = 3, x TW = 55 g, range: 20-85 g) had good outcome, Weiss' criteria were 0-7, and p53 reactivity was negative in all. Among the aneuploid tumors (n = 9, x TW = 298 g, range: 7-1000 g), 6 had good outcome, 2 presented metastasis and 1 was lost to follow-up; Weiss' criteria were 2-8 and p53 reactivity was positive in 3 tumors (2 of them of malignant evolution). AgNORs number was not different in cases of good or poor outcome (3.65 +/- 1.9 vs 2.83 +/- 1.1). Our findings indicate that diploid and near-diploid cases had always a good outcome regardless of tumor weight. In aneuploid cases, tumor weights < 100 g had good outcome, while those > 750 g had poor prognosis. Malignant tumors were aneuploid and had reactivity to p53-protein. Good outcome in aneuploid tumors < 100 g is probably due to early treatment. The expression of p53-protein appears as a promising marker of poor prognosis. Weiss' criteria and AgNORs were not useful in the present series.

The role of deoxyribonucleic acid image cytometric and interphase cytogenetic analyses in the differential diagnosis, prognosis, and clinical follow-up of hydatidiform moles: A report from the central molar registration in the Netherlands

Objectives: To assess the value of deoxyribonucleic acid ploidy in the differential diagnosis and clinical follow-up of hydatidiform moles, the histopathologic features, deoxyribonucleic acid ploidy, and clinical follow-up were compared in 347 cases: 143 complete moles, 52 partial moles, and 152 abortions, of which 56 cases were hydropic abortions with histologic features of triploidy but lacked trophoblastic hyperplasia. Study Design: In all cases deoxyribonucleic acid image cytometry was performed, and in 85 of these cases interphase cytogenetics was also performed. Results: With use of deoxyribonucleic acid image cytometry and interphase cytogenetics, a bimodal polyploid deoxyribonucleic acid pattern was present in 97% of complete moles, 27% of partial moles, and 4% of abortions. All these cases of partial mole were reclassified to complete mole on the basis of this deoxyribonucleic acid pattern and the histopathologic features in spite of the presence of fetal blood cells, amnion, or yolk sac. Deoxyribonucleic acid triploidy was found in 95% of the remaining partial moles, in 77% of hydropic abortions with histologic features of triploidy, and in 14% of the remaining abortions. Reliable differentiation between deoxyribonucleic acid triploid partial moles and hydropic abortions with histologic features of triploidy was not possible on basis of the histopathologic features (trophoblastic hyperplasia) or 3.5c exceeding rates. Deoxyribonucleic acid diploidy was found in 1% of complete moles, 23% of hydropic abortions with features of triploidy, and 78% of the remaining abortions. Deoxyribonucleic acid tetraploidy was rarely found (1% of complete moles, 2% of partial moles, 1% of abortions). Persistent gestational trophoblastic disease developed in 33% of the bimodal deoxyribonucleic acid polyploid cases (all complete moles), in 1% of the diploid cases (concerning one of the two diploid complete moles), and in 1% of the triploid cases (partial moles). Conclusion: Deoxyribonucleic acid analysis is essential in the diagnosis of hydatidiform moles to decide on clinical follow-up.

Analysis of DNA ploidy using fresh frozen tissues of head and neck squamous cell carcinomas

The DNA ploidy of fresh frozen tissues of head and neck squamous cell carcinomas was determined by flow cytometry to investigate whether DNA ploidy is correlated with various clinical and pathological parameters. The subjects were 51 patients who had been treated radically by our department. The DNA ploidy pattern was classified into three types, diploid, single aneuploid and multiploid, according to the DNA index and the DNA histogram. This is our original classification. No particular correlation could be detected between the DNA ploidy pattern and sex, age, primary tumour site or disease stage. The degree of tissue differentiation tended to be poorer in aneuploid tissues than in diploid tissues. The efficacy of chemotherapy was higher in aneuploid cases than in diploid cases. The recurrence rate was significantly lower in diploid cases than in multiploid cases. When disease stage, degree of histological differentiation, efficacy of the chemotherapy and the DNA ploidy pattern were subjected to multivariate analysis for correlation with the prognosis, the DNA ploidy pattern showed the highest correlation. The results suggest that the DNA ploidy as analyzed by flow cytometry can be used as an important prognostic factor.

Flow cytometric analysis of tumor DNA profile related to response to treatment and survival in small-cell lung cancer.

Flow cytometric (FCM) analysis of tumor DNA ploidy and S-phase fraction (SPF) has been widely used to predict prognosis and treatment response in many malignant tumors, but rarely in small-cell lung cancer (SCLC). In the present study, tumor DNA ploidy and SPF were measured from paraffin-embedded tumor biopsy samples of 36 small-cell lung cancer patients treated with combination chemotherapy and radiotherapy. Aneuploidy was detected in 69% of the tumors. There was a statistically non-significant trend towards more aneuploidy among extensive disease (ED) patients as compared to patients with limited disease (LD): 80% versus 65%, respectively (p = 0.69). The mean SPF was 21.3% (+/-7.6) in patients with LD and 29.0% (+/-5.3) in patients with ED, the difference (7.6%) being statistically significant (p = 0.008, 95% CI for the difference 2.2-13.1). No significant differences was detected in the survival of aneuploid and diploid patients or patients with low (< or = 24.9%) and high (> 24.9%) SPF. Similarly, no significant difference was observed between aneuploid and diploid cases in relation to response to treatment or response duration. It is concluded that the difference detected in the SPF with LD and ED of SCLC may indicate the biological aggressiveness of extensive SCLC.

DNA ploidy and proliferating cell nuclear antigen image analysis of peritoneal and pleural effusions. A possible diagnostic role.

To determine the role of DNA and proliferating cell nuclear antigen (PCNA) image analysis (IA) in enhancing the diagnostic sensitivity of conventional cytology (CC). The histopathologic and clinical data on 87 consecutive pleural and peritoneal effusions were used to evaluate the accuracy of CC and DNA IA results. CC showed a sensitivity, specificity, positive predictive value (PPV) and negative predictive value (NPV) of 65%, 100%, 100% and 62%, respectively. Aneuploidy peaks were seen in 49 cases; 47 of them were true positives. Thirty of 38 diploid cases were true negatives. The sensitivity, specificity, PPV and NPV were 85%, 94%, 96% and 80%, respectively. There were positive correlations between DNA ploidy profile and PCNA proliferative index (PI), (R = .697) and significant differences in PCNA PI between malignant and benign effusions (P < .001). The DNA IA PI by PCNA can be used as a complementary diagnostic tool with CC in cytologically inconclusive cases.

IN RELATION TO NUCLEAR DNA PLOIDY PATTERN, p53 GENE MUTATION AND CLINICOPATHOLOGICAL FINDINGS IN COLORECTAL ADENOMAS AND CANCERS

The expression of p53 mutation and its association with nuclear DNA ploidy patterns and clinicopathological findings were studied incolorectal adenomas and cancers. The subjects were 81 cases of colorectal cancer and 19 cases of colorectal adenoma, in which the p53 gene mutation was measured by RT-PCR-SSCP or flowcytometry. There were 4 cases of mild atypia, 8 cases of moderate, and 7 cases of severe in the colorectal adenomas, and 21 casee of Dukes's classification A, 19 cases of B, 24 cases of C and 17 cases of D in the colorectal cancers. In DNA ploidy pattern, there were 39 (48.1%) diploidy cancers and 42 (51.9%) aneuploidy cancers, significant relations to pathological features including depth of invasion, lymph node metastasis nad stage (p<0.05) were noted in the aneuploid cases. No p53 gene mutation was detected in mild and moderate atypia adenomas, but it was detected in a severe atypia adenoma (1/7). As to the cancers, p53 gene mutation was detected in two out of five focal cancers (40.0%), and in 46 out of all 81 colorectal cancers (56.8%). There was no relation to pathological findings. It was inferred that p53 gene mutation might participate with carcinogenesis. In the correlation with DNA ploidy patterns, p53 gene mutation occurred in 17 out of 39 diploid cases (43.6%) and in 29 out 42 aneuploid cases (67.7%). And p53 gene mutation was detected in nine out of ten recurrent cases. It was suggested that the expression of p53 mutation might cause DNA abnormality and might be an important factor for recurrence.

Association between flow cytometric S-phase fraction and apoptotic rate in breast cancer.

Tumor heterogeneity may adversely affect the flow cytometric measurement of S-phase fraction (SPF) in breast cancer specimens, and 10-20% of breast cancer specimens are not evaluable by flow cytometry due to technical factors such as debris, high coefficients of variation, poor specimen quality, or small sample size. Therefore, we performed this study on 207 specimens of breast cancer in order to determine if the apoptotic rate (AR) could serve as a useful adjunct to flow cytometric SPF measurements in breast cancers. The average AR in each specimen was determined by microscopic examination of tumor tissue that was specifically stained for apoptotic bodies by a commercially available TUNEL (Tdt-mediated dUTP digoxigenin nick end labelling) assay kit. The mean AR (4.5 +/- 3.0, n = 37) in the high SPF (> 10%) group was significantly (P < 0.01) higher than the mean AR (1.3 +/- 1.2, n = 72) in the low SPF (< 6%) group. Although the distributions of AR values in the two groups had substantial overlap, AR values greater than 5.5 per high power field (h.p.f.) were not observed in the low SPF cases but were present in 13 out of 37 cases with a high SPF. Simple linear regression analyses relating SPF to the mean AR in 57 DNA diploid cases and 41 DNA aneuploid cases yielded a minimal correlation (r2 = 0.21) between the two parameters only in the DNA aneuploid group. We conclude that an elevated AR has an association with high SPF in breast cancers, but the association is too weak to permit the general use of AR as a predictor of SPF. Our study also identified a subset of breast cancers with both a high SPF (> 10%) and a high AR (> 5.5/h.p.f.) that may warrant further investigation to determine its clinical significance.

Impact of tumor cell kinetics on the outcome of induction chemotherapy in oral carcinoma patients

Pretherapeutic S-phase fractions of 68 carcinomas of the tongue and the floor of mouth were analysed by DNA flow cytometry immediately before intra-arterial induction chemotherapy with cisplatin and epirubicin or 5-fluorouracil. Patients with diploid tumors had an excellent 5-year survival rate of 89% in contrast to only 31% in the aneuploid group. Cell proliferation had no impact on the outcome of diploid cases. Node-positive patients with aneuploid tumors had a poor 18% 5-year survival rate, independent of S-phase fractions. Survival was significantly worse in nodenegative patients with high proliferating aneuploid tumors (45%) compared to the low-proliferative N-0 group (83%).