DNA image cytometry. A prognostic tool in squamous cell carcinoma of the esophagus?

In 45 patients who underwent an esophagus resection due to a squamous cell carcinoma, in addition to the TNM classification and usual morphologic criteria, the paraffin-embedded material underwent deparaffinization, was rehydrated, and was mechanically and enzymatically processed into a single-cell solution. For evaluating the DNA histogram this was analyzed with the help of automatic single-cell cytophotometric study. The method, contrary to that of flow cytometric study, allows for the selective analysis of tumor cells due to the electronically, previously given selection criteria, whereas artifacts, stroma, and infection cells remain excluded from analysis. The multivariate analysis shows that the prognosis is only correlated with DNA content of the tumor cells. Patients with diploid tumors had a median survival time of 32 months as compared with the 22 months of patients with hypotriploid tumors, and 6.5 months with hypertriploid tumors. DNA cytometric analysis should be included when diagnosing patients with esophagus carcinoma.     

Independent prognostic value of ploidy in colorectal cancer. A prospective study using image cytometry

In a prospective study, the DNA content of Feulgen-stained nuclei obtained from fresh samples of 211 colorectal adenocarcinomas was evaluated by means of image analysis. The DNA histogram classification took into account aneuploidy and S-phase fraction for diploid cases. No significant relationship was found between ploidy and sex, age, preoperative carcinoembryonic antigen (CEA), size of the tumor, histologic differentiation, or Dukes' stage. Aneuploidy was more frequently encountered in distal tumors. Preoperative CEA, histologic differentiation, Dukes' stage, and ploidy were individually associated with overall survival. In Dukes' A, B, and C tumors, patients with normal and elevated CEA had no significant difference in overall survival. A relationship was apparent between disease-free survival and site, histologic differentiation, Dukes' stage, and ploidy. Multivariate overall survival analysis did not reveal independent prognostic significance of ploidy when all Dukes' stages were considered. In contrast, Dukes' stage, differentiation, and ploidy were good indicators of higher risk of colorectal cancer-related death in patients undergoing curative surgery. Dukes' stage and ploidy were also indicators for recurrence. Thus, routine histopathologic characteristics should be used in combination with quantitative cytologic features for the definition of a relevant prognostic index in colorectal cancer.     

Image cytometry for DNA analysis in endometrial carcinoma correlated with other prognostic parameters

We evaluated by static cytometry DNA ploidy parameters in 30 stage I-IV endometrial carcinomas and correlated these data with standard clinical-pathological features and disease-free period. We observed a direct correlation between either non-diploid DNA content and deeper myometrial invasion (p<0.02) or D.I.greater than or equal to 1.2 and M2-M3 tumors (p<0.009). The Kaplan Meier survival curves illustrate a more rapid relapse of disease associated with non-diploidy, high 5cExR, high level of proliferation and D.I.greater than or equal to 1.2. while Cox regression model gave relative hazards for disease recurrence of 4, 6, 3.7 and 2.1 for non-diploidy, D.I.greater than or equal to 1.2, high 5cExR and high level of proliferation respectively. This prospective study confirmed the prognostic value of DNA Index, and its usefulness in clinical practice especially in stage I cases, otherwise characterised by favourable standard prognostic factors, is discussed.     

Flow cytometry and quantitative immunohistochemical study of cell cycle regulation proteins in invasive breast carcinoma: prognostic significance

BACKGROUND: Between January 11, 1991 and January 8, 1992, 104 patients with previously untreated, invasive, primitive breast carcinoma were admitted to the authors' hospital. METHODS: For each patient, flow cytometry DNA analyses on frozen samples and on immunohistochemical staining were performed, including Ki-67, cyclin A, p53, and p21(waf1) (p21), with assessment of the percentages of positive nuclei were assessed. Correlations with classic clinicopathologic data and survival (overall, metastasis free, or recurrence free) and a multivariate analysis were performed. RESULTS: After a multivariate analysis according to a Cox model that was stratified by age, tumor size, tumor grade, lymph node status, and receptor status, among the factors studied, the presence of p21 was the unique remaining prognostic factor for patients with invasive breast carcinoma. Because of the lack of a correlation between p21 and proliferative factors (Ki-67, S-phase, and cyclin A), the authors combined p21 with those markers and found that, for the different combinations, after statistical analysis, only p21 combined with S-phase or with cyclin A and lymph node status were salient survival prognostic factors. CONCLUSIONS: Immunohistochemical study of proteins involved in the cell cycle and assessment of proliferative activity using flow cytometric DNA analysis aided the authors in singling out correlations of cyclin A and S-phase, along with p21, with metastasis free survival and overall survival in patients with invasive breast carcinoma. These promising results will require confirmation in a larger series of patients.     

Feulgen cytometry in simultaneous endometrial and ovarian carcinoma

The modal DNA value was measured in six patients with simultaneous endometrial and ovarian carcinoma by Feulgen static cytometry. One patient with low-grade endometrial and ovarian carcinoma manifested diploid indices at both sites. Another patient demonstrated aneuploid ovarian carcinoma and diploid endometrial carcinoma, indicating that these were separate neoplasms. The remaining four patients with Stage III disease had aneuploid endometrial and ovarian carcinomas with identical DNA indices. These data support a single neoplastic process with metastasis in the latter four patients. There was good correlation with the clinicopathologic impression on the likelihood of synchronous primaries versus metastatic neoplasms. It was concluded that DNA analysis is a useful adjunct in assessing the probability that spatially separate neoplasms represent metastasis.     

A comparative study of DNA ploidy in 115 fresh-frozen breast carcinomas by image analysis versus flow cytometry.

BACKGROUND. Qualitative and quantitative analysis of cellular DNA content may be clinically useful in the prognostic evaluation of certain types of malignant tumors, including breast carcinoma. Flow cytometric (FCM) analysis has been the most frequently used procedure for DNA analysis, but it requires a reasonably large tissue sample. Computer-based image analysis (IA) now allows imprint, cytospin, and needle aspiration smear preparations and other small tissue samples to be used. METHODS. To resolve concern about the diagnostic efficacy of small tissue samples in the use of IA, the authors performed a comparative study of FCM analysis and IA using 115 fresh-frozen breast carcinomas. Feulgen-stained imprint preparations for IA and single-cell suspensions from the same fresh-frozen tissue for FCM analysis were used, and the respective histograms were compared. RESULTS. The results were concordant in 90.4% (104 of 115) of the cases, but 11 specimens yielded discordant data. IA provided histograms with a somewhat lower resolution and a relatively high coefficient of variation for the G0/G1 peak, thus rendering occasional tumors, which were near-diploid aneuploid by FCM analysis (four cases), not amenable to diagnosis by aneuploid characterization. In three additional cases, FCM analysis showed aneuploid hyperdiploid (two cases) and multiploid (one case) histograms, but IA only demonstrated a diploid peak. Conversely, in four other cases, aneuploid peaks were recognized only by IA. CONCLUSIONS. Computerized IA has significant advantages over FCM analysis, including lower cost, the ability to analyze very small specimens, the capability of detecting rare high ploidy cells, the capacity to classify cellular populations according to specific morphologic type, and the fact that no destructive enzyme or chemical digestion is required for specimen preparation, thereby preserving the integrity of fragile cells.     

Diagnostic and prognostic use of DNA image cytometry in cervical squamous intraepithelial lesions and invasive carcinoma

In the fight against cervical malignancy and its precursors, several adjuvant diagnostic methods have been proposed to increase the accuracy of cytologic and histologic diagnoses. Because chromosomal aneuploidy has been accepted as an early key event in tumorigenesis caused by genetic instability, the cytometric equivalent of chromosomal aneuploidy detected by DNA image cytometry (DNA-ICM) may serve as a marker of neoplasia. During the last decade, the appearance of a new generation of hardware with high processing and storage capacities, together with the development of appropriate software, has facilitated the development of high-performance DNA-ICM systems. International consensus on the clinical application of DNA-ICM has been reached. According to the statements of Task Force 8 of the International Consensus Conference on the Fight Against Cervical Cancer, indications for DNA-ICM include the identification of prospectively malignant cells in squamous intraepithelial lesions (SILs) and atypical squamous cells of undetermined significance (ASCUS). The European Society of Analytical Cellular Pathology consensus reports on DNA-ICM have provided standardized technical details on performance, terms, and algorithms for diagnostic data interpretation and quality-assurance procedures. Increasing biologic evidence and clinical data have confirmed the utility of DNA-ICM as an adjuvant method suitable for determining the diagnosis and prognosis of cervical intraepithelial lesions and invasive carcinoma. Patients with ASCUS and low-grade SIL diagnoses that reveal DNA euploidy may return for normal screening intervals, whereas the detection of DNA aneuploidy indicates that these lesions should be removed. Formerly a research tool, today, standardized DNA-ICM has become a useful and low-cost laboratory method to establish objectively and reproducibly an early diagnosis of prospectively progressive cervical intraepithelial lesions at a high-quality level. DNA-ICM may further contribute to the monitoring of treatment in patients with invasive cervical malignancies.     

Flow cytometry and squamous cell carcinoma of the maxillary sinus: A possible prognostic indicator for multimodality intervention

Malignant tumors of the paranasal sinuses constitute less than 1% of all malignancies. Unfortunately, few prognostic factors have been identified regarding the efficacy of interventional therapy. Patients with carcinoma of the maxillary sinus frequently present with an advanced stage of disease and multimodality therapy is often proposed. Although some tumors appear to be more sensitive to preoperative chemotherapy and radiation therapy, a method of predicting and evaluating tumor behavior has not been recognized. Introduced as a means of identifying cell populations with abnormalities in total DNA content, flow-cytometric DNA analysis provides a quick and reliable means of tumor characterization for many malignancies. Although nuclear DNA content has been implicated as a prognostic factor in an increasing number of tumor types, current data on the role of DNA content in head and neck carcinoma is conflicting and incomplete. To evaluate the role of flow-cytometric DNA analysis in predicting therapeutic alternatives, 22 patients with squamous cell carcinoma of the maxillary sinus were reviewed. Patient outcome and histopathologic grade were retrospectively compared with flow-cytometric evaluations of paraffin-embedded formalin-fixed tumor specimens. Four of sixteen tumors (25%) were found to be aneuploid and were also associated with an increased survival rate (p < 0.01). This initial data emphasizes the possible usefulness of DNA measurement for characterization of squamous cell carcinoma of the maxillary sinus and stresses the need for further evaluation.     

Independent prognostic importance of microvessel density in endometrial carcinoma.

Angiogenesis is thought to be an important factor for tumour growth and metastatic spread, and microvessel counts may provide useful prognostic information for several tumour types. To investigate the prognostic impact of angiogenesis in endometrial carcinoma patients, the intratumour microvessel density, which was determined immunohistochemically, has been related to survival. Sixty patients with endometrial carcinoma with long (median 19 years) and complete follow-up have been studied. Patients with increased mean microvessel density (MVDmean > 68 mm2) had a significantly shorter 5-year survival compared with the rest (57% vs 90%, P = 0.004). In multivariate survival analyses, MVDmean had an independent prognostic impact (P = 0.03) when FIGO stage, histological type, histological grade as well as nuclear p53 protein expression was adjusted for. These findings indicate that intratumour microvessel density may contribute additional prognostic information to that obtained from the known risk factors and may be helpful in identifying endometrial carcinoma patients at high risk for disease progression.     

子宫内膜癌115例临床病理与预后分析

目的探讨子宫内膜癌的临床,病理特点以及影响预后因素。方法对我院1997年~1999年间初治为手术治疗的115例子宫内膜癌患者的临床资料进行回顾性的分析。结果平均发病年龄56.7岁,在获访的90例中,手术病理分期Ⅰ期5年生存率97.6%,Ⅱ期的5年生存率90.9%,Ⅲ期61.9%,Ⅳ期0。Ⅰ、Ⅱ期与Ⅲ、Ⅳ期的5年生存率差异有显著性(P<0.05)。病理分级Ⅰ级的盆腔及腹主动脉旁淋巴结转移率为3.1%,(1/32),Ⅱ级为5.6%(3/54)Ⅲ级为33.3%(6/20),Ⅰ,Ⅱ级与Ⅲ级淋巴结转移的差异有显著性(P<0.05)。结论子宫内膜癌手术病理分期能客观判断预后,组织学分级与盆腔及腹主动脉旁淋巴结转移相关。     

Flow cytometry as a prognostic predictor in gastric linitis plastica

Gastric linitis plastica (GLP) is a diffusely infiltrating carcinoma of the stomach that is usually diagnosed in an advanced stage and associated with poor prognosis. Recent studies to evaluate ploidy of these tumors are not conclusive. We undertook a retrospective study of 43 surgically treated patients with GLP (27 males, 16 females, mean age 65 years) to see if ploidy could be used to predict outcome. Flow cytometric DNA analysis was performed on paraffin-embedded tissue using the modified Hedley technique. Mean follow-up interval was 11 months (1–72 months) with 18 (42%) alive at end of study. The remaining 25 (58%) died with a mean survival of 7 months. Lymph node status was positive in 31 (70%) and negative in 12 (30%) of patients. Twenty-nine (67%) of tumors were diploid; 14 (33%) were aneuploid. Statistical analysis revealed overall surcvival correlated significantly (P = 0.04) only with lymph node status. Diploid tumors had 18 (60%) positive and 11 (40%) negative lymph nodes, whereas aneuploid tumors had 13 (93%) positive and 1 (7%) negative nodes. DNA content correlated significantly (P = 0.05) with lymph node status, but not with overall survival. Tumors with positive lymph nodes were 18 (51%) diploid and 13 (42%) aneuploid; tumors with negative nodes were 11 (92%) diploid and 1 (8%) aneuploid. Conclusions: The majority of GLP tumors manifest diploid characteristics, and the presence or absence of lymph node metastasis is a major determinant in overall survival. © 1994 Wiley-Liss, Inc.     

子宫内膜癌的预后影响因素分析

背景与目的:子宫内膜癌的预后影响因素较多,但其中仅有少数因素对预后构成独立影响。本研究的目的在于探讨子宫内膜癌的独立预后影响因素。方法:对我院1990年1月至2000年12月间初治时行手术治疗的265例子宫内膜癌患者的临床资料进行回顾性研究,预后相关因素采用单因素分析及多因素相关回归分析,并进行逐步筛查。结果:本组病例的5年无瘤生存率及总生存率分别为83.3%和84.3%。单因素分析显示:临床分期、手术-病理分期、病理分级、组织学类型、肌层浸润深度、宫颈受累、淋巴结转移、腹腔液性质、脉管瘤栓及附件转移与5年无瘤生存率及总生存率有显著性相关(P<0.05),年龄、合并症因素与预后无显著性相关(P>0.05)。经多因素分析后得出,手术-病理分期、病理分级、肌层浸润深度及宫颈受累4个因素对子宫内膜癌患者的5年无瘤生存率及总生存率均产生显著性影响(P<0.05),临床分期仅对5年无瘤生存率有显著性影响(P<0.001),而对总生存率无显著性影响(P=0.074)。肌层浸润>50%者远处转移率(12.9%)明显高于≤50%者(0.6%)(P<0.001)。宫颈受累者的淋巴结转移率(21.1%)明显高于宫颈未受累者(3.6%)(P<0.001)。结论:FIGO分期、病理分级、肌层浸润深度及宫颈受累是子宫内膜癌独立的预后影响因素。在估计预后方面,手术-病理分期     

Flow cytometry for detection and evaluation of urinary bladder carcinoma.

Flow Cytometry (FCM) DNA assays of bladder irrigation specimens are now recognized as a clinically useful and reliable means of detecting and monitoring carcinoma of the bladder. This technique, which identifies carcinoma by the presence of an aneuploid population of cells, can be carried out on specimens obtained in an outpatient or hospital setting and is easily performed in any medium-sized laboratory. It is most sensitive to superficial and high grade tumors. Overall, nearly 80% of superficial carcinomas of bladder will have positive flow cytometry, comparing very favorably with conventional cytology. Until now, the widest clinical application of FCM has been in monitoring the conservative treatment of stage 0-1 flat and papillary carcinomas, but newly developed dual parameter measurements are capable of quantifying proliferative activity, oncogene expression, growth factor receptors, and other cellular features that may better characterize the biologic potential of these tumors and can be expected to aid in the selection and timing of treatment.     

Prospective multicenter validation confirms the prognostic superiority of the endometrial carcinoma prognostic index in international Federation of gynecology and obstetrics stage 1 and 2 endometrial carcinoma.

PURPOSE: To validate the prognostic value of the endometrial carcinoma prognostic index (ECPI; combined myometrium invasion, flow cytometric DNA ploidy, and morphometric mean shortest nuclear axis [MSNA]) versus classic prognosticators. PATIENTS AND METHODS: Prospective multicenter ECPI analysis was conducted in 463 endometrial carcinomas with a median of 6.5 years (range, 1 to 10 years) follow-up, review of pathology features, and univariate (Kaplan-Meier) and multivariate (Cox) analyses. RESULTS: Initial routine and review diagnoses varied considerably (invasion depth, 11%; type, 20%; grade, 34%; vessel invasion, 72%); the review diagnoses were stronger prognostically. In International Federation of Gynecology and Obstetrics stage 1 (after histopathologic examination; pFIGO-1; n = 372; 38 deaths occurred as a result of disease [10.2%]), DNA ploidy was prognostic in hysterectomies (P <.00001) but not in curettages (P =.06). ECPI was a stronger prognostic indicator than other features. ECPI, MSNA, and DNA ploidy were also prognostic in pFIGO-1B and -1C subgroups. Multivariate analysis in pFIGO-1 showed that uterine MSNA < or = versus > 7.93 microm (hazard ratio [HR], 3.4) and grade (as 1 + 2 v 3; HR, 2.6) added to the ECPI (HR, 32), but only in patients with an unfavorable ECPI of > 0.87. Adjuvant radiotherapy was not an independent prognostic factor in any of the subgroups. In pFIGO-2 (n = 46), ECPI, DNA-ploidy, and age (< or = 64, > 64 years) were significant. In FIGO-3 (n = 31) and FIGO-4 (n = 14), none of the classic or other features analyzed was of prognostic value, which explains why in previous studies using different mixtures of FIGO stages, DNA ploidy prognostic results varied. CONCLUSION: In endometrial carcinoma, DNA-ploidy is prognostic in hysterectomy and not in curettage samples. The ECPI is prognostically much stronger than the classic features widely used for therapy triage in pFIGO-1 and -2.     

60岁以上子宫内膜癌患者的临床病理特点及预后因素分析

目的分析60岁以上老年子宫内膜癌患者临床病理特点及预后因素.方法从1981年1月至2002年12月,北京大学第一临床医院共收治子宫内膜癌患者323例,收集其显著的临床病理资料.把患者分为两组,第一组年龄≥60岁,共108例,第二组为对照组,年龄＜60岁,共215例.比较两组患者的临床病理特点、治疗措施及预后.单因素及多因素分析影响子宫内膜癌患者的预后因素.结果第一组患者手术病理期别为Ⅲ、Ⅳ期的占31.5%,高于第二组的20%,(P＜0.05).两组患者的组织类型、病理分级及治疗措施相似.第一组患者的5年生存率为83.52%,第二组为86.73%,两组差异无显著性.多因素回归分析显示,手术病理期别早、组织病理高中分化、行次广泛或广泛子宫切除术的患者预后好.年龄不是独立的预后因素.结论60岁以上老年子宫内膜癌患者手术病理期别为Ⅲ、Ⅳ期者较多,年龄不是子宫内膜癌患者独立的预后因素,对老年患者仍应采取积极的治疗措施.     

子宫内膜腺鳞癌患者预后的多因素分析

目的 :探讨影响子宫内膜腺鳞癌预后的相关因素 ,并划分高危人群。方法 :收集中山大学肿瘤防治中心 1980年 1月 1日～ 1999年 12月 3 1日收治以手术或手术为主综合治疗的子宫内膜腺鳞癌患者 78例。应用Cox多因素模型分析预后影响因素 ,建立预后指数 (prognosticindex ,PI)方程 ,根据预后指数将患者分为不同的危险组 ,计算不同危险组的期望生存曲线。结果 :患者 5年生存率为 61 1% ,多因素分析结果显示手术病理分期 (FIGO分期 )、病理分级、肿瘤浸润深度是影响预后的主要危险因素 ,P <0 0 5。预后指数PI =1 0 89(FIGO分期 ) 0 970 (病理分级 ) 1 3 67(浸润深度 )。根据PI值将患者划分为低危组、中危组和高危组 ,3组患者的 5年生存率分别为10 0 %、77 74%、0 ,P <0 0 0 1。 3组的 5年生存率两两之间差异有统计学意义 ,P <0 0 5。结论 :手术病理分期、肿瘤病理分级和肿瘤浸润深度是影响子宫内膜腺鳞癌患者预后的主要因素 ;PI值计算简单 ,可根据PI值对患者进行合理化分层 ,为临床个体化治疗和预测预后提供参考。     

Clinical implications and prognostic value of five biomarkers in endometrial carcinoma

Objective

The aim was to identify the relationship between ER, PR, P53, Ki-67, PTEN, the association with clinicopathological parameters and the correlation with survival.

Methods

We studied 190 cases of primary endometrial carcinoma in which ER, PR, Ki-67, P53, PTEN antigens were investigated with the use of immunohistochemical methods. To evaluate the correlations among immunohistochemical staining and the age, menopause status, histological type, FIGO stage, grading, depth of invasion, lymph nodes involvement and serum tumor marker. Survival analysis was assessed within single and combined biomarkers types.

Results

The percentage of Ki-67 and P53 positive endometrial tumors was significantly higher in ER negative vs ER positive tumors (both P = 0.000). The same trend was evident in PR positive and negative group. The percentage of PTEN positive tumors was significantly higher in PR positive versus PR negative tumors (P = 0.021) but was no difference in different ER status. ER and PR status were significant predictors with FIGO staging, grading and recurrence. There was no clear association between PTEN positivity and clinicopathological parameters except more relevance with endometrioid histotype (P = 0.013). Positive Ki-67 or P53 was found to be strictly related to more aggressive features. There was statistically significant difference in different status of P53 and Ki-67 in survival time.

Conclusion

ER and PR positive tumors showed a statistically significant association with better clinical outcome, PR has more significant influence on prognosis. The percentage of positive Ki-67 or P53 was significantly higher in hormone-independent group versus in hormone-dependent group and combined Ki-67 and P53 may have more effect on prognosis in former group.