Combination of aneuploidy and high S-phase fraction indicates increased risk of relapse in stage I endometrioid endometrial carcinoma.

INTRODUCTION: Endometrioid endometrial carcinoma is a cancer type with generally excellent prognosis when diagnosed at an early stage, but there is a subset of patients with relapsing disease in spite of early diagnosis and surgical treatment. There is a need to find prognostic markers to identify these patients with increased risk of relapse. Depth of myometrial invasion, histological grade, and presence of lymphovascular invasion are known risk factors. DNA content (ploidy) and proliferation measured as S-phase fraction (SPF) have been discussed as prognostic markers but need additional evaluation. MATERIAL AND METHODS: We evaluated relapse-free survival (RFS) with respect to ploidy and SPF, which was analyzed by flow cytometry on fresh tumor tissue, in a cohort of 1001 women treated for stage I endometrioid endometrial carcinoma in northern Sweden during the period of 1993-2010, with a median follow up time of 12.0 years. Data were obtained from historical records. RESULTS: In simple analysis, both aneuploidy and high SPF were associated to increased risk of relapse with hazard ratios (HR) 2.37 (95% CI 1.52-3.70) and 1.94 (95% CI 1.24-3.02), respectively. Our data also confirmed stage, tumor grade, and ploidy as independent prognostic markers in an age adjusted cox regression multivariable analysis but we did not find SPF to contribute to prognosis. However, the combination of aneuploidy and high SPF identified a group of patients with increased risk of relapse, HR 2.02 (95% CI 1.19-3.44). CONCLUSION: In this study, which is the largest study of ploidy and SPF in stage I endometrioid endometrial carcinoma using fresh frozen tissue, aneuploidy was shown to be an independent prognostic marker. Furthermore, the combination of aneuploidy and high SPF could be used to identify patients with increased risk of relapse.

A flow cytometric cell-cycle assay using methyl green.

Methyl green (MG), a conventional, low-cost histological stain, was used to design a flow cytometric cell-cycle/DNA-ploidy assay. On fluorometry, MG absorbed maximally at 633-nm, showed negligible fluorescence in free-state, but emitted brightly when bound to DNA. Optimal dye and cell concentrations for staining and effects of time and photobleaching on stained cells were determined for a lyse-permeabilize-stain protocol. Linearity of DNA-binding, coefficients-of-variation of G0/G1-peaks and minimal carryover were confirmed. Assay results correlated highly with a propidium iodide-based kit in 29 acute lymphoblastic leukemia specimens. The MG-based DNA-ploidy assay represented an accurate and inexpensive alternative to conventional PI-based assays.

Cell proliferation, measured as flow cytometric S-phase fraction, is a strong prognostic indicator in FIGO stage I endometrioid endometrial carcinoma: a population-based study.

INTRODUCTION: In early-stage endometrial carcinoma, there is controversy regarding the prognostic value of the flow cytometric variables DNA ploidy (diploid vs. aneuploid) and S-phase fraction. In Sweden, the former is included in national guidelines despite poor scientific support and the latter is not used clinically. This study investigates the prognostic properties of these variables, together with classical histopathological variables, in multivariate analysis in a stringently stratified material. MATERIAL AND METHODS: Consecutive, population-based patient material restricted to International Federation of Gynecology and Obstetrics (FIGO) 2009 stage I endometrioid endometrial carcinoma (n = 1140) was retrospectively collected from routinely reported data from medical records. Data on age, FIGO stage, degree of differentiation, S-phase fraction, DNA ploidy status, and adjuvant treatment were included in the study. Cumulative incidence curves with other causes of death as a competing risk were used for univariable analysis for the primary endpoint endometrial cancer death. Cox proportional hazards regression analysis was used for multivariate modeling of all endpoints, and for univariable analysis for the secondary endpoints overall survival and time to progression. RESULTS: An S-phase fraction value of >5.5% was associated with worse outcome (for endometrial cancer death: hazard ratio 2.25; 95% CI 1.38-3.67; p = 0.001, adjusted) and DNA ploidy status was not, for all endpoints tested. CONCLUSIONS: In FIGO stage I endometrioid endometrial carcinoma, DNA ploidy status had no prognostic value, whereas the S-phase fraction may be used to identify those with a higher risk of adverse clinical outcome.

Clinical and Analytical Validation of Two Methods for Ki-67 Scoring in Formalin Fixed and Paraffin Embedded Tissue Sections of Early Breast Cancer.

Proliferation determined by Ki-67 immunohistochemistry has been proposed as a useful prognostic and predictive marker in breast cancer. However, the clinical validity of Ki-67 is questionable. In this study, Ki-67 was retrospectively evaluated by three pathologists using two methods: a visual assessment of the entire slide and a quantitative assessment of the tumour margin in 411 early-stage breast cancer patients with a median follow-up of 26.8 years. We found excellent agreement between the three pathologists for both methods. The risk of recurrence for Ki-67 was time-dependent, as the high proliferation group (Ki-67 ≥ 30%) had a higher risk of recurrence initially, but after 4.5 years the risk was higher in the low proliferation group. In estrogen receptor (ER)-positive patients, the intermediate Ki-67 group initially followed the high Ki-67 group, but eventually followed the low Ki-67 group. ER-positive pN0-1 patients with intermediate Ki-67 treated with endocrine therapy alone had a similar outcome to patients treated with chemotherapy. A cut-off value of 20% appeared to be most appropriate for distinguishing between the high and low Ki-67 groups. To summarize, a simple visual whole slide Ki-67 assessment turned out to be a reliable method for clinical decision-making in early breast cancer patients. We confirmed Ki-67 as an important prognostic and predictive biomarker.

Genistein anticancer efficacy during induced oral squamous cell carcinoma: an experimental study.

BACKGROUND: About 7 million people die from various types of cancer every year representing nearly 12.5% of deaths worldwide. This fact raises the demand to develop new, effective anticancer, onco-suppressive, and chemoprotective agents for the future fighting of cancers. Genistein exhibits pleiotropic functions in cancer, metabolism, and inflammation. It functions as an antineoplastic agent through its effect on the cell cycle, apoptotic processes, angiogenesis, invasion, and metastasis. AIM OF THE STUDY: The current study aimed to study the genistein onco-suppressive effects during 7,12-dimethylbenz[a]anthracene (DMBA)-induced oral carcinogenesis in hamsters' buccal pouch utilizing flow cytometry analysis (FMA), as a fast-diagnosing tool, in addition to the histopathology. MATERIAL AND METHODS: The buccal mucosa of adult male Syrian hamsters was painted with paraffin oil only (group 1), DMBA mixed in mineral oil (group 2), or orally administrated genistein along with painting DMBA (group 2B). The buccal mucosa was utilized for flow cytometric analysis and histopathological examination. RESULTS: Grossly, DMBA-induced carcinogenesis started at the 9th week. Progressive signs appeared in the following weeks reaching to large ulcerative oral masses and exophytic nodules at the 21st week. Histologically, invasive well-differentiated oral squamous cell carcinoma (OSCC) appeared in the underlying tissues from the 12th week, showing malignant criteria. Genistein had delayed clinicopathological change, which started 6 weeks later, than the DMBA-painted hamsters, as mild epithelial dysplastic changes. This became moderate during the last 6 weeks, without dysplastic changes. Flow cytometry revealed that DMBA led to considerable variation in DNA proliferation activity, aneuploid DNA pattern, in 47.22% of hamsters and significantly raised the S-phase fragment (SPF) values, which drastically reduced after genistein treatment. CONCLUSION: Taken together, genistein could be employed as an onco-suppressive agent for carcinogenesis. Moreover, FMA could be used as an aiding fast tool for diagnosis of cancer.

Flow Cytometric DNA Ploidy Analysis in Haemato-Lymphoid Neoplasms: An Analysis of 132 Cases.

Background: FxCycleTM Violet (FCV) based flow cytometric (FCM) DNA ploidy analysis is a rapid and simple tool that can substantiate in characterizing the biological behaviour across the spectrum of haematological malignancies and correlates with cytogenetic studies. Materials and Methods: In this prospective study, we performed simultaneous immunophenotyping with FCV based on ploidy analysis in n=132 consecutive new samples, comprising n=110 samples of haemato-lymphoid neoplasms, including acute leukemias (n=67, 60.9%), CML with myeloid blast crisis (n=1, 0.9%), MDS with excess blasts (n=2, 1.8%), mature B cell/ T cell neoplasms (n=37, 33.7%), multiple myeloma (n=3, 2.7%) along with n=22 normal samples. The FCM DNA data was compared with corresponding conventional karyotyping results, wherever available. Results: In FCM ploidy analysis (n=110), the overall DNA index (DI) ranged from 0.81 to 2.17 and S-Phase fraction (SPF) from 0.1-31.6%. Diploidy was seen in n = 90 (81.8%), low-hyperdiploidy in n = 10 (9.1%), high-hyperdiploidy in n = 7 (6.4%) with one case each (0.9% each) having near-tetraploidy, high-hypodiploidy and low-hypodiploidy. The DI of all viable cell populations in normal samples ranged from 0.96-1.05. Conventional karyotyping was performed in n=76/110 cases (70%) with n= 11/76 (15%) culture failures. The modal chromosome number ranged from 45 to 63. A concordance of 95.4% (n=62/65) was noted with corresponding FCM DI. Conclusion: FCV-based ploidy is a sensitive technique that provides complementary information and ascertains a strong correlation with conventional cytogenetics across all haemato-lymphoid neoplasms. It can detect aneuploidy in all B-ALL and myeloma cases, even in hemodiluted samples with cytogenetic culture failure; supplement the diagnoses of erythroleukemia, and provide a useful screen for a higher grade lymph node disease in lymphoma cases with SPF > 3%.

S-phase - an independent prognostic marker in upper tract urothelial carcinoma.

OBJECTIVES: To evaluate S-phase fraction as a predictor of invasiveness and cancer-specific survival in upper tract urothelial carcinoma (UTUC). PATIENTS AND METHODS: One hundred and fifteen patients having undergone radical nephroureterectomy were analysed with histology in radical nephroureterectomy specimens as reference test and S-phase fraction as index test. Ploidy and S-phase were determined using flow cytometry. Differences in S-phase fraction were calculated between stages, grades (WHO 1999 and 2004 classifications), ploidy and patients that died of UTUC and those who did not. Five- and 10-year-cancer-specific survivals were calculated. Areas under the ROC curve (AUCs) of S-phase fraction in relation to tumour stage and to death from UTUC were measured. Multiple Cox regression was performed. RESULTS: Independent prognostic markers of death from UTUC were S-phase fraction and stage. Correlation between S-phase fraction and risk of dying from UTUC was strong, with a 17% greater risk of death from UTUC with every 1% increase in S-phase fraction, hazard ratio = 1.17, 95% CI = 1.10-1.25, p < 0.001, Spearman's rho ρ = 0.65. AUCs for S-phase fraction as predictors of stage and death from UTUC were 0.8 (95% CI = 0.705-0.894) and 0.77 (95% CI = 0.67-0.87), respectively. Cancer-specific survival was statistically significantly different between stages, ploidy and WHO 1999 grades, but not between WHO 2004 grades. This was also reflected in S-phase fraction, which differed in LG-G1 compared with LG-G2 and in HG-G2 compared with HG-G3. CONCLUSION: S-phase fraction was a good test for predicting both invasiveness and cancer-specific survival. Using both WHO 1999 and 2004 classifications, rather than one system alone, had a higher predictive value of cancer-specific survival.

Extranodal presentation of a lymphoma with precursor B-cell phenotype and translocation t(8;14) in South Africa.

INTRODUCTION: A rare entity of a B-cell malignancy with precursor B-cell phenotype and concomitant translocation t(8;14) or variant MYC translocation exists. These cases show clinical, pathological and molecular overlap between precursor B-lymphoblastic leukaemia or lymphoma and Burkitt leukaemia or lymphoma (BLL). CASE PRESENTATION: We report a case from February 2019 at the Charlotte Maxeke Johannesburg Academic Hospital, South Africa, of a 9-month-old infant with a predominantly extracranial soft tissue mass showing extradural extension. There was no involvement of the peripheral blood or bone marrow. Fine needle aspiration and Tru-Cut biopsy of the soft tissue scalp mass showed the tumour to be of precursor B-cell phenotype. Contrastingly, an immunophenotypic assessment revealed a high S-phase fraction and raised concern for BLL. This prompted testing for the translocation t(8;14) by fluorescence in-situ hybridisation analysis, which confirmed this aberration. MANAGEMENT AND OUTCOME: Based on the published experience of other centres, the patient was initiated on a BLL protocol. Despite an excellent clinical response, the patient succumbed to neutropenic sepsis six months after diagnosis. CONCLUSION: Leukaemia or lymphoma with translocation t(8;14) or variant MYC translocation and precursor B-cell phenotype is a rare entity with a varied clinical presentation. This poses a challenge for diagnosis and classification and a clinical dilemma for the choice of treatment.

Prognostic relevance of DNA flow cytometry in breast cancer revisited: The 25-year experience of the Portuguese Institute of Oncology of Lisbon.

The potential prognostic significance of DNA flow cytometric measurements (DNA ploidy and S-phase fraction) in breast cancer remains in dispute. Inconclusive data, primarily due to the lack of consistent standardization and quality control programs, have limited its translation into clinical practice. The aim of the present review, based on the 25-year experience of the Portuguese Institute of Oncology of Lisbon, is to assess the clinical relevance and application of DNA flow cytometry for the prognosis of breast cancer. Overall, data from Portuguese Institute of Oncology of Lisbon indicate that DNA flow cytometry provides significant prognostic information that is biologically relevant and clinically useful for the management of patients with breast cancer. Furthermore, this data has demonstrated the independent value of DNA aneuploidy as a prognostic indicator of poor clinical outcome in various subgroups of patients with early or locally advanced breast cancer at short- and long-term follow-up. Notably, aneuploidy identifies subsets of patients with grade (G)1 or G2 tumours who exhibit a poor clinical outcome. These patients may benefit from adjuvant chemotherapy, particularly those with luminal A and luminal B/human epidermal growth factor-2-negative endocrine-responsive breast cancer. In conclusion, data from Portuguese Institute of Oncology of Lisbon reinforces the clinical importance and utility of DNA flow cytometric analysis, particularly DNA ploidy, in the prognostic assessment and therapeutic planning for patients with breast cancer.

Correlation between histological grading and ploidy status in potentially malignant disorders of the oral mucosa: A flow cytometric analysis.

BACKGROUND: Histopathological grading of oral dysplastic lesions is the method of choice for evaluating malignant and potentially malignant disorders. Owing to inter- and intra-observer variability, determination of the DNA ploidy status of lesions may serve as an adjunct in the prediction of malignant transformation. AIM: To correlate histopathological grading and ploidy status in potentially malignant and malignant disorders of the oral mucosa. SETTINGS AND DESIGN: A pilot study was done with 30 patients (10 patients with oral potentially malignant disorders predominantly leukoplakia, 10 patients with oral malignant lesions and 10 patients with normal mucosa). MATERIALS AND METHODS: Incisional biopsy was done after isolating the biopsy site with 1% Toluidine blue staining. Two sections of the tissue were removed and sent for histopathological and Flow-cytometric analysis respectively. Histopathological diagnosis was obtained and compared with Flow-cytometric results which were graded as diploid and aneuploid. Further, the S - phase fraction, DNA index were also calculated to evaluate the severity of malignant transformation or malignancy. STATISTICAL ANALYSIS: The results were analyzed using Pearson Chi-Square Test. RESULTS: There exists a significant correlation between histopathology and ploidy status in both potentially malignant and malignant group. (P = 0.002). CONCLUSION: The data from this study has shown that DNA Ploidy analysis can be used as a valuable tool in assessing the carcinomatous progression of potentially malignant and malignant lesions.

Prognostic value of proliferation markers: immunohistochemical ki-67 expression and cytometric s-phase fraction of women with breast cancer in libya.

BACKGROUND: We evaluated the association of the immunohistochemical Ki-67 expression, and S-phase fraction with clinicopathological variables and patient outcome. PATIENTS AND METHODS: Histological samples from 100 primary Libyan breast carcinoma patients were retrospectively studied with monoclonal antibody to Ki-67. S-phase fraction was determined by DNA image cytometry. RESULTS: The median Ki-67 percentage for all tumors was 27.5%, ranging from 1 to 80% and the median S-phase fraction (SPF) was 11%, ranging from 0 to 62 %. Tumors with high Ki-67 expression were found in 76% of patients and with high SPF values in 56%. Ki-67 expression was more frequent in tumors with high SPF than low SPF. High Ki-67 and high SPF were associated with advanced stages, poor differentiation of tumors, positive lymph nodes, and distant metastasis. The Ki-67 was associated with hormone receptor negative tumors. The SPF was higher in young patients (<50 years) than in older patients. In the overall population (median follow-up 49 months), patients with high Ki-67 and high SPF had shorter survival time and predicted recurrence than patients with low Ki-67 and low SPF. In a Cox multivariate analysis, high SPF (p= 0.007), hormonal status (p= 0.001) and clinical stage (p=0.005) were independent predictors of disease-specific survival. The Ki-67 (p=0.065) in borderline significance proved to be independent predictor of disease-free survival. The SPF showed more statistically significance with a high grade of malignancy and survival time than Ki-67. CONCLUSIONS: The SPF value is useful cell proliferation marker to assess tumor prognosis. These markers may reflect the aggressive behavior of Libyan breast cancer and predict of the recurrence. It is therefore important to take these markers into consideration to select a high risk subgroup of the patients for intensive treatment.

Diagnostic and prognostic values of S-phase fraction and aneuploidy in patients with bone marrow aplasia.

AIM: It is often difficult and challenging task to differentiate aplastic anemia (AA) from hypoplastic myelodysplastic syndrome (HMDS) among the patients with bone marrow aplasia. This is possibly because of the considerable clinical, cytological and histological similarities between these two disorders. As prognostic and therapeutic approach to AA and HMDS are different, it is imperative to differentiate them at the time of initial diagnosis. Various attempts have been made in the past to differentiate AA from HMDS. In the present study, we explored the value of certain new parameters i.e. S-phase fraction (SPF) and aneuploidy that could be used for this purpose. MATERIALS AND METHODS: The study included 46 consecutive patients with aplastic anemia, 15 patients with HMDS along with 32 age and sex-matched control subjects. S-phase fraction and aneuploidy analysis was carried out by flow cytometry using Mod Fit-LT V3.0 software. RESULTS: The mean SPF value was significantly lower (p=0.02) in patients with AA and higher (p=0.01) in HMDS as compared to that of the control. Aneuploidy was present in 15.2% patients with AA and in 33.3% HMDS cases. During follow-up, 4 patients with AA developed MDS, out of these, three patients had aneuploidy as well as high SPF value at the time of diagnosis. Two patients with HMDS who had aneuploidy and high SPF, converted into AML. Eleven patients died during the study, in whom 8 had aneuploidy and high SPF value. CONCLUSION: We conclude that high SPF value and presence of aneuploidy favour the diagnosis of HMDS rather than AA. SPF and aneuploidy may be important parameters in patients with AA to predict their propensity to evolve into myelodysplastic syndrome and acute myeloid leukemia. SPF value may also be useful in the early diagnosis of HMDS before morphologically evidence of dysplasia is apparent.

Analysis and clinical significance of DNA ploidy, S-phase fraction and proliferating index in colorectal carcinoma / 中国医师进修杂志

ObjectiveTo study the changes and clinical significance of DNA ploidy,S-phase fraction(SPF),proliferating index(PI) in colorectal carcinoma.MethodsIn 40 cases of colorectal carcinoma (colorectal carcinoma group ),40 cases corresponding cancer-adjacent tissue (cancer-adjacent tissue group ),12 cases of precancerous lesions (precancerous lesions group) and 10 cases of normal colorectal mucosa coli (normal colorectal mucosa coli group),DNA ploidy,SPF and PI were detected with flow cytometry and compared.ResultsDNA diploid was 7 cases,DNA heteroploid was 33 cases in colorectal carcinoma group,35,5 cases in cancer-adjacent tissue group,10,2 cases in precancerous lesions group,10,0 case innormal colorectal mucosa coli group,there were significant differences among them(P＜ 0.01 ).SPF and PI in colorectal carcinoma group (35.36 ± 7.45,42.92 ± 6.81 ) were significantly higher than those in cancer-adjacent tissue group (20.82 ±5.51,31.34 ±4.88),precancerous lesions group (21.13 ± 5.07,31.70 ±5.59) and normal colorectal mucosa coli group ( 19.93 ± 3.73,32.01 ± 4.99),there were significant differences among them(P＜ 0.01 ).DNA ploidy was significantly correlated with Dukes staging (P=0.027) and the differentiation of colorectal carcinoma (P =0.030).ConclusionsDNA ploidy,SPF,PI may get some molecular biologycharacteristic and proliferation activity of colorectal carcinoma at molecule level,which may be helpful for treatment and evaluation of prognosis of colorectal carcinoma.

S-phase Fraction as an Independent Prognostic Factor in Invasive Breast Carcinoma -A Study of Long-term Follow-up / 한국유방암학회지

PURPOSE: To evaluate the significance of the S-phase fraction (SPF) and DNA ploidy, determined by DNA flow cytometry, as prognostic markers in invasive breast cancer. METHODS: Between October 1986 and June 1999, 143 breast carcinoma patients, treated by surgery, were analyzed. Flow cytometry was performed for the identification of the SPF and DNA ploidy, with immunohistochemistry performed on paraffin embedded material for the hormone receptor. Two SPF classes were defined on the basis of the median value (10) by using a log rank test (high SPF>10, low SPF<10). The correlation between SPF and the clinicopathological factors (tumor size, lymph node status, histological grade and steroid receptor status) and between the SPF and 5 yr disease-free survival (DFS) were investigated. RESULTS: DNA ploidy was not associated with tumor size, lymph node status, histological grade, overall survival and DFS. In a univariate analysis, high SPF values were associated with shorter 5 yr DFS in individual groups. In the node negative group, the 5 yr DFS of the low SPF group was higher than that of the high SPF group, but in the node positive group, the SPF values showed statistical significance with the 5 yr DFS. In a multivariate analysis, the SPF was independently associated with the 5 yr DFS in the node negative group. CONCLUSION: These results suggested the SPF is an independent prognostic factor in lymph node negative, estrogen receptor positive and progesterone receptor negative breast cancers.

Relationship between Flow Cytometric Nuclear DNA Quantification and Clinicopathologic Parameters in Endometrial Cancer / 대한산부인과학회지

OBJECTIVE: The aim of this study was to investigate the relationship of the DNA ploidy, S-phase fraction to other clinicopathologic factors including age, stage, architecture grade, nuclear grade, lymph node involvement, myometrial invasion in patients with endometrial cancer. METHODS: A prospective analysis was performed on 66 endometrial cancer cases treated at our hospital from Jan. 2000 to Nov. 2004. Of these 66 cases, 41 cases were analyzed by flow cytometry. Fresh tissues for analysis were obtained by dilatation and curettage or surgery as hysterectomy. All specimens for histopathologic grading and stage were classified according to WHO criteria and FIGO stage. DNA ploidy groups were divided into two groups, diploidy and aneuploidy. Fraction more than 6% was classified as high percentage S-phase fraction (SPF). DNA ploidy and SPF were analyzed by flow cytometry in fresh surgical specimens from endometrial cancer. RESULTS: Of the 41 cases, 5cases were aneuploidy, and 16 cases were high percentage SPF. With regard to DNA ploidy and clinicopathologic prognostic factors, aneuploidy was significantly increased as stage, histological type, nuclear grade, architecture grade, and myometrial invasion increased. With regard to DNA ploidy and clinical prognostic factors, aneuploidy was not increased as age, lymph node involvement increased. With regard to SPF and clinicopathologic prognostic factors, high percentage SPF (>6%) was significantly increased as stage, histological type, and nuclear grade increased. With regard to SPF and clinicopathologic prognostic factors, high percentage SPF (>6%) was not increased as age, lymph node involvement, architecture grade, and myometrial invasion increased. CONCLUSION: The DNA ploidy by flow cytometry has shown to have a close relationship to stage, histological type, myometrial invasion, and nuclear and architecture grade. Also, the SPF has shown to have a close relationship to stage, histological type, and nuclear grade. Our results were consistent with the concept that aneuploidy or high percentage SPF could predict the poor prognosis of disease course. The flow cytometric DNA quantification in endometrial cancer may provide major information about tumor prognosis.

The Prognostic Value of the S-phase Fraction and Ki-67 Expression in Early Breast Cancer

PURPOSE: The aim of this study was to compare the S-phase fraction (SPF) and Ki-67 with other biologic factors, and to assess the prognostic value of Ki-67 and SPF in 108 breast cancer patients. MEHOODS: The SPF and Ki-67 level were determined in formalin-fixed, paraffin-embedded tissue specimens from 108 patients with early breast cancer who underwent surgery between January 1997 and December 2000 at the Wonkwang University Hospital. The clinicopathological characteristics of the early breast cancer such as the tumor size, node status, histological grade, hormone receptor, various tumor markers and cancer recurrence were compared with the SPF and Ki-67 values. RESULTS: The median SPF was 9.03% (range 0~43%). The SPF correlated with CD31 (P=0.020) and DNA diploidy (P=0.000). Ki-67 correlated with the histological grade (P= 0.010) and p53 (P=0.035). No correlation was found between the SPF and Ki-67. Eight cases recurred during the follow-up period. Strong expression of Ki-67, p53, DNA aneuploidy and a young age were correlated with recurrence (P=0.001, P=0.029, P=0.021 and P=0.002, respectively). However, the SPF was not related to recurrence. CONCLUSION: In early breast cancer, Ki-67 expression is correlated with the histological grade and p53 expression. In addition, strong Ki-67 expression is associated with a recurrence. Further studies regarding the prognostic significance of the proliferation markers, such as Ki-67 and SPF, will be needed to confirm these results.

Prognostic Value for the S-phase fraction in T1 T2, Node Negative Breast Cancer / 한국유방암학회지

PURPOSE: To evaluate the prognostic value of the S-phase fraction (SPF) and the correlation with other clinicopathologic factors in node negative breast cancer. METHODS: From 1995 to 1998, seventy one breast carcinoma tumors with T1-2N0M0 staging were prospectively sampled as fresh tumors for flow cytometric DNA analysis. We determined the nuclear DNA content, and the SPF was calculated from DNA histograms. We evaluated the relationship between the SPF and other clinicopathologic factors (age, tumor size, tumor grade and, steroid receptor status). The five year distant relapse free survival (DRFS) and overall survival (OS), according to the SPF, were determined. RESULTS: The SPF ranged from 0.1 to 50.9% (median: 13.4%). The SPF was dichotomized using the median value to divide patients into 38 patients (53.5%) having tumors with the low SPF and 33 patients (46.5%) having tumors with the high SPF. The patient's age and the tumor size were not significantly associated with the SPF. High SPF was associated with high tumor grade, but this did not reach statistical significance. There was a significant correlation between high SPF and estrogen receptor negativity; 34.4% of ER positive tumor had high SPF, whereas 58.3% of ER negative tumor had high SPF(p = 0.042). The mean follow up duration was 65.0 months (median: 62.3). Among 71 patients, there were 4 (5.6%) cases of local recurrence, 7 (9.9%) cases of systemic recurrence and 10 (14.1%) cases of disease related death. The patients with high SPF showed a poorer 5 years DRFS and OS than did the patients with low SPF (87.9% vs. 91.2%; 80.4% vs. 94.5%, respectively), but the difference had no statistical significance. CONCLUSION: The present data was insufficient to use SPF information for the selection of the type of adjuvant therapy, but SPF is a promising prognostic factor for node negative breast cancer. Further study with a sufficient number of patients is needed and this should lead to a better understanding of SPF in node negative breast cancer.

Correlation of Proliferating Cell Nuclear Antigen (PCNA) Expression and S-phase Fraction, Survival Rate in Primary Non-Small Cell Lung Cancer / 결핵

BACKGROUND: To study the prognosis of patients with lung cancer, many investigators have reported the methods to detect cell proliferation in tissues including PCNA, thymidine autoradiography, flow cytometry and Ki-67. PCNA, also known as cyclin, is a cell related nuclear protein with 36KD intranuclear polypeptide that is maximally elevated in S phase of proliferating cells. In this study, PCNA was identified by paraffin-embedding tissue using immunohistochemistry which has an advantage of simplicity and maintenance of tissue architecture. The variation of PCNA expression is known to be related with proliferating fraction, histologic type, anatomic(TNM) stage, degree of cell differentiation, S-phase fraction and survival rate. We analyzed the correlation between PCNA expression and S-phase fraction, survival. METHODS: To investigate expression of PCNA in primary lung cancer, we used immunohistochemical stain to paraffin-embedded sections of 57 resected primary non-small cell lung cancer specimen and the results were analyzed according to the cell type, cell differentiation, TNM stage, S-phase fraction and survival. RESULTS: PCNA expression was dMded into five group according to degree of staging(-, +, ++, +++,++++). Squamous cell type showed high positivity than in adenocarcinoma. Nonsignificant difference related to TNM stage was noticed. Nonsignificant difference related to degree of cell differentiation was noticed. S-phase fraction was increased wit advance of PCNA positivity, but t could not reach the statistic significance. The 2 year survival rate and median survival time were -50% 13 months, +75% 41.3 months, ++73% 33.6 months, +++67% 29.0 months, ++++25% 9 months with statistic significance (P<0.05, Kaplan-Meier, generalized Wilcox). CONCLUSION: From this study. PCNA expression was high positive n squamous cell cancer. And, there was no relationship between PCNA positivity and TNM stage, cellular differentiation or S-phase fraction. But, the patients with high positive PCNA staining showed poor survival rate than the patients with lower positive PCNA. It was concluded that PCNA immunostaining is a simple and useful method for survival prediction in paraffin embedded tissue of non-small cell lung cancer.

DNA Ploidy and S-phase Fraction in Breast Cancer

BACKGROUND: Histological differentiation, status of hormonal receptors, size of tumor, and status of axillary lymph nodes are known prognostic factors in breast cancer. Recently, the DNA ploidy and the S-phase fraction have been considered as relating to parameters affecting the prognosis of cancer patients. METHODS: The authors measured the DNA ploidy and the S-phase fraction and evaluated their significance as prognostic factors in breast cancer. Their correlation with tumor size, lymph-node involvement, pathologic differentiation, status of hormonal receptors, menopause, and survival duration were also analyzed. The DNA ploidy and the S-phase fraction were checked with an FAC scan (Becton Dikinson, U.S.A.) and a cell fit (Becton Dikinson, U.S.A.) using the fresh breast-cancer tissue from 104 patients. RESULTS: The ploidy analysis showed that 32% of the tumors were diploid and 68% were aneuploid. There was significance in the pathologic differentiation, but no significance in the other prognostic factors. The median S-phase fraction was 8.9%, and it was used as a cutoff point. Forty-nine percent (49%) of the tumors were greater than 8.9%, and 51% were less than it. There was significance in the status of hormonal receptors. There was no significance in the relation between these factors and the disease-free survival rate. CONCLUSIONS: Even though these results suggest that the DNA ploidy and the S-phase fraction determined by using flow cytometry are not clinically useful independent prognostic factors, it will be necessary to measure them in a large number of patients and then follow those patients so that a multivariate survival analyses can be performed to evaluate the clinical significance of these factors.

Flow Cytometric DNA Content Analysis in Breast Cancer Comparison study of fresh and paraffin-embedded tissues

DNA content of 25 cases of breast carcinoma was analyzed by flow cytometry in both fresh and formalin-fixed, paraffin-embedded tissue. Aneuploidy in fresh tissue and paraffin-embedded tissues was 72% and 32%, respectively. There was a 52% agreement in analysis of DNA ploidy between fresh and paraffin-embedded tissues. Most of the discrepancies resulted from loss of aneuploid peaks on the histograms of paraffin-embedded tissue. Mean S-phase fraction was slightly higher in a paraffin-embedded tissue than that in the fresh tissue; 19.2 9.1% versus 16.1 8.8% and there was no significant correlation between the S-phase fractions. In statistical analysis, the histologic and nuclear grades were not correlated with ploidy or mean S-phase fraction. Therefore it is strongly recommended to use the fresh tissue in flow cytometric DNA content analysis of breast cancer.