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Purpose: The efficacy of postoperative adjuvant chemotherapy in patients with stage II colorectal cancer has been a subject of debate. This study aimed to evaluate the prognostic and predictive significance of DNA ploidy and stroma ratio in patients diagnosed with stage II colorectal cancer (CRC). Methods: Clinical data and tumor tissues from 179 patients with stage II CRC were collected retrospectively. DNA ploidy (P) and stroma (S) were assessed using automatic image analysis tools powered by machine learning. Results: Patients were categorized into three risk groups: PS-low (diploid and low stroma, PS-L), PS-intermediate (non-diploid or high stroma, PS-M), and PS-high (non-diploid and high-stroma, PS-H). According to the univariable model, the PS-H group exhibited significantly poorer 5-year overall survival rates at 73.0% compared to 87.8%, with a hazard ratio (HR) of 2.281 (95% CI: 0.946-5.502, P = 0.066), as well as lower 5-year disease-free survival rates at 69.4% versus 86.6%, HR = 2.323 (95% CI: 1.016-5.308, P = 0.046) among stage II colorectal cancer patients. Notably, chemotherapy was associated with improved overall survival [HR = 83.460 (95% CI: 0.179-38925.833), P = 0.003] and disease-free survival [HR = 8.628 (95% CI: 1.059-70.265), P = 0.044] in individuals within the PS-high group. Conclusion: While ploidy and stroma alone do not possess predictive power regarding survival outcomes for stage II colorectal cancer patients, those receiving chemotherapy within the PS-H group demonstrated enhanced survival rates. Therefore, combining assessments of ploidy and stroma may serve as an adjunctive tool in clinical decision-making processes to guide chemotherapy treatment strategies for patients diagnosed with stage II colorectal cancer. [ABSTRACT FROM AUTHOR]

Background: Gastric cancer peritoneal metastasis lacks effective predictive indices. This article retrospectively explored predictive values of DNA ploidy, stroma, and nucleotyping in gastric cancer peritoneal metastasis. Methods: A comprehensive analysis was conducted on specimens obtained from 80 gastric cancer patients who underwent gastric resection at the Department of Gastrointestinal Surgery of Wuhan University Renmin Hospital. Tumor tissues were sectioned and stained. DNA ploidy, stroma, and nucleotyping were quantified using microscopy and digital analysis software. Data analysis was employed by Pearson Chi-square, continuous correction Chi-square, and binary logistic regression. Results: Using both univariate and multivariate analysis, pathological T stage and nucleotyping exhibited a positive correlation with peritoneal metastasis. DNA ploidy and stroma showed a positive correlation in univariate analysis. Chi-square tests demonstrated a positive correlation of DNA ploidy, stroma, and nucleotyping with peritoneal metastasis. The combined application of these three indicators displayed heightened predictive value for peritoneal metastasis. Non-diploid status, high stroma, and chromosomal heterogeneity emerged as positive factors for peritoneal metastasis in gastric cancer. Conclusions: DNA ploidy, stroma, and nucleotyping prove to be predictive factors for peritoneal metastasis, with enhanced predictive efficacy when combined in pairs. [ABSTRACT FROM AUTHOR]

Purpose: This study aimed to investigate the role of DNA ploidy and proliferation index in distinguishing ameloblastoma (AB) from ameloblastic carcinoma (AC). Methods: The study included 29 ACs, 6 conventional ABs that transformed into ACs, and a control cohort of 20 conventional ABs. The demographics and clinicopathologic details of the included cases were summarised and compared. The Ki-67 proliferation index was scored using the QuPath open-source software platform. DNA ploidy analysis was performed using high-resolution flow cytometry. Results: The cohort of ABs presented at an overall younger age compared to both primary and secondary ACs. There was a statistically significant difference between the median duration of the tumour when comparing primary and secondary ACs, with ACs presenting with longer durations than the AB cohort. All cases of AC showed a relatively high median proliferation index of 41.7%, with statistically significant higher scores compared to ABs. DNA ploidy analysis showed that all cases in the AB cohort were diploid. Two diploid cases of AB that transformed into ACs were aneuploid when the corresponding secondary AC was analysed. Fourteen cases of AC were diploid and 12 were aneuploid, with no statistically significant association found between DNA ploidy status of primary and secondary ACs. A statistically significant difference was noted when the DNA ploidy status of ABs was compared to that of ACs. When comparing the Ki-67 proliferation score of ACs to their DNA ploidy status, no statistically significant association was noted. Conclusion: DNA ploidy analysis and proliferation index via Ki-67 IHC are useful ancillary tests that may be used to support a diagnosis of AC and may assist in distinguishing between challenging cases of AB and AC. [ABSTRACT FROM AUTHOR]

Abstract Background The DNA ploidy and the S phase fraction (SPF) are associated with advanced tumors progression. The present study aimed to investigate the clinicopathological behavior of oral squamous cell carcinoma(OSCC) and the prognostication of the DNA ploidy and the SPF on its clinical outcomes. Methods This a prospective longitudinal study of 93 OSCC patients (186 specimens). Using high resolution flowcytometry, DNA analysis and SPF were performed for the primary tumor, field of cancerization and a control normal field. The patients were followed clinically for the development of recurrence or second primary tumor(SPT). Results All the 93 patients are associated with wide field of cancerization. Out of the total number of cases, 28 (30%) patients developed recurrence. Of them 17 (18.3%) had regional recurrence, and eight (8.6%) had local recurrence. Second primary tumors were identified in 14 (15.1%) of the cases. Nodal metastasis, overall recurrence and SPTs were significantly associated with DNA aneuploidy and high SPF. High SPF and DNA hyperploidy showed a 28% increased risk of local recurrence (P.value = 0.032). Conclusion In a series of 93 oral cancers who had wide field of cancerization, 30% of the patients developed recurrence, and 15% developed second primary tumors. The DNA ploidy and the SPF were independent prognosticators for the prediction of OSCC outcomes.

Background: The DNA ploidy and the S phase fraction (SPF) are associated with advanced tumors progression. The present study aimed to investigate the clinicopathological behavior of oral squamous cell carcinoma(OSCC) and the prognostication of the DNA ploidy and the SPF on its clinical outcomes. Methods: This a prospective longitudinal study of 93 OSCC patients (186 specimens). Using high resolution flowcytometry, DNA analysis and SPF were performed for the primary tumor, field of cancerization and a control normal field. The patients were followed clinically for the development of recurrence or second primary tumor(SPT). Results: All the 93 patients are associated with wide field of cancerization. Out of the total number of cases, 28 (30%) patients developed recurrence. Of them 17 (18.3%) had regional recurrence, and eight (8.6%) had local recurrence. Second primary tumors were identified in 14 (15.1%) of the cases. Nodal metastasis, overall recurrence and SPTs were significantly associated with DNA aneuploidy and high SPF. High SPF and DNA hyperploidy showed a 28% increased risk of local recurrence (P.value = 0.032). Conclusion: In a series of 93 oral cancers who had wide field of cancerization, 30% of the patients developed recurrence, and 15% developed second primary tumors. The DNA ploidy and the SPF were independent prognosticators for the prediction of OSCC outcomes. [ABSTRACT FROM AUTHOR]

Introduction: DNA ploidy (P), stroma fraction (S), and nucleotyping (N) collectively known as PSN, have proven prognostic accuracy in stage II colorectal cancer (CRC). However, few studies have reported on the prognostic value of the PSN panel in stage III colon cancer patients receiving capecitabine and oxaliplatin adjuvant chemotherapy. Objectives: This study aimed to validate PSN's prognostic impact on stage III colon cancer, identifying candidates for optimized adjuvant chemotherapy duration. Design: A retrospective analysis was conducted on a cohort of stage III colon cancer patients from April 2008 to June 2020. Methods: Postoperative pathological samples from stage III colon cancer patients who underwent radical surgery and postoperative adjuvant chemotherapy at Sun Yat-sen University Cancer Center were retrospectively collected. Automated digital imaging assessed PSN, categorizing risk groups. Kaplan–Meier, Cox regression, and time-dependent receiver operating characteristic analysis compared model validity. Results: Significant differences in 5-year disease-free survival (DFS) and overall survival (OS) were noted among PSN-based low-, moderate-, and high-risk groups (DFS: 92.10% versus 83.62% versus 79.80%, p = 0.029; OS: 96.69% versus 93.99% versus 90.12%, p = 0.016). PSN emerged as an independent prognostic factor for DFS [hazard ratio (HR) = 1.409, 95% confidence interval (CI): 1.002–1.981, p = 0.049] and OS (HR = 1.720, 95% CI: 1.127–2.624, p = 0.012). The PSN model, incorporating perineural invasion and tumor location, displayed superior area under the curve for 5-year (0.692 versus 0.553, p = 0.020) and 10-year (0.694 versus 0.532, p = 0.006) DFS than TNM stage. In the PSN high-risk group, completing eight cycles of adjuvant chemotherapy significantly improved 5-year DFS and OS compared to four to seven cycles (DFS: 89.43% versus 71.52%, p = 0.026; OS: 96.77% versus 85.46%, p = 0.007). Conclusion: The PSN panel effectively stratifies stage III colon cancer, aiding in optimized adjuvant chemotherapy duration determination. [ABSTRACT FROM AUTHOR]

Purpose: Post prostatectomy PSA kinetics and General Grade Groups (GGG) are the strongest prognostic markers of biochemical recurrence (BCR) and prostate cancer (PCa)-specific mortality after radical prostatectomy. Despite having low-risk PCa, some patients will experience BCR, for some, clinically significant BCR. There is a need for an objective prognostic marker at the time of prostatectomy to improve risk stratification within this population. In this study, we investigated the prognostic potential of DNA ploidy. Materials and Methods: Prostatectomy samples from 97 patients with GGG1 and GGG2 with a low-risk CAPRA-S score were included in this study. PCa tissue with the worst Gleason pattern underwent tissue disaggregation, cell isolation and staining with a DNA stoichiometric stain. Using image cytometry, DNA ploidy was measured and a Ploidy Score (PS) was generated. Results: Among the 97 patients, 79 had no BCR, 18 experienced BCR, of which 14 had a PSA doubling time (PSA-DT) >1 year (low-risk group) and 4 had a PSA-DT of <1 year (high-risk group). Using Logistic regression analysis, only pathological T stage (pT) and PS independently predicted BCR with PS being the most significant (p = 0.001). The number of aneuploid cells was significantly higher in the high-risk group compared to the other groups (p = 1.7x10-11). PS combined with GGG diagnosis further stratified risk groups of biochemical recurrence free survival within CAPRA-S low-risk cohort. Conclusion: DNA ploidy is an independent prognostic marker of BCR in low-risk PCa after radical prostatectomy, which could early on identify potentially aggressive PCa recurrences and introduce a more personalized approach to salvage treatments. [ABSTRACT FROM AUTHOR]

Background: This prospective observational study investigated the determinants of malignant transformation (MT) in localized oral leukoplakia (OL) and proliferative verrucous leukoplakia (PVL). Methods: Demographic, clinical, histological, and DNA ploidy status data were collected at enrolment. Survival analysis was performed (MT being the event of interest). Results: One‐hundred and thirty‐three patients with OL and 20 patients with PVL entered the study over 6 years (mean follow‐up 7.8 years). The presence of OED, DNA ploidy, clinical presentation, and lesion site were associated with MT in patients with OL in a univariate analysis. In a multivariate model, OED was the strongest predictor of MT in patients with OL. Adding DNA ploidy increased the model's predictive power. None of the assessed predictors was associated with MT in patients with PVL. Conclusions: DNA ploidy might identify a subset OL with low risk or minimal risk of MT, but it does not seem to be a reliable predictor in patients with PVL. [ABSTRACT FROM AUTHOR]

Malignant and potentially malignant epithelial lesions are often associated with various abnormalities such as epithelial dysplasia, abnormal DNA content, loss of heterozygosity, and chromosomal number aberrations. Screening and early detection of such abnormalities facilitates proper care and also helps to prevent further progression of potentially malignant lesions to malignancy. In such way, the presence of DNA aneuploidy in oral potentially malignant disorders (OPMDs) may serve as an indicator for the malignant transforming potential. Various assessment methods have been proposed to find the DNA ploidy status of cells. This current systematic review is mainly designed to assess the importance of ploidy status in OPMD while measuring the feasibility of using this biomarker for evaluating the hazard of malignant transformation. As an upshot of this systematic review, we can conclude that use of DNA ploidy status can serve as an independent bio-marker for predicting the malignant transformation of lesions. Furthermore, as a future scope the use of DNA ploidy analysis in normal mucosa of smokers will help to assess the malignancy risk and this technique might also help to predict the genetic predisposition of patients with malignancy.

Malignant and potentially malignant epithelial lesions are often associated with various abnormalities such as epithelial dysplasia, abnormal DNA content, loss of heterozygosity, and chromosomal number aberrations. Screening and early detection of such abnormalities facilitates proper care and also helps to prevent further progression of potentially malignant lesions to malignancy. In such way, the presence of DNA aneuploidy in oral potentially malignant disorders (OPMDs) may serve as an indicator for the malignant transforming potential. Various assessment methods have been proposed to find the DNA ploidy status of cells. This current systematic review is mainly designed to assess the importance of ploidy status in OPMD while measuring the feasibility of using this biomarker for evaluating the hazard of malignant transformation. As an upshot of this systematic review, we can conclude that use of DNA ploidy status can serve as an independent bio-marker for predicting the malignant transformation of lesions. Furthermore, as a future scope the use of DNA ploidy analysis in normal mucosa of smokers will help to assess the malignancy risk and this technique might also help to predict the genetic predisposition of patients with malignancy. [ABSTRACT FROM AUTHOR]

Simple Summary: In this work, we evaluated the prognostic value of the chromatin structure in patients with early-stage lung cancer. We assessed the associations of DNA ploidy, nucleotyping, and tumor–stroma ratio (TSR) with 5-year disease-free survival rates. Clarifying whether patients with homogeneous and heterogeneous chromatin can benefit from adjuvant chemotherapy can guide the decision-making regarding chemotherapy after lung cancer resection, improve the survival rate of patients, and reduce the incidence and cost of adverse events related to lung treatment. (1) Background: Chromatin structure typing has been used for prognostic risk stratification among cancer survivors. This study aimed to ascertain the prognostic values of ploidy, nucleotyping, and tumor–stroma ratio (TSR) in predicting disease progression for patients with early-stage non-small cell lung cancer (NSCLC), and to explore whether patients with different nucleotyping profiles can benefit from adjuvant chemotherapy. (2) Methods: DNA ploidy, nucleotyping, and TSR were measured by chromatin structure typing analysis (Matrix Analyser, Room4, Kent, UK). Cox proportional hazard regression models were used to assess the relationships of DNA ploidy, nucleotyping, and TSR with a 5-year disease-free survival (DFS). (3) Results: among 154 early-stage NSCLC patients, 102 were non-diploid, 40 had chromatin heterogeneity, and 126 had a low stroma fraction, respectively. Univariable analysis suggested that non-diploidy was associated with a significantly lower 5-year DFS rate. After combining DNA ploidy and nucleotyping for risk stratification and adjusting for potential confounders, the DNA ploidy and nucleotyping (PN) high-risk group and PN medium-risk group had a 4- (95% CI: 1.497–8.754) and 3-fold (95% CI: 1.196–6.380) increase in the risk of disease progression or mortality within 5 years of follow-up, respectively, compared to the PN low-risk group. In PN high-risk patients, adjuvant therapy was associated with a significantly improved 5-year DFS (HR = 0.214, 95% CI: 0.048–0.957, p = 0.027). (4) Conclusions: the non-diploid DNA status and the combination of ploidy and nucleotyping can be useful prognostic indicators to predict long-term outcomes in early-stage NSCLC patients. Additionally, NSCLC patients with non-diploidy and chromatin homogenous status may benefit from adjuvant therapy. [ABSTRACT FROM AUTHOR]

Objective To investigate the application value of liquid-based cytology, DNA ploidy and fluorescence in situ hybridization (FISH) in the diagnosis of upper tract urothelial carcinoma (UTUC). Methods From January 2013 to June 2021, 246 cases with suspected UTUC on CT or MRI images in the Army Medical Center of PLA were recruited and retrospectively analyzed. Among them, 190 cases were pathologically diagnosed as UTUC, 43 cases non-tumor lesions, and 13 cases other types of tumors. All the 246 cases were examined with liquid-based cytology of voided urine, 178 cases were also with DNA ploidy analysis and 167 cases with FISH. The diagnostic accuracy of liquid-based cytology, DNA ploidy and FISH alone or in combination was calculated and compared. Results The sensitivities of liquid-based cytology, DNA ploidy and FISH in the diagnosis of UTUC were 44.7%, 82.2%, and 57.9%, respectively; the specificities were 97.7%, 82.4%, and 92.3%, respectively. The diagnostic sensitivities of the above 3 methods for high-grade UTUC were 48.1%, 87.7%, and 60.7%, respectively, and for low-grade UTUC 25.0%, 52.4%, and 37.5%, respectively. When any one of the 3 methods is positive, the diagnostic sensitivity was increased to 93.8% (compared with the 3 methods being used alone, all P < 0.05). When any 2 of them were positive at the same time as the criteria for diagnosing UTUC (both negative or only 1 positive was regarded as negative), the positive predictive value and specificity of this combined diagnostic method were both 100%. Conclusion Urinary liquid-based cytology, DNA ploidy and FISH all are clinically valuable in the diagnosis of UTUC. The combination of the 3 methods can effectively improve the sensitivity or specificity of diagnosis, thereby helping to reduce the risk of missed diagnosis or misdiagnosis.

Abstract 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.

Background: For clinically low-risk stage III colorectal cancer, the decision on cycles of adjuvant chemotherapy after surgery is disputed. The present study investigates the use of additional biomarkers of ploidy and stroma-ratio(PS) to stratify patients with low-risk stage III colorectal cancer, providing a basis for individualized treatment in the future. Methods: This study retrospectively enrolled 198 patients with clinical-low-risk stage III colorectal cancer (T1-3N1M0) and analyzed the DNA ploidy and stroma ratio of FFPE tumor tissues. The patients were divided into PS-low-risk group (Diploidy or Low-stroma) and PS-high-risk group (Non-diploid and High-stroma). For survival analyses, Kaplan–Meier and Cox regression models were used. Results: The results showed that the 5-year DFS of the PS-high-risk group was significantly lower than that in the PS-low-risk group (78.6 vs. 91.2%, HR = 2.606 [95% CI: 1.011–6.717], P = 0.039). Besides, in the PS-low-risk group, the 5 year OS (98.2 vs. 86.7%, P = 0.022; HR = 5.762 [95% CI: 1.281–25.920]) and DFS (95.6, vs 79.9%, P = 0.019; HR = 3.7 [95% CI: 1.24–11.04]) of patients received adjuvant chemotherapy for > 3 months were significantly higher than those received adjuvant chemotherapy for < 3 months. We also found that the PS could stratify the prognosis of patients with dMMR tumors. The 5-year OS (96.3 vs 71.4%, P = 0.037) and DFS (92.6 vs 57.1%, P = 0.015) were higher in the PS-low-risk dMMR patients than those in the PS-high-risk dMMR patients. Conclusion: In this study, we found that PS can predict the prognosis of patients with stage III low-risk CRC. Besides, it may guide the decision on postoperative adjuvant chemotherapy. [ABSTRACT FROM AUTHOR]

Abstract Background Colorectal cancer liver metastases (CRLM) has not been identified as a unified disease entity due to the differences in the severity of metastatic disease and tumor aggressiveness. A screen for specific prognostic risk subgroups is urgently needed. The current study aimed to investigate the prognostic value of DNA ploidy, stroma fraction and nucleotyping of initially resectable liver metastases from patients with CRLM. Methods One hundred thirty-nine consecutive patients with initially resectable CRLM who underwent curative liver resection from 2006 to 2018 at Sun Yat-sen University Cancer Center were selected for analysis. DNA ploidy, stroma fraction and nucleotyping of liver metastases were evaluated using automated digital imaging systems. Recurrence-free survival (RFS) and overall survival (OS) were analyzed using the Kaplan-Meier method and Cox regression models. Results DNA ploidy was identified as an independent prognostic factor for RFS (HR, 2.082; 95% CI 1.053–4.115; P = 0.035) in the multivariate analysis, while stroma-tumor fraction and nucleotyping were not significant prognostic factors. A significant difference in 3-year RFS was observed among the low-, moderate- and high-risk groups stratified by a novel parameter combined with the tumor burden score (TBS) and DNA ploidy (72.5% vs. 63.2% vs. 37.3%, P = 0.007). The high-risk group who received adjuvant chemotherapy had a significantly better 3-year RFS rate than those without adjuvant chemotherapy (46.7% vs. 24.8%; P = 0.034). Conclusions Our study showed that DNA ploidy of liver metastases is an independent prognostic factor for patients with initially resectable CRLM after liver resection. The combination of DNA ploidy and TBS may help to stratify patients into different recurrence risk groups and may guide postoperative treatment among the patients.

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%.

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%), highhyperdiploidy 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%. [ABSTRACT FROM AUTHOR]

The ability to predict malignant transformation in oral potentially malignant disorders would inform targeted treatment, provide prognostic information and allow secondary prevention. DNA ploidy and loss of heterozygosity assays are already in clinical use, and loss of heterozygosity has been used in prospective clinical trials. This review appraises published evidence of predictive ability and explores interpretation of heterogeneous studies, with different diagnostic methods, criteria and intention. Both methods have a sound biological foundation and have predictive value independent of dysplasia grading and clinical parameters. The application of these two techniques cannot be directly compared because of differences in expression of results and application to populations of different risk. Predicting malignant transformation accurately on an individual patient basis is not yet possible with either technique. However, they are valuable applications to stratify patients for inclusion in trials, identify the lowest risk patients and exclude risk when biopsy results are indeterminate for dysplasia. [ABSTRACT FROM AUTHOR]

Background: Colorectal cancer liver metastases (CRLM) has not been identified as a unified disease entity due to the differences in the severity of metastatic disease and tumor aggressiveness. A screen for specific prognostic risk subgroups is urgently needed. The current study aimed to investigate the prognostic value of DNA ploidy, stroma fraction and nucleotyping of initially resectable liver metastases from patients with CRLM. Methods: One hundred thirty-nine consecutive patients with initially resectable CRLM who underwent curative liver resection from 2006 to 2018 at Sun Yat-sen University Cancer Center were selected for analysis. DNA ploidy, stroma fraction and nucleotyping of liver metastases were evaluated using automated digital imaging systems. Recurrence-free survival (RFS) and overall survival (OS) were analyzed using the Kaplan-Meier method and Cox regression models. Results: DNA ploidy was identified as an independent prognostic factor for RFS (HR, 2.082; 95% CI 1.053–4.115; P = 0.035) in the multivariate analysis, while stroma-tumor fraction and nucleotyping were not significant prognostic factors. A significant difference in 3-year RFS was observed among the low-, moderate- and high-risk groups stratified by a novel parameter combined with the tumor burden score (TBS) and DNA ploidy (72.5% vs. 63.2% vs. 37.3%, P = 0.007). The high-risk group who received adjuvant chemotherapy had a significantly better 3-year RFS rate than those without adjuvant chemotherapy (46.7% vs. 24.8%; P = 0.034). Conclusions: Our study showed that DNA ploidy of liver metastases is an independent prognostic factor for patients with initially resectable CRLM after liver resection. The combination of DNA ploidy and TBS may help to stratify patients into different recurrence risk groups and may guide postoperative treatment among the patients. [ABSTRACT FROM AUTHOR]

Background: Methods to analyze progression of carcinogenesis and stage of cancer are vital nowadays due to the high prevalence of these lesions. DNA ploidy analysis is one such important method in early diagnosis and improving prognosis. Aims and Objectives: The aim of this study was to correlate histopathological grading and DNA ploidy in oral leukoplakia, oral submucous fibrosis (OSMF), and oral squamous cell carcinoma (OSCC) cases. Materials and Methods: Our study included 80 subjects, grouped into 4 groups of 20 each of OSCC, leukoplakia, OSMF, and healthy individuals. Histopathological grading was carried out in study cases, DNA ploidy was estimated using flow cytometry, and both the findings were correlated. Results: Among the 20 cases of leukoplakia group, 6 cases showed aneuploidy and 14 showed diploidy. In the 20 cases of OSF group, 2 cases showed aneuploidy and 18 showed diploidy, and in the 20 cases of OSCC group, 10 showed aneuploidy and 10 showed diploidy. Most of the aneuploidy cases showed severe dysplasia. Conclusion: Analysis of DNA ploidy status can serve as a diagnostic tool for early detection of malignancies owing to the subjective nature of traditional histopathological grading.

Tobacco use may initiate the process of oral carcinogenesis with clinically undetectable changes. Smoking cessation may prevent its progression. The objective of this study was to evaluate the association between DNA ploidy and micronucleus (MN) frequency in chronic smokers. Three groups were evaluated: Smoker Group, Former Smoker Group and Control Group. Exfoliative cytology was performed on the lateral border of the tongue and mouth floor. MN and DNA ploidy analyses were performed, as well as the correlation between the variables. The data showed a difference between the groups for the total MN (p = 0.0227), and the Smoker group had the highest mean (4.22 ± 4.12). The three groups did not differ statistically from each other on ploidy evaluation (p-value > 0.05). There was also an association between aneuploidy and increased MN frequency in the Former Smoker group (p = 0.0036). In conclusion, these results point out that there is a relationship between the frequency of MN and aneuploidy in former smokers. Moreover, smoking cessation, even for a short period of time, may promote the decrease of MN frequency caused by tobacco use. [ABSTRACT FROM AUTHOR]

Oral leukoplakia (OL) is the most common oral potentially malignant disorder, with a global prevalence of 2%‐3%, variable malignant transformation rate and incompletely understood aetiology. Considering the subjectivity in oral dysplasia grading, other evaluation methods have been tested as predictors of malignant transformation. DNA ploidy status and loss of heterozygosity signatures have been shown to be good predictive markers of malignant transformation. However, effective markers to predict which lesions will progress to invasive carcinoma and by which mechanisms remain unclear. Recent evidence suggests that dysplasia progression to carcinoma occurs through neutral clonal evolution (i.e. randomly). We focus on the genetic basis of OL, encompassing the gross chromosomal alterations and single‐gene mutations, and discuss such alterations in the context of aetiology, clinical presentation and progression. The deeper we understand the genetic basis of OL, the more we approach a better comprehension of the complex and poorly understood process of oral carcinogenesis. [ABSTRACT FROM AUTHOR]

Purpose: DNA ploidy measurement has earlier been suggested as a potentially powerful prognostic tool in many cancer types, but the role in renal tumors is still unclear. Methods: To clarify its prognostic impact, we analyzed the DNA content of 1320 kidney tumors, including clear cell, papillary and chromophobe renal cell carcinoma (RCC) as well as renal oncocytoma and compared these data with clinico-pathological parameters and patient prognosis. Results: A non-diploid DNA content was seen in 37% of 1276 analyzable renal tumors with a striking predominance in chromophobe carcinoma (74.3% of 70 cases). In clear cell carcinoma, a non-diploid DNA content was significantly linked to high-grade (ISUP, Fuhrman, Thoenes; p < 0.0001 each), advanced tumor stage (p = 0.0011), distant metastasis (p < 0.0001), shortened overall survival (p = 0.0010), and earlier recurrence (p < 0.0001). In papillary carcinoma, an aberrant DNA content was significantly linked to high Fuhrman grade (p = 0.0063), distant metastasis (p = 0.0138), shortened overall survival (p = 0.0010), and earlier recurrence (p = 0.0003). Conclusion: In summary, the results of our study identify a non-diploid DNA content as a predictor of an unfavorable prognosis in clear cell and papillary carcinoma. [ABSTRACT FROM AUTHOR]

Background: Oral potentially malignant disorders are a clinical conundrum as there are no reliable methods to predict their behaviour. We combine conventional oral epithelial dysplasia grading with DNA ploidy analysis to examine the validity of this approach to risk assessment in a cohort of patients with known clinical outcomes. Methods: Sections from diagnostic biopsies were assessed for oral epithelial dysplasia using the WHO grading system, and DNA ploidy analysis was performed using established methods. Patients reviewed for a minimum of 5 years who did not develop oral squamous cell carcinoma were classified as "non‐transforming" cases. Patients that developed oral squamous cell carcinoma ≥ 6 months after the initial diagnostic biopsy were classified as having "malignant transformation." Results: Ninety cases were included in the study. Seventy cases yielded informative DNA ploidy results. Of these 70 cases, 31 progressed to cancer. Oral epithelial dysplasia grading and DNA ploidy status were both significantly associated with clinical outcome (P < 0.05). Severe dysplasia had a hazard ratio of 3.50 (CI: 1.46, 8.45; P = 0.005) compared to cases with mild dysplasia. Aneuploidy had a hazard ratio of 2.09 (CI: 1.01, 4.32; P = 0.046) compared to cases with a diploid/tetraploid status. Receiver operating characteristic analysis gave an area under the curve of 0.617 for DNA ploidy status and 0.688 when DNA ploidy status was combined with dysplasia grading. Conclusion: Our findings suggest that combining dysplasia grading with DNA ploidy status has clinical utility which could be used to develop novel management algorithms. [ABSTRACT FROM AUTHOR]

The combination of DNA ploidy and automatically estimated stroma fraction has been shown to correlate with recurrence and cancer death in colorectal cancer. We aimed to extend this observation and evaluate the prognostic importance of this combined marker in prostate cancer. DNA ploidy status was determined by image cytometry and the stroma fraction was estimated automatically on hematoxylin and eosin stained sections in three tumor samples from each patient to account for tumor heterogeneity. The optimal threshold for low (≤56%) and high (>56%) stroma fraction was identified in a discovery cohort (n = 253). The combined marker was validated in an independent patient cohort (n = 259) with biochemical recurrence as endpoint. The combined marker predicted biochemical recurrence independently in the validation cohort. Multivariable analysis showed that the highest risk of recurrence was observed for patients with samples that had both non‐diploid ploidy status and a high stroma fraction (hazard ratio: 2.51, 95% confidence interval: 1.18–5.34). In conclusion, we suggest the combination of DNA ploidy and automatically estimated stroma fraction as a prognostic marker for the risk stratification of prostate cancer patients. It may also be a potential generic marker as concurrent results have been described in colorectal cancer. What's new? The stroma that surrounds tumor cells can have significant effects on tumor growth and behavior. Another hallmark of cancer is aneuploidy caused by genomic instability, which offers an important prognostic marker in several cancer types. In this study, the authors found that combining these two markers—stromal fraction and DNA ploidy—provided a significant predictor of biochemical recurrence in prostate cancer. The low cost, high throughput, and accuracy of digital analysis using this combined method thus promise to provide improved prognostic biomarkers in prostate cancer. [ABSTRACT FROM AUTHOR]

Background: Methods to analyze progression of carcinogenesis and stage of cancer are vital nowadays due to the high prevalence of these lesions. DNA ploidy analysis is one such important method in early diagnosis and improving prognosis. Aims and Objectives: The aim of this study was to correlate histopathological grading and DNA ploidy in oral leukoplakia, oral submucous fibrosis (OSMF), and oral squamous cell carcinoma (OSCC) cases. Materials and Methods: Our study included 80 subjects, grouped into 4 groups of 20 each of OSCC, leukoplakia, OSMF, and healthy individuals. Histopathological grading was carried out in study cases, DNA ploidy was estimated using flow cytometry, and both the findings were correlated. Results: Among the 20 cases of leukoplakia group, 6 cases showed aneuploidy and 14 showed diploidy. In the 20 cases of OSF group, 2 cases showed aneuploidy and 18 showed diploidy, and in the 20 cases of OSCC group, 10 showed aneuploidy and 10 showed diploidy. Most of the aneuploidy cases showed severe dysplasia. Conclusion: Analysis of DNA ploidy status can serve as a diagnostic tool for early detection of malignancies owing to the subjective nature of traditional histopathological grading. [ABSTRACT FROM AUTHOR]

Potato harbors great amount of diversity in terms of its ploidy. The progress of breeding programme is often limited by the hybridization barriers confronted due to differences in ploidy. Hence, understanding the DNA ploidy is of prime importance. The DNA ploidy can be estimated using flow cytometry. Great choices of nuclear isolation buffers (NIB) are available for sample preparation in flow cytometry. Selecting the best suitable among them is critical and also govern the success of experiment. Here, we tested five modified NIBs to identify the best suitable one for potato sample preparation. Investigation reveals best suitability of modified HPI buffer for flow cytometry in potato. The same buffer was used to estimate the genome size and ploidy of randomly selected potato samples. [ABSTRACT FROM AUTHOR]

Background: It was tested whether the difference in carcinogenesis between noxa and human papillomavirus (HPV)‐driven head and neck squamous cell carcinoma (HNSCC) is associated with a variation in genomic instability. Methods: Conventional and molecular cytogenetics in HPV‐positive and HPV‐negative HNSCC cell lines. Results: Numerical aneuploidy determined by multicolor fluorescence in situ hybridization and DNA ploidy was very similar for both entities with most chromosomes being present either in quadruplicate or triplicate, and only few are still diploid with, however, a striking similarity in the overall pattern. A clear difference was seen concerning the translocations formed, with no difference in the total amount but with a significantly higher genomic instability of HPV‐positive cell lines at chromosome 3 as compared to HPV‐negative cells. Conclusion: The different processes of carcinogenesis of HPV‐positive and HPV‐negative HNSCC appear to result in a similar pattern of numerical but a clear difference in structural chromosomal aberrations. [ABSTRACT FROM AUTHOR]

It is believed that the majority of oral cancers develop from oral potentially malignant lesions (OPML). Though they can be easily detected during screening, risk stratification is difficult. During screening clinicians often find it difficult to distinguish OPMLs from benign lesions, and predicting OPML at risk of malignant transformation is particularly challenging. DNA aneuploidy has been known to be a marker of malignancy in a number of sites including the oral cavity. We performed a systematic review to evaluate the effectiveness of DNA-ICM using brushings in differentiating OPMLs from benign/inflammatory lesions during screening and in predicting malignant transformation. MEDLINE, Pubmed, EMBASE electronic databases were systematically searched using a combination of keywords and subject headings. A total of 11 articles satisfied our inclusion criteria. These studies reported a wide range of sensitivity (16-96.4%) and specificity (90-100%) due to the differences in study design, definitions of high risk or low risk lesions and DNA-ICM protocol used. No long-term longitudinal studies were identified to assess the role of DNA-ICM using brushings in predicting malignant transformation. No studies evaluated the role of DNA-ICM in community screening settings. A number of studies combined DNA-ICM with other techniques like cytology or argyrophilic nucleolar organizer region counts leading to improved test results. In spite of DNA aneuploidy being accepted as a marker of malignancy, there is limited evidence of DNA-ICM using brushings being successful as an adjunct oral cancer screening tool. Longitudinal studies and large community screening studies need to be undertaken to draw stronger conclusion. [ABSTRACT FROM AUTHOR]

Background Chromosomal abnormalities play a key role in neoplastic potential for flat urothelial lesions, irrespective of whether cytologic atypia is present or not. Having information about lesions with malignant potential in the urinary bladder can help clinicians in taking logical actions concerning management after urinary bladder-conserving resections. Aim The aim of the study was to differentiate various flat urothelial lesions by detecting its genetic abnormalities and proliferative activity using UroVysion fluorescence in-situ hybridization and flow cytometry. Patients and methods Ninety-eight paraffin-embedded tissue sections from 35 patients with infiltrative bladder cancer and flat urothelial lesions [reactive atypia, hyperplasia, dysplasia, carcinoma in situ (CIS), squamous metaplasia, glandular metaplasia, histologically normal urothelium in patients with tumor and control] were assessed by flow cytometry and fluorescence in-situ hybridization using UroVysion targeted against the band 9p21 locus and pericentromeric areas of chromosomes 3, 7, and 17. Results Gain of at least one of the chromosomes was observed in 60% of samples being more than 90% in CIS, dysplasia, and infiltrating bladder tumors. The mean percentage of cells with polysomy in reactive atypia, flat urothelial hyperplasia, glandular metaplasia, keratinizing squamous metaplasia, and histologically normal urothelium in patients with tumors ranged from 12 to 20%, but in CIS and dysplasia it was 67 and 44%, respectively. Deletion of the P16 locus was detected in 49% of lesions. Flow cytometry displayed aneuploidy pattern in 60% of lesions with higher percentage in CIS, dysplasia, and infiltrating bladder tumors. High S-phase fraction (SPF) was observed in 44% of samples. High S phase showed significant association with polysomy and DNA ploidy (P =0.02 and 0.01, respectively). In addition, there was association between polysomy expression with del 9p21 (P =0.03) and between del 9p21 with DNA ploidy and SPF (P =0.003 and 0.02). Conclusions Our analysis has shown that combined evaluation of flat urothelial lesions using UroVysion, DNA ploidy, and SPF help in the recognition of these lesions, especially those with higher genetic abnormalities and having higher S phase because follow up of these lesions showed a higher tendency for malignant transformation. Furthermore, estimation of continuing chromosomal instability such as polysomy, DNA aneuploidy, and high SPF might deliver further interpretation of urinary bladder cancer risk assessment away from the position and histopathology of precancerous lesions. [ABSTRACT FROM AUTHOR]

Abstract: Background and Objectives: Quantitation of cell DNA content, DNA ploidy, has been established as a research and prognostic technique for decades. A variety of instruments have been used although only a few commercially available systems have established quality assurance and published outcome data. The aim of this study was to compare two automated systems. Methods: Nuclear monolayers were obtained from 112 oral biopsies by enzyme digestion and Feulgen staining. These were scanned on both the Fairfield and the Ploidy Work Station (PWS) systems. The overall ploidy diagnosis, number of epithelial nuclei, coefficient of variation (CV) and 5c exceeding rate (5CER) were compared by quantile–quantile plots, t test, Wilcoxon and Spearman's tests. Results: The PWS system identified more nuclei (p < 0.0001) at a lower CV (p < 0.0001). Using the PWS system, fewer samples were classified as indeterminate. No difference between 5CER was found between systems (p > 0.54). There was complete concordance between the two systems in terms of DNA ploidy diagnosis. Conclusions: The PWS system is comparable to the Fairfield system for determination of DNA ploidy and has advantages that may lead to improved performance. [ABSTRACT FROM AUTHOR]

Background: Epithelioid hemangioendothelioma (EHE) involving serous effusion is extremely rare, and the diagnosis can be challenging. DNA ploidy quantitation of EHE in effusion fluids has not been previously described in the English-language literature. Methods: Specimens of cytological diagnosed with EHE in effusion fluids between 2002 and 2009 were retrieved from the pathology files at MD Anderson Cancer Center. A total of four cases of EHE involving or arising from effusion fluids were found, and we reviewed cytospin, smears, cell block sections, and immunostained slides. DNA image analysis for ploidy and proliferation evaluation was performed on a destained, papanicolaou-stained slide from each case. Results: The tumor cells were epithelioid with prominent cytoplasmic vacuolization and intracytoplasmic inclusions, which could resemble reactive mesothelial cells, mesothelioma, or adenocarcinoma. The tumor cells were positive for endothelial markers. DNA image analysis in three of the four cases revealed predominantly diploid and tetraploid subpopulations, with few aneuploid cells and fairly low proliferation indices, and these patients had fairly prolonged survival. Conclusions: DNA image analysis is useful for differentiating EHE from reactive mesothelial cells and high-grade carcinoma. For accurate diagnosis of EHE in effusion fluids, cytologic features should be considered together with clinical history and ancillary studies. [ABSTRACT FROM AUTHOR]

Objective: To evaluate the performance of an automated DNA‐image‐cytometry system as a tool to detect cervical carcinoma. Methods: Of 384 liquid‐based cervical cytology samples with available biopsy follow‐up were analyzed by both the Imager System and a high‐risk HPV test (Cobas). Results: The sensitivity and specificity of Imager System for detecting biopsy proven high‐grade squamous intraepithelial lesion (HSIL, cervical intraepithelial neoplasia [CIN]2‐3) and carcinoma were 89.58% and 56.25%, respectively, compared to 97.22% and 23.33% of HPV test but additional HPV 16/18 genotyping increased the specificity to 69.58%. The sensitivity and specificity of the Imager System for predicting HSIL+ (CIN2‐3+) lesions among atypical squamous cells of undetermined significance samples were 80.00% and 70.53%, respectively, compared to 100% and 11.58% of HPV test whilst the HPV 16/18 genotyping increased the specificity to 77.89%. Among atypical squamous cells‐cannot exclude HSIL, the sensitivity and specificity of Imager System for predicting HSIL+ (CIN2‐3+) lesions upon follow up were 82.86% and 33.33%%, respectively, compared to 97.14% and 4.76% of HPV test and the HPV 16/18 genotyping increased the specificity to 19.05%. Among low‐grade squamous intraepithelial lesion cases, the sensitivity and specificity of the Imager System for predicting HSIL+ (CIN2‐3+) lesions were 66.67% and 35.71%%, respectively, compared to 66.67% and 29.76% of HPV test while HPV 16/18 genotyping increased the specificity to 79.76%. The overall results of imager and high‐risk HPV test agreed in 69.43% (268) of all samples. Conclusions: The automated imager system and HPV 16/18 genotyping can enhance the specificity of detecting HSIL+ (CIN2‐3+) lesions. [ABSTRACT FROM AUTHOR]

Background: Oral squamous cell carcinoma (OSCC) is the most frequently occurring malignant tumour in the oral cavity. OSCC arises because of multiple genetic alterations. Cell cycle aberrations and aneuploidy are reportedly among the main characteristics of cancer cells and are associated with aggressive growth and poor prognosis.Methods: The study sample included 47 non-metastasised and 39 metastasised primary OSCC, with matched positive cervical lymph nodes and 17 normal oral mucosa samples. Tissue microarrays (TMAs) were prepared with a minimum of three cores from each case. TMA sections were cut and immunostained with MCM2, Ki-67, geminin and cyclin D1 antibodies. DNA image analysis was performed on the whole tissue section before TMAs were created.Results: The results revealed that there were no differences in cell cycle protein expression in different areas of the tumours or between the metastatic and non-metastatic carcinomas. None of the cell cycle proteins showed significant differences between the lymph node metastasis and the primary OSCC, except for Ki-67. Geminin/Ki-67 ratio showed significant difference between metastatic and non-metastatic tumours. Aneuploidy was detected in all (100%) cases of OSCC. Similarly, all lymph node samples (39 cases) were aneuploid.Conclusion: The results suggest that although there was dysregulation of cell cycle regulatory proteins, only Ki-67 and the MCM2/Ki-67 and geminin/Ki-67 ratios may have prognostic significance in oral cancer. DNA ploidy alone was not specific and may not be a good tool to evaluate prognosis or metastatic progression in oral cavity carcinomas. [ABSTRACT FROM AUTHOR]

Objective We sought to determine whether DNA ploidy correlates with the four molecular subgroups of endometrial carcinoma (EC) as determined using ProMisE (Proactive Molecular Risk Classifier for Endometrial Cancer). Methods 90 cases of EC previously characterized by clinicopathological parameters, outcomes, and ProMisE molecular subgroup ( POLE EDM, MMR-D, p53 wt or p53 abn) were assessed for DNA ploidy using image cytometry. Associations of ploidy with traditional clinicopathological parameters were also tested. Results Abnormal DNA ploidy status differed amongst the ProMisE groups (p < 0.001) and was found in 80.9% (17/21) of p53 abn, 37.0% (10/27) of p53 wt, 28.6% (4/14) of POLE EDM and 14.3% (4/28) of MMR-D. Abnormal DNA content was significantly associated with lower BMI (p = 0.034) and grade 3 tumors (p = 0.001). In the entire cohort, abnormal DNA content was significantly associated with worse progression free survival (p = 0.0094) but not disease specific survival (p = 0.249) or overall survival (p = 0.187). When examining ploidy within each of the ProMisE groups, abnormal DNA content correlated with worse overall survival (p = 0.041) and progression free survival (p = 0.011) in the MMR-D group. No statistically significant relationship was seen in the remaining 3 groups. Conclusion Abnormal DNA ploidy status did correlate with the molecular subgroups of EC; abnormal DNA content was seen in the large majority of p53 abn cases. Abnormal ploidy however was also seen in smaller numbers in the p53 wt, POLE EDM and MMR-D groups; therefore abnormal DNA content was not a specific marker for any one molecular group. The addition of ploidy to the ProMisE molecular categories conferred additional prognostic value within the MMR-D group, which merits further study. [ABSTRACT FROM AUTHOR]

Assessing the nuclear C–DNA content and estimating DNA ploidy in succulent species, such as dragon fruit ( Hylocereus spp., Cactaceae), is a challenging task. Dragon fruit tissues are rich in polysaccharides that are released during the nuclei isolation procedure, inducing the formation of clusters of nuclei and hence leading to inaccurate results. Our flow cytometry protocol for dragon fruit species is simple, rapid and reproducible. Our innovations include: modifications to the nuclear isolation buffer to ‘match’ the cytosol composition of the cells; filtration of the samples instead of centrifugation, thereby preventing co-precipitation of nuclei and polysaccharides; and staining of the nuclei with propidium iodide. DNA ploidy of dragon fruit samples were estimated using a reference standard − either external or internal − with a known ploidy level and a known nuclear DNA content. The best results were obtained using new emerging and actively growing stem tips, which have lower concentrations of polysaccharides and a lower frequency of endopolyploid nuclei, thereby simplifying the nuclei isolation procedure. [ABSTRACT FROM AUTHOR]

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. [ABSTRACT FROM AUTHOR]

Background: The role of DNA ploidy in genomic instability of cancer cells and prognosis has been described in a number of studies. The role of the centrosome in cell cycle has also been reported. Aim: In this study, we aimed to investigate the correlation between the centrosome and DNA ploidy in breast cancer in a search for a cytologic predictive and prognostic marker. Materials and Methods: Cell prints were prepared from cell culture of mesothelial cells, fibroblast cell line MRC5 and breast cancer cell lines MCF7 and T47D. Indirect immunofluorescence was used with anti-γ-tubulin and centrosomes were quantified using a fluorescence microscope. DNA ploidy was scored with the DNA index analyzed by flow cytometry. Results: The normal mesothelial cells (94% of the cells with one detected centrosome) and MRC5 diploid cells (68% with two centrosomes) were used as quality controls. A correlation between the number of centrosomes and DNA ploidy was found in MCF7 cell lines (64% of the cells with a number of centrosomes ≥ 3). It was not observed in invasive breast cancer samples; however, the frequency of cells with centrosomes ≥ 3 was found to be slightly higher in DNA aneuploid samples than in DNA diploid samples (15% vs 13.3%). Conclusion: Quantification of centrosome appears to be correlated to DNA ploidy in breast cancer cell lines and slightly associated to DNA aneuploidy in invasive breast cancer. Studies analyzing a larger number of samples as well as morphological abnormalities of the centrosome are needed.

Nos últimos dez anos, grandes mudanças ocorreram no tratamento do MM com a utilização de novas drogas. Frente a estas novas opções de tratamento é essencial reconhecermos parâmetros clínicos ou biológicos que orientem a melhor escolha terapêutica. Mais recentemente foi validado um novo e simples sistema de estadiamento, International Staging System (ISS), baseado nos valores dabeta2 microglobulina e albumina sérica. Os pacientes são classificados em três grupos de risco: Estádio I: beta2M 3,5 g/dl. Mediana de sobrevida de 62 meses; Estádio II: beta2 M 3,5 - < 5,5 mg/l. Mediana de sobrevida 49 meses; Estádio III: beta2 > 5,5 mg/l. Mediana de sobrevida de 29 meses. Atualmente, a citogenética e achados moleculares estão sendo amplamente reconhecidos como fatores de prognóstico. A deleção do cromossomo 13/13q-, translocação t(4;14), deleção p53 e, mais recentemente, a amplificação da banda cromossômica 1q21 estão associadas a prognóstico reservado.Over the last 10 years, great changes have occurred in the treatment of multiple myeloma (MM) due to the use of new drugs. Considering the new options, it is essential to recognize clinical and biological parameters to arrive at the best therapeutic choice. More recently the new International Staging System (ISS) for multiple myeloma was validated which utilizes two straight forward laboratory parameters: the beta2 microglobulin (beta2M) and albumin levels. Stage I: beta2M < 3.5 mg/L and albumin level > 3.5 g/dL with a median survival of 62 months; stage II: beta2M < 3.5 and albumin < 3.5 g/dL or beta2M > 3.5 to < 5.5 g/dL with a median survival of 49 months; stage III: > 5.5 g/dL with a median survival of 29 months. The importance of cytogenetics and molecular features as prognostic factors is being recognized. Deletion of chromosome 13 or 13q, the t(4:14) translocation, p53 deletion and amplification of chromosome band 1q21 are all associated with poor prognosis.

OBJECTIVE: To study the importance of NB84, synaptophysin and AgNOR and explore the quantitative association of these factors with diagnosis and outcome as well as the association between NB84 and AgNOR and other tumor and stromal factors in twenty-eight peripheral neuroblastic tumors. METHODS: We assessed AgNORs, NB84, synaptophysin and several other markers in tumor tissues from 28 patients with primary neuroblastic tumors. The treatment included: surgery for stage 1, chemotherapy and bone marrow transplantation for most of stages 3 and 4. Histochemistry, immunohistochemistry and morphometry were used to evaluate the amount of tumor staining for AgNOR, NB84 and synaptophysin; the outcome for our study was survival time until death due to recurrent neuroblastic tumors. RESULTS: Only stage (pOBJETIVO: Estudar a importância dos marcadores NB84 e AgNOR e explorar as relações quantitativas entre esses marcadores com o diagnóstico e prognóstico assim como as relações entre NB84 e AgNOR e outros marcadores tumorais e estromais em 28 tumores neuroblásticos periféricos. MÉTODOS: Examinamos AgNOR, NB84 e sinaptofisina e vários outros marcadores em tecidos tumorais de vinte e oito pacientes com tumors neuroblásticos primários. Tratamento dos pacientes incluiu: cirurgia para o estágio 1, quimioterapia e transplante de medula óssea para a maioria dos pacientes nos estágios 3 e 4. Utilizamos histoquímica, imunohistoquímica e morfometria para avaliar a intensidade e extensão de expressão do AgNOR, NB84 e sinaptofisina, tendo o prognóstico dos pacientes incluído o tempo de sobrevida até a morte por recurrência dos tumores neuroblásticos. RESULTADOS: Estadiamento (p

Amifostine is a cytoprotective drug that was initially used to control and treat nuclear radiation injury and is currently used to provide organ protection in cancer patients receiving chemotherapy. Clinical studies have also found that amifostine has some efficacy in the treatment of cytopenia caused by conditions such as myelodysplastic syndrome and immune thrombocytopenia, both of which involve megakaryocyte maturation defects. We hypothesized that amifostine induced the differentiation of megakaryocytes and investigated this by exposing the human Dami megakaryocyte leukemia cell line to amifostine (1 mmol/L). After 12 days of amifostine exposure, optical microscopy showed that the proportion of Dami cells with diameters >20 μm had increased to 24.63%. Transmission electron microscopy identified the development of a platelet demarcation membrane system, while flow cytometry detected increased CD41a expression and decreased CD33 expression on the Dami cell surface. Ploidy analysis found that the number of polyploid cells with >4N DNA content increased to 27.96%. We did not detect any elevation in the mRNA or protein levels of megakaryocytic differentiation-associated transcription factors GATA-binding factor 1 ( GATA-1) and nuclear factor, erythroid 2 ( NF-E2), but nuclear import assay revealed an increased nuclear translocation of these proteins. These findings indicate that amifostine induced the differentiation of Dami cells into mature megakaryocytes via a mechanism involving increased nuclear translocation of the transcription factors, NF-E2 and GATA-1. [ABSTRACT FROM AUTHOR]

Aim: Atypical lymphocytes usually described as lymphocytes with altered shape, increased DNA amount, and larger size. For analysis of cause of genesis and source of atypical lymphocytes during African swine fever virus (ASFV) infection, bone marrow, peripheral blood, and in vitro model were investigated. Materials and Methods: Atypical lymphocytes under the influence of ASFV were studied for morphologic, cytophotometric, and membrane surface marker characteristics and were used in vivo and in vitro models. Results: This study indicated the increased size, high metabolic activity, and the presence of additional DNA amount in atypical lymphocytes caused by ASFV infection. Furthermore, in atypical lymphocytes, nuclear-cytoplasmic ratio usually decreased, compared to normal lymphocytes. In morphology, they looking like lymphocytes transformed into blasts by exposure to mitogens or antigens in vitro. They vary in morphologic detail, but most of them are CD2 positive. Conclusions: Our data suggest that atypical lymphocytes may represent an unusual and specific cellular response to ASFV infection. [ABSTRACT FROM AUTHOR]

Background Carcinoma of the urinary bladder is a significant cause of morbidity and mortality. Development of cancer is a process that takes place over many years, sometimes decades, and can be successfully managed if precancer is detected clinically. Urothelial dysplasia and carcinoma in situ (CIS) are related to recurrence and progression of urothelial carcinoma. Distinguishing CIS and dysplasia from reactive atypia is often difficult on the basis of histological features alone. Because this distinction is therapeutically and prognostically critical, confirming the morphologic impression using immunohistochemistry (IHC) is being increasing practiced in routine clinical practice. Aim We attempted to determine whether an IHC panel would help in this differential diagnosis and the role of DNA ploidy in this aspect. Patients and methods The ploidy and immunoprofile of 21 cases of CIS and 64 non-CIS conditions (14 urothelial biopsies with dysplasia, 13 biopsies with flat hyperplasia, 16 biopsies with reactive atypia, 11 biopsies with normal urothelium from patients with a history of bladder cancer, and 10 normal ureter sections from nephrectomies) were determined using an image analyzer for ploidy and antibodies against CK20, p53, CD44, and ErbB-2 for IHC. Results Although the individual specificity of CK20, p53, and ErbB-2 was high (97, 92, and 91%, respectively), their sensitivity for CIS detection was lower, being 79, 80, and 79%, respectively. Whereas 81% of CIS cases showed positivity for at least two of those three markers, only one case of reactive urothelium showed positivity for two of them. The discriminatory performance of CD44 was good, with a sensitivity and specificity for reactive atypia of 100 and 81%, respectively, versus CIS. The DNA aneuploidy was 71% in CIS, 36% in dysplasia, 6% in reactive atypia, and 18% in the normal urothelium of patients with tumors. Conclusion In conclusion, a panel of CK20, p53, CD44, and ErbB-2 helps to differentiate urothelial CIS from nonneoplastic urothelial atypias. Positive staining for at least two of the three antibodies (CK20, p53, and ErbB-2) is strongly associated with CIS. DNA aneuploidy helps in differentiating CIS from reactive atypia. However, the histologic findings should be a primary determinant in the diagnosis of flat urothelial lesions, with IHC and DNA ploidy playing a supportive confirmatory role. [ABSTRACT FROM AUTHOR]

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Malignant tumors have telomerase activity, which is thought to play a critical role in tumor growth. However, the relation between telomerase activity and genomic DNA status in tumor cells is poorly understood. In the present study, we examined telomerase activity in 13 clear cell type renal cell carcinomas (CRCCs) with similar clinicopathologic features by telomeric repeat amplification protocol assay (TRAP). Based on TRAP assay results, we divided the CRCCs into two groups: a high telomerase activity group and a low/no telomerase activity group. We then analyzed genomic aberration, DNA ploidy, and telomere status in these two groups by comparative genomic hybridization (CGH), laser scanning cytometry (LSC), and telomere-specific fluorescence in situ hybridization (T-FISH), respectively. CGH showed the high telomerase activity group to have fewer genomic changes than the low/no telomerase activity group, which had many genomic aberrations. Moreover, with LSC, DNA diploid cells were found more frequently in the high telomerase activity group than in the low/no telomerase activity group. In addition, T-FISH revealed strong telomere signal intensity in the high telomerase activity group compared with that of the low/no telomerase activity group. These results suggest that telomerase activity is linked to genomic DNA status and that high telomerase activity is associated with genomic stability, DNA ploidy, and telomere length in CRCC.

The present study investigated the predictive and prognostic impact of DNA ploidy together with other well-known prognostic factors in a series of non-endometrioid, high-risk endometrial carcinomas. From a complete consecutive series of 4,543 endometrial carcinomas of International Federation of Gynecology and Obstetrics (FIGO) stages I-IV, 94 serous carcinomas, 48 clear cell carcinomas and 231 carcinosarcomas were selected as a non-endometrioid, high-risk group for further studies regarding prognosis. The impact of DNA ploidy, as assessed by flow cytometry, was of particular focus. The age of the patients, FIGO stage, depth of myometrial infiltration and tumor expression of p53 were also included in the analyses (univariate and multivariate). In the complete series of cases, the recurrence rate was 37%, and the 5-year overall survival rate was 39% with no difference between the three histological subtypes. The primary cure rate (78%) was also similar for all tumor types studied. DNA ploidy was a significant predictive factor (on univariate analysis) for primary tumor cure rate, and a prognostic factor for survival rate (on univariate and multivariate analyses). The predictive and prognostic impact of DNA ploidy was higher in carcinosarcomas than in serous and clear cell carcinomas. In the majority of multivariate analyses, FIGO stage and depth of myometrial infiltration were the most important predictive (tumor recurrence) and prognostic (survival rate) factors. DNA ploidy status is a less important predictive and prognostic factor in non-endometrioid, high-risk endometrial carcinomas than in the common endometrioid carcinomas, in which FIGO and nuclear grade also are highly significant and important factors. [ABSTRACT FROM AUTHOR]

Festulolium is a hybrid between Festuca and Lolium species that has valuable agronomic traits from both grass species. The purpose of our breeding program is to produce hexaploid festulolium that introduces tolerance to summer depression into Italian ryegrass (Lolium multiflorum) by crossing it with tall fescue (Festuca arundinacea). However, we found the DNA ploidy of hexaploids was not stable and was reduced in successive generations. We aimed to find out how to obtain stable high-ploidy festulolium. F1 hybrids of L. multiflorum and F. arundinacea were produced. The F3 generation was produced from putative hexaploid F2 individuals by open pollination. The F4 to F6 generations were obtained by polycrossing. The DNA ploidy levels of F2 to F6 individuals were estimated by flow cytometry. Cytological characteristics of the F5 and F6 individuals were investigated by FISH and GISH. The DNA ploidy level of hexaploid festulolium was reduced and stabilized at almost the same level as a tetraploid. Seed fertility was inversely correlated with an increase in ploidy level. GISH revealed no preferential Lolium transmission. FISH with a telomere probe revealed that counting the exact number of chromosomes in festulolium was difficult. DNA ploidy level was strongly correlated with the number of chromosomes. [ABSTRACT FROM AUTHOR]