High Ki-67 Score Research Articles

AOSOP15 PHASE 2 STUDY COMPARING CISPLATIN/ETOPOSIDE AND WEEKLY PACLITAXEL/CARBOPLATIN REGIMENS WITH CONCURRENT THORACIC RADIOTHERAPY IN PATIENTS WITH LOCALLY ADVANCED NON-SMALL-CELL LUNG CANCER

Introduction. Matrix-producing carcinoma (MPC) of the breast is a rare subtype of breast carcinoma. As a rare entity, clinicopathological features and its response to therapy are still unclear. To better understand the biological features of MPC, we investigated its immunohistochemical expression profiles. Methods. We undertook immunohistochemical study for oestrogen receptor (ER),progesterone receptor (PR),HER2,Ki67, cytokeratin (CK) 5, CK14, epidermal growth factor receptor (EGFR), androgen receptor (AR), p53, c-MET, and laminin on 11 cases of MPC. Results. All cases were triple negative breast cancer. Six cases showed a high Ki67 score (>50%). Most cases were immunoreactive for CK5 (10/11), CK14 (9/11), and EGFR (10/11). Three cases were positive for AR. Nine cases were positive for p53. Positive immunoexpression of c-MET was observed in one case, and of laminin in ten cases. The one metastatic lymph node showed the same matrix-producing pattern but expressed CK5, EGFR, and Ki67 differently from the primary site. Discussion. These data suggest that MPC has the immunohistochemical features of basal-like breast carcinoma, and these expression profiles might be helpful in biomarker selection for appropriate targeted therapies. Funding. None declared. The authors declared no conflicts of interest.

Abstract PD01-04: Deep kinome sequencing identifies a novel D189Y mutation in the Src family kinase LYN as a possible mediator of antiestrogen resistance in ER+ breast cancer

Abstract Estrogen receptor-positive (ER+) breast tumors adapt to hormone deprivation and acquire resistance to aromatase inhibitors (AIs). The proliferative rate of tumor cells measured by Ki67 after short-term treatment with an AI has been proposed as a surrogate endpoint of long term outcome. The purpose of this study was to identify kinase mutations associated with resistance to endocrine therapy as identified by a high Ki67 score after two weeks of pre-surgical therapy with letrozole. We performed deep-kinome sequencing on 4 ER+/HER2– breast tumors that retained high Ki67 scores (14.8 to 24.5%) following short-term letrozole treatment. Genomic DNA from the surgically excised tumors was deep sequenced using a capture approach with 120-bp biotinylated oligonucleotides hybridizing to 612 genes, including 517 kinases with ≥300x coverage. All 4 tumors contained a ‘hot spot’ mutation in PIK3CA, the gene encoding the p110α catalytic subunit of PI3K. One tumor also contained a novel D189Y somatic mutation in LYN, a member of the Src family kinases (SFKs; variant frequency of 8%). D189Y (565G>T) is located in the LYN SH2 domain. Reverse-phase protein array (RPPA) analysis available in 10 tumors in this study revealed a significant correlation (p = 0.006) between Y416 P-Src (which detects all SFKs) and a high post-letrozole Ki67 score. A siRNA screen targeting 779 kinases identified LYN as one of the top hits whose knockdown significantly reduced growth of MCF-7 breast cancer cells with acquired resistance to estrogen deprivation. We next analyzed the Cancer Genome Atlas (TCGA) SNP data to detect copy number changes for 444 tumors where corresponding gene expression data were available. Copy number increases (>0.8 log2 ratio over normal matched DNA) in LYN were present in approximately 10% of breast cancers, with the highest copy number gains observed in luminal B tumors. In contrast, other SFKs showed less frequent copy number increases (YES1 <1%, SRC 2%, FYN 1%). Analysis of the 550 breast cancers in the TCGA identified four additional LYN mutations (E159K, K188N, G418R, A503D), with the first two located in the SH2 domain. Finally, we investigated the role of D189Y LYN in endocrine-resistant breast cancer using ER+ MCF-7 cells transduced with GFP (control), wild-type (WT) LYN, or D189Y LYN vectors. Overexpression of WT or mutant LYN resulted in increased phosphorylation of Src (at Y416), IGF-IR, EGFR, STAT3, AKT, and MAPK. D189Y LYN overexpression also induced phosphorylation of IRS-1. Although stable transduction of either WT or D189Y LYN accelerated MCF-7 cell growth in estrogen-depleted medium, the mutant was more potent than WT LYN at inducing this effect. Further, D189Y LYN overexpression rendered treatment with the ER downregulator fulvestrant or the PI3K inhibitor BKM120 less effective. These results suggest, first, that LYN may play a role in escape from estrogen deprivation in a subset of ER+ breast cancers. Second, ER+ breast cancers harbor multiple molecular alterations capable of mediating hormone-independent growth. Citation Information: Cancer Res 2012;72(24 Suppl):Abstract nr PD01-04.

Abstract S3-6: Profiling of triple-negative breast cancers after neoadjuvant chemotherapy identifies targetable molecular alterations in the treatment-refractory residual disease

Abstract Background: Neoadjuvant chemotherapy (NAC) is increasingly used in patients with triple-negative breast cancer (TNBC). NAC can induce a pathologic complete response (pCR) in ∼30% of patients which portends a favorable prognosis. In contrast, patients with residual disease (RD) in the breast at surgical resection exhibit worse outcomes. Objective: We hypothesized that profiling residual TNBCs after NAC would identify molecularly targetable lesions in the chemotherapy-resistant component of the tumor and that the persistent tumor cells would mirror micro-metastases which ultimately recur in such patients. Methods: We utilized targeted next generation sequencing (NGS) for 182 oncogenes and tumor suppressors in a CLIA certified lab (Foundation Medicine, Cambridge, MA) and gene expression profiling (NanoString) of the RD after NAC in 102 patients with TNBC. The RD was stained for Ki67, which has been reported to predict outcome after NAC in unselected breast cancers. Results: Thirteen tumors were not evaluable due to low tumor cellularity. Of 89 evaluable post-NAC tumors, 57 (64%) were basal-like; 19% HER2-enriched; 6% luminal A; 6% luminal B and 5% normal-like. Mean depth of coverage was 635 (range: 135–1207). Of 81 tumors evaluated by NGS, 72/81 (89%) demonstrated mutations in TP53, 22 were MCL1-amplified (27%), and 17 were MYC-amplified (21%). Alterations in the PI3K/mTOR pathway (AKT1-3, PIK3CA, PIK3R1, RAPTOR, PTEN, and TSC1) were identified in 27 tumors (33%). Cell cycle genes were altered in 25 tumors (31%), including amplifications of CDK2, CDK4, and CDK6, CCND1-3, and CCNE1 as well as RB loss. Alterations in the DNA repair pathway (BRCA1/2, ATM; 16 tumors; 20%) and the Ras/MAPK pathway (KRAS, RAF1, NF1; 10 tumors; 12%) were also common. Sporadic growth factor receptor amplifications occurred in EGFR, KIT, PDGFRA, PDGFRB, MET, FGFR1, FGFR2, and IGF1R. NGS identified 7 patients with ERBB2 gene amplification in the RD which was confirmed by FISH in both the pre- and post-treatment tissue, suggesting NGS could assist in the identification of ERBB2-overexpressing tumors misclassified at the time of diagnosis. In general, the gene amplifications identified by NGS corresponded to enhanced gene expression levels. Amplifications of MYC were independently associated with poor recurrence-free survival (RFS) and overall survival (OS). An interaction effect on survival was observed between MEK activation (assayed by a gene expression signature) and MYC amplification, suggesting cooperation between these pathways. Alterations in DNA repair also identified a subgroup with poor RFS and OS. In contrast, high post-NAC Ki67 score did not predict poor RFS or OS in this predominantly TNBC cohort. Conclusions: The diversity of lesions in residual TNBCs after NAC underscores the need for powerful and broad molecular approaches to identify actionable molecular alterations and, in turn, better inform personalized therapy of this aggressive disease. Incorporation of this platform into clinical studies and eventually standards of care should aid in the prioritization of patients with RD after NAC into rational adjuvant studies. Citation Information: Cancer Res 2012;72(24 Suppl):Abstract nr S3-6.

Profiling of residual breast cancers after neoadjuvant chemotherapy identifies DUSP4 deficiency as a mechanism of drug resistance

Neoadjuvant chemotherapy (NAC) induces a pathological complete response (pCR) in ~30% of patients with breast cancer. However, many patients have residual cancer after chemotherapy, which correlates with a higher risk of metastatic recurrence and poorer outcome than those who achieve a pCR. We hypothesized that molecular profiling of tumors after NAC would identify genes associated with drug resistance. Digital transcript counting was used to profile surgically resected breast cancers after NAC. Low concentrations of dual specificity protein phosphatase 4 (DUSP4), an ERK phosphatase, correlated with high post-NAC tumor cell proliferation and with basal-like breast cancer (BLBC) status. BLBC had higher DUSP4 promoter methylation and gene expression patterns of Ras-ERK pathway activation relative to other breast cancer subtypes. DUSP4 overexpression increased chemotherapy-induced apoptosis, whereas DUSP4 depletion dampened the response to chemotherapy. Reduced DUSP4 expression in primary tumors after NAC was associated with treatment-refractory high Ki-67 scores and shorter recurrence-free survival. Finally, inhibition of mitogen-activated protein kinase kinase (MEK) synergized with docetaxel treatment in BLBC xenografts. Thus, DUSP4 downregulation activates the Ras-ERK pathway in BLBC, resulting in an attenuated response to anti-cancer chemotherapy.

HER2 and TOP2A in high-risk early breast cancer patients treated with adjuvant epirubicin-based dose-dense sequential chemotherapy

BackgroundHER2 and TOP2A parameters (gene status, mRNA and protein expression) have individually been associated with the outcome of patients treated with anthracyclines. The aim of this study was to comprehensively evaluate the prognostic/predictive significance of the above parameters in early, high-risk breast cancer patients treated with epirubicin-based, dose-dense sequential adjuvant chemotherapy.MethodsIn a series of 352 breast carcinoma tissues from patients that had been post-operatively treated with epirubicin-CMF with or without paclitaxel, we assessed HER2 and TOP2A gene status (chromogenic in situ hybridization), mRNA expression (quantitative reverse transcription PCR), as well as HER2 and TopoIIa protein expression (immunohistochemistry).ResultsHER2 and TOP2A amplification did not share the same effects on their downstream molecules, with consistent patterns observed in HER2 mRNA and protein expression according to HER2 amplification (all parameters strongly inter-related, p values < 0.001), but inconsistent patterns in the case of TOP2A. TOP2A gene amplification (7% of all cases) was not related to TOP2A mRNA and TopoIIa protein expression, while TOP2A mRNA and TopoIIa protein were strongly related to each other (p < 0.001). Hence, TOP2A amplified tumors did not correspond to tumors with high TOP2A mRNA or TopoIIa protein expression, while the latter were characterized by high Ki67 scores (p = 0.003 and p < 0.001, respectively). Multivariate analysis adjusted for nodal involvement, hormone receptor status, Ki67 score and HER2/TOP2A parameters revealed HER2/TOP2A co-amplification (21.2% of HER2 amplified tumors) as an independent favorable prognostic factor for DFS (HR = 0.13, 95% CI: 0.02-0.96, p = 0.046); in contrast, increased HER2/TOP2A mRNA co-expression was identified as an independent adverse prognostic factor for both DFS (HR = 2.41, 95% CI: 1.31-4.42, p = 0.005) and OS (HR = 2.83, 95% CI: 1.42-5.63, p = 0.003), while high TOP2A mRNA expression was an independent adverse prognostic factor for OS (HR = 2.06, 95% CI: 1.23-3.46, p = 0.006). None of the parameters tested was associated with response to paclitaxel.ConclusionsThis study confirms the favorable prognostic value of HER2/TOP2A co-amplification and the adverse prognostic value of high TOP2A mRNA expression extending it to the adjuvant treatment setting in early high-risk breast cancer. The strong adverse prognostic impact of high HER2/TOP2A mRNA co-expression needs further validation in studies designed to evaluate markers predictive for anthracyclines.Trial registrationAustralian New Zealand Clinical Trials Registry ACTRN12611000506998.

The relationship between RACK1 expression and clinicopathologic information of esophageal squamous cell carcinoma

To investigate the expression and relationship of receptor for activated C kinase 1 (RACK1) and clinical characteristics in esophageal squamous cell carcinoma (ESCC). Western Blotting was conducted to detect the RACK1 expression in ESCC cell lines. Immunohistochemistry was performed to assay the expression of RACK1 and Ki67 in tumor tissues and adjacent normal epithelium from 113 ESCC patients in tissue microarray. The relationship between the RACK1 level and such clinicopathologic profiles as age, gender, location, smoking, differentiation degree and TNM (tumor, node, metastasis) stage were analyzed. The expression of RACK1 protein was significantly down-regulated in ESCC tissues as compared with the normal adjacent epithelium (χ(2) = 63.363, P < 0.01). An upregulated expression of RACK1 was observed in 72.5% (29/40) ESCC tissues of patients without a smoking history. And it was significantly higher than that in 46.6% (34/73) of patients with a smoking history (χ(2) = 7.040, P = 0.008). In addition, the rate of up-regulated of RACK1 was significantly higher in stage I and II group (63.8%, 44/69) than that in stage III group (43.2%, 19/44) (χ(2) = 4.616, P = 0.032). Moreover, the ESCC tissues with a higher Ki67 score showed a lower level of RACK1 than that with a lower Ki67 score (χ(2) = 8.261, P = 0.016). The expression of RACK1 is down-regulated in ESCC tissues and associated with smoking. The expression of RACK1 was associated with smoking, TNM staging and Ki67 score of ESCC.

Abstract P5-01-10: Tumor Optic Properties Measured Using Diffuse Optic Spectroscopy Imaging Correlate with Proliferation and Glucose Metabolism in Breast Cancer Patients

Abstract (Background) Diffuse Optic Spectroscopy Imaging (DOSI) is non-invasive imaging technology that employs near-infrared (NIR) light to quantitatively characterize the hemodynamic and metabolic properties of breast cancer tumors. (Methods) We utilized DOSI to measure baseline tumor concentrations of oxy-hemoglobin (ctO2Hb), deoxy-hemoglobin (ctHHb), total hemoglobin (ctTHb), oxygen saturation (stO2), as well as water and lipid content of tumors from sixteen patients with primary breast cancer prior to neoadjuvant chemotherapy. Core-needle biopsy specimens were also collected from these patients and analyzed for Ki67, Glut-1, and Fatty acid synthese (FAS), biomarkers of cancer proliferation, glucose metabolism and fatty acid metabolism, respectively. These optic and biological biomarkers were statistically compared to each other and to overall therapy response. (Results) Ki67 score was positively correlated with baseline levels of ctO2Hb (µM) (r = 0.51, p = 0.04), ctTHb (µM) (r = 0.51, p = 0.05), and stO2 (%) (r = 0.57, p = 0.02), and negatively correlated with lipid content (%) (r = -0.52, p = 0.04). Tumors with positive Glut-1 status (n=8) showed significantly higher baseline levels of ctO2Hb (26.2±12.5 v.s 19.9±10.1, p = 0.04), ctTHb (41±13.3 v.s 27.5±11.7, p = 0.05), and stO2 (81.1±6.6 v.s 70.2±9.9, p = 0.02) and lower baseline levels of lipid (49±17.1 v.s 66.2±10.4, p = 0.03) compared to those with negative Glut-1 status (n = 8). There were no correlation between FAS and the optic properties. Five (31.3%) of 16 tumors achieved pathologic complete response (pCR) after completion of chemotherapy followed by surgery. Tumors with pCR showed higher stO2, higher Ki67 score and higher likelihood of Glut-1 expression than those with non-pCR (p = 0.009, 0.01, 0.03, respectively). Only p-values with statistical significance were described. (Conclusion) Higher tumoral expression levels of Ki67 and Glut-1 were correlated with higher oxygenation and lower lipid concentration and associated with a pathologic complete response to chemotherapy. Non-invasive optic properties measured using DOSI are potential surrogate markers for tumor proliferation and glucose metabolism. Citation Information: Cancer Res 2010;70(24 Suppl):Abstract nr P5-01-10.

Exploring the peritoneal surface malignancy phenotype—a pilot immunohistochemical study of human pseudomyxoma peritonei and derived animal models

Peritoneal surface malignancies are characterized by the propensity for tumor growth on peritoneal surfaces without development of extraperitoneal metastases, but the molecular basis for this phenomenon is incompletely understood. Five human tumors and corresponding orthotopic animal models of human pseudomyxoma peritonei and peritoneal mucinous carcinomatosis from colorectal carcinoma were extensively characterized by immunohistochemical analysis of molecular markers of tissue differentiation (carcinoembryonal antigen, CK20, CK7, and vimentin), proliferation and metastasis (Ki-67, vascular endothelial growth factor, and S100A4), mucins (MUC1, MUC2, MUC4, MUC5AC), and adhesion molecules (E-cadherin, N-cadherin, P-cadherin, claudin 1, claudin 3, and claudin 4). Macro- and microscopic growth patterns of implanted human tissues were preserved through passages in the animals, as were with few exception immunohistochemical staining profiles, supporting the relevance of the models as tools for studying the human disease. Tissue differentiation marker expression was in accordance with previously published results and high Ki-67 score confirmed high proliferative capacity, whereas absence of metastatic capacity was supported by low expression levels of the studied metastasis markers. These mucinous tumors expressed high levels of MUC2 and MUC4, whereas MUC1 was not expressed and MUC5AC expression was variable. Similarly, specific adhesion molecules from the cadherin and claudin families were shown to be of relevance in the investigated samples. The results indicate that mucinous peritoneal surface malignancies of intestinal origin are characterized by the presence of specific molecular markers and represent a step toward understanding the complexity of this intriguing phenotypic entity.

Predictors of Tumor Progression During Neoadjuvant Chemotherapy in Breast Cancer

PURPOSE Although most breast cancer patients who receive neoadjuvant chemotherapy (NCT) have a tumor response, a small proportion experience progressive disease (PD). Predictors of response have been reported, but predictors for progression have not been identified. We sought to identify predictors of tumor progression during NCT with the ultimate aim of identifying patients who might benefit from a first-line surgical approach or from novel targeted therapies. PATIENTS AND METHODS Data were obtained from reviewing medical records of patients with stage I to III breast cancer who received NCT (anthracycline and/or taxane based). Statistical analysis was performed to compare patients with any response or stable disease with patients with PD. RESULTS One thousand nine hundred twenty-eight patients received NCT; 1,762 patients (91%) had some response, 107 (6%) had stable disease, and 59 (3%) had PD at some point during NCT. Factors predictive of PD included African American race (P = .002), tumor (T) status (P = .002), and American Joint Committee on Cancer clinical stage (P = .02). Histopathologic features of PD were high tumor grade (P = .005), high Ki-67 score (P = .002), and negative estrogen receptor (ER)/progesterone receptor (PR) status (P < .001/P < .001). Pre-NCT T status, race, and ER status were independent predictors of progression in multivariate analysis. Disease progression was a negative predictor of distant disease-free survival and overall survival in multivariate analysis (P < .001). CONCLUSION Factors predictive of PD include race, advanced tumor stage, high nuclear grade, high Ki-67 score, and ER/PR negativity. Because many of these variables are also associated with response to NCT, novel molecular predictors are needed to identify patients at risk for progression on standard NCT.

Analysis of immunohistochemical predictors of outcome in pseudomyxoma peritonei

22067 Background: PMP is a rare malignant process presenting with a broad range of biological behavior, varying from the indolent course of disseminated peritoneal adenomucinosis (DPAM) to the aggressive peritoneal mucinous carcinomatosis (PMCA). However, there is overlap in the tumor biology not captured by this histological classification, and better prognostic tools need to be identified. In this study we investigated the value of immunohistochemical markers (IHC) in predicting histological subtypes and mortality in PMP patients. Methods: We retrospectively reviewed the charts of 65 patients treated between October 1, 2002 and September 31, 2006. All patients were treated with aggressive cytoreduction and IPHC with mitomycin C. Tumors were categorized into DPAM, PMCA and PMCA-I, an intermediate subtype. IHC expression of Cox-II, Her-2, EGFR, MUC-2, and Ki-67 as well as ploidy status were determined. Using Chi-Square test we determined the predictive value of these markers for PMP histological classification, while impact on survival was investigated using Kaplan-Meier survival analysis (KM). Results: Of the 65 PMP patients, the histological classification was 41% DPAM, 34% PMCA-I and 25% PMCA. With a median follow up of 16 months, patients with DPAM had a lower mortality rate of 4% compared to 17.4% PMCA-I and 25% PMCA (Table 1). The expression of EGFR, aneuploidy, and a Ki-67 score > 50% correlated progressively with the more aggressive histological subtypes (Table1). Fifteen percent of patients (n=10) died due to progressive disease. Mortality was associated with Ki-67 (p=0.03) and aneuploidy (p=0.05), but not with histological classifications evident on survival analysis. Conclusion: Expression of EGFR, high Ki-67 score, and aneuploidy appear to predict for the more aggressive histological variants (PMCA/PMCA-I). Additionally, high Ki-67 score and aneuploidy were associated with mortality independent of PMP subtype, and may represent a more sensitive tool for predicting outcome. Table 1 DPAM (N = 27) PMCA-I (N = 22) PMCA (N = 16) p-value Median Age 46 46 46 0.97 Mortality 2 (7%) 4 (18.2%) 4 (25%) 0.27 COX-II 10 (37%) 8 (36.4%) 7 (43.7%) 0.94 EGFR 10 (37%) 17 (77%) 13 (81.2%) 0.004 HER2 16 (59%) 12 (54%) 10 (66%) 0.56 Ki-67 > 50% 1 (4%) 5 (22%) 6 (37%) 0.01 Aneuploidy 1 (4%) 7 (32%) 6 (37%) 0.01 No significant financial relationships to disclose.

Lymph vascular invasion in invasive mammary carcinomas identified by the endothelial lymphatic marker D2-40 is associated with other indicators of poor prognosis

BackgroundImmunohistochemical studies of lymphatic vessels have been limited by a lack of specific markers. Recently, the novel D2-40 antibody, which selectively marks endothelium of lymphatic vessels, was released. The aim of our study is to compare lymphatic and blood vessel invasion detected by hematoxylin and eosin (H&E) versus that detected by immunohistochemistry, relating them with morphologic and molecular prognostic factors.MethodsWe selected 123 cases of invasive mammary carcinomas stratified into three subgroups according to axillary lymph node status: macrometastases, micrometastases, and lymph node negative. Lymphatic (LVI) and blood (BVI) vessel invasion were evaluated by H&E and immunohistochemistry using the D2-40 and CD31 antibodies, and related to histologic tumor type and grade, estrogen and progesterone receptors, E-cadherin, Ki67, p53, and Her2/neu expression.ResultsLVI was detected in H&E-stained sections in 17/123 cases (13.8%), and in D2-40 sections in 35/123 cases (28.5%) (Kappa = 0.433). BVI was detected in H&E-stained sections in 5/123 cases (4.1%), and in CD31 stained sections in 19/123 cases (15.4%) (Kappa = 0.198). LVI is positively related to higher histologic grade (p = 0.013), higher Ki67 expression (p = 0.00013), and to the presence of macrometastases (p = 0.002), and inversely related to estrogen (p = 0.0016) and progesterone (p = 0.00017) receptors expression.ConclusionD2-40 is a reliable marker of lymphatic vessels and is a useful tool for lymphatic emboli identification in immunostained sections of breast carcinomas with higher identification rates than H&E. Lymphatic vessel invasion was related to other features (high combined histologic grade, high Ki67 score, negative hormone receptors expression) associated with worse prognosis, probable reflecting a potential for lymphatic metastatic spread and aggressive behavior.

Immunohistochemical analysis of host reaction to heavyweight-, reduced-weight-, and expanded polytetrafluoroethylene (ePTFE)-based meshes after short- and long-term intraabdominal implantations

Prosthetic meshes induce a variety of inflammatory changes in the host, which may lead to excessive scarring with detrimental clinical consequences, especially in the long term. This study aimed to characterize the degree of short- and long-term inflammatory changes induced by common prosthetic meshes. Twenty 4 x 4-cm samples each of expanded polytetrafluoroethylene (ePTFE), heavyweight polypropylene (hPP), ePTFE/heavyweight polypropylene (ePTFE/hPP), and reduced-weight polypropylene/regenerated cellulose (rPP) were implanted intraperitoneally in 40 rabbits for 4 or 12 months. After explantation, samples of mesh/tissue complex were analyzed for the degrees of cellular apoptosis (enzyme-linked immunoassay [ELISA]) and cellular turnover (mouse monoclonal antibody). In the short term, the degree of apoptosis in the hPP mesh was significantly higher than in the ePTFE and rPP groups. Similarly, it was higher in the ePTFE/hPP group than in either the ePTFE or the rPP group. The amount of Ki-67-positive cells was significantly higher in the hPP group than in the ePTFE or rPP group. The cell turnover in the ePTFE/hPP group was similar to that in the hPP group, but significantly higher than in either the ePTFE or the rPP group. The rPP group, in turn, had a higher Ki-67 score than the ePTFE group. In the long term, both the degree of apoptosis and Ki-67 positivity were significantly lower in the rPP and ePTFE groups than in either the ePTFE/hPP or the hPP group. A significant decrease in Ki-67 scores between the short and long-term groups was found only in the rPP group. In the short term, heavyweight polypropylene-based meshes were associated with significantly higher cell proliferation and death. A significantly higher degree of apoptosis and cell turnover were associated with heavyweight polypropylene-based meshes even 1 year after implantation, indicating ongoing inflammation and scar remodeling. On the other hand, ePTFE and reduced-weight polypropylene meshes were associated with nearly physiologic levels of inflammatory markers. Overall, an exaggerated and persistent host foreign body response to heavyweight polypropylene-based meshes indicates poor biocompatibility, with potential detrimental clinical sequela.

Bio-pathologic Characteristics Related to Chromosome 11 Aneusomy and Cyclin D1 Gene Status in Surgically Resected Stage I and II Breast Cancer: Identification of an Adverse Prognostic Profile

We aimed at developing a more detailed understanding of cyclin D1 in early stage human breast cancer and defining the biologic profiles with different prognostic value correlating cyclin D1 gene amplification and chromosome 11 aneusomy with bio-pathologic variables of known clinical importance. Cyclin D1 gene amplification and chromosome 11 aneusomy were investigated using fluorescence in situ hybridization whereas cyclin D1, PgR, HER-2, Bcl2, p53, and Ki-67 expressions were analyzed by immunohistochemistry in 121 stage I or II breast cancer patients uniformly treated with cyclophosphamide/metotrexate/5-fluorouracil-based chemotherapy. Cyclin D1 was amplified in 6.6% and overexpressed in 32.2% of cases. Amplification was not associated with any selected bio-pathologic variables, whereas the chromosome 11 aneusomy level significantly increased in tumors with higher histologic grade (P < 0.01), higher tumor size (P < 0.003), p53 nuclear accumulation (P < 0.04), and ERalpha negativity (P < 0.049). Multiple correspondence analysis showed 4 different biologic tumor profiles. The first, characterized by high Ki-67 score, p53+, cyclin D1+, HER-2+, aneusomy level > 30%, ratio (cyclin D1 gene/CEP11) > 2, was associated with tumor relapse defining the most unfavorable biologic profile. Kaplan-Meier's method showed significantly shorter disease-free survival in patients with at least 3 variables positive out of the 6 detected by multiple correspondence analysis. In multivariate analysis, the identified biologic profile emerged as the only significant prognostic indicator. Our findings are of particular clinical interest for early stage breast cancer patients, because the assessment of biologic factors predictive of tumor aggressiveness may influence postoperative therapeutic strategies.

Correlation of proliferative markers (Ki-67 and PCNA) with survival and lymph node metastasis in oral squamous cell carcinoma: a clinical and histopathological analysis of 113 patients

The purposes of this study were to examine the correlations between proliferation markers and survival rate in oral squamous cell carcinoma (OSCC) patients, and to evaluate the efficacy of proliferation markers in predicting lymph node metastasis. The patients’ age, gender, T score, clinical stage, PCNA and Ki-67 index were analysed. Univariate analysis showed that T score had a significant influence on survival, and stage 4 group had a significantly lower survival rate. Lymph node metastasis was also a significant predictor of survival. Using a cut-off point of 25%, those patients with lower Ki-67 scores had survival advantage over those with higher Ki-67 scores. PCNA did not show any differences in survival with a cut-off point of 50%. Ki-67 and PCNA were significantly higher in the primary tumours associated with lymph node metastasis (pN+) than in those without lymph node metastasis (pN0). Multivariate analysis showed that clinical stage and Ki-67 were independent prognostic factors for survival in OSCC patients. From this result, it can be postulated that the cancer staging based on the TNM stage was a powerful prognostic variable and Ki-67 had a significant effect on the cumulative survival rate.

The mitogen-activated protein kinases (MAPK) p38 and JNK are markers of tumor progression in breast carcinoma

Objective. To investigate the activation of mitogen-activated protein kinases (MAPK) in breast carcinoma effusions and to analyze its relationship to anatomic site and clinical parameters. Methods. Activated MAPK (p-ERK, p-JNK, and p-p38) expression was studied in 42 effusions and 51 corresponding solid tumors (23 primary, 28 metastases) using immunohistochemistry (IHC). Hormone receptor and HER2 status, proliferation (Ki-67), and apoptosis (p85-PARP fragment) were assessed. MAPK levels, activity, and activation ratio (phospho/pan-MAPK ratio) were analyzed in 19 effusions using immunoblotting (IB). Results. Nuclear expression of p-p38 and p-JNK was significantly higher in effusions compared with both primary tumors ( P < 0.001 for p-JNK, P = 0.011 for p-p38) and lymph node metastases ( P = 0.003 for p-JNK, P = 0.025 for p-p38) but was not accompanied by apoptosis. IB showed pan-ERK and p-ERK in 18/19 effusions, pan-JNK and p-JNK in 18/19 and 17/19 effusions, respectively, and pan-p38 and p-p38 in 19/19 and 17/19 specimens, respectively. In univariate survival analysis of all cases, advanced disease stage ( P = 0.041), previous chemotherapy ( P = 0.004), and radiation ( P = 0.001) and higher Ki-67 scores ( P = 0.01) correlated with worse overall survival (OS). In Cox multivariate analysis, stage ( P = 0.018), chemotherapy ( P = 0.024), radiation ( P = 0.017), and ER status ( P = 0.002) were independent prognosticators of OS. Quantitative analysis of IB data showed that higher p38 activation ratio correlates with shorter OS ( P = 0.01). Conclusions. This study presents first evidence of in vivo activation of MAPK in breast carcinoma effusions. The elevated JNK and p38 activation in effusions may be a stress-related mechanism providing breast carcinoma cells with survival advantages rather than a drive towards apoptosis. p38 and Ki-67 may be new prognostic markers for patients with breast cancer effusions.

Is Ki67 a marker for the transformation of laryngeal dysplasia to carcinoma?

Conclusion. Ki67 is not a reliable marker of malignant transformation in laryngeal dysplasia.Objectives. No reliable means of predicting which cases of laryngeal dysplasia will undergo malignant transformation currently exists. Our aim was to evaluate Ki67, a marker of cell proliferation, as a potential marker for the transformation of laryngeal dysplasia to squamous cell carcinoma.Patients and methods. Eighty consecutive cases of previously untreated patients with a histological diagnosis of laryngeal dysplasia from 1987 to 1993 were identified from the pathological archives. Standard immunohistochemical techniques were used to identify Ki67-positive cells and activity was scored on a scale of 0–4 using defined criteria.Results. Of the 80 cases there were 24 females and 56 males with a mean age of 56 years (range 29–80 years). Twenty cases subsequently transformed to a squamous cell carcinoma. For each Ki67 score (0–4), the rate of malignant transformation was: 0, 1 of 6 patients (17%); 1, 7 of 33 patients (21%); 2, 5 of 22 patients (23%); 3, 4 of 13 patients (31%); and 4, 3 of 6 patients (50%). A higher Ki67 score seemed to correlate with a higher likelihood of malignant transformation but this did not reach statistical significance (p = 0.17, Pearson χ2 test). Considering a score of 3 or 4 as positive for predicting malignant transformation produced a test of relatively high specificity (80%) but poor sensitivity (35%).

Prognostic factors in ovarian carcinoma stage III patients. Can biomarkers improve the prediction of short- and long-term survivors?

The aim of the study was to determine if biomarker expression could help discriminate between short-term and long-term survivors in women with advanced ovarian cancer. Fifty-one patients with stage III ovarian cancer were selected for the study, which included 28 short-term survivors (death from ovarian cancer within 18 months) and 23 long-term survivors (alive for more than 5 years). There was no difference between the two groups with respect to FIGO substage, age, World Health Organization score, and first-line platinum therapy. Classic clinical pathologic parameters were examined together with p53, Bcl-2, Ki-67, PDGFRalpha, P-glycoprotein, BRCA1, and DNA ploidy. Immunohistochemistry was used for scoring biomarker expression and image cytometry for DNA ploidy. All patients had primary debulking surgery followed by first-line platinum therapy. On multivariate analysis, the presence of ascites, debulking surgery and repeat laparotomy, clear-cell histology, elevated CA125, and high Ki-67 score were all found to be of prognostic importance. The long-term survivors were characterized by primary optimal cytoreduction surgery (<1 cm residual disease), attempt at maximal tumor debulking by experienced gynecological oncologic surgeons, and the absence of ascites. Normal CA125 level before platinum therapy and negative Ki-67 expression also predicted a more favorable prognosis.

Prognostic factors in ovarian carcinoma stage III patients. Can biomarkers improve the prediction of short- and long-term survivors?

The aim of the study was to determine if biomarker expression could help discriminate between short-term and long-term survivors in women with advanced ovarian cancer. Fifty-one patients with stage III ovarian cancer were selected for the study, which included 28 short-term survivors (death from ovarian cancer within 18 months) and 23 long-term survivors (alive for more than 5 years). There was no difference between the two groups with respect to FIGO substage, age, World Health Organization score, and first-line platinum therapy. Classic clinical pathologic parameters were examined together with p53, Bcl-2, Ki-67, PDGFRα, P-glycoprotein, BRCA1, and DNA ploidy. Immunohistochemistry was used for scoring biomarker expression and image cytometry for DNA ploidy. All patients had primary debulking surgery followed by first-line platinum therapy. On multivariate analysis, the presence of ascites, debulking surgery and repeat laparotomy, clear-cell histology, elevated CA125, and high Ki-67 score were all found to be of prognostic importance. The long-term survivors were characterized by primary optimal cytoreduction surgery (&lt;1 cm residual disease), attempt at maximal tumor debulking by experienced gynecological oncologic surgeons, and the absence of ascites. Normal CA125 level before platinum therapy and negative Ki-67 expression also predicted a more favorable prognosis.

Ki67 antigen and PCNA proliferation markers predict survival in anorectal malignant melanoma.

To find a possible correlation of Ki67 antigen and proliferating cell nuclear antigen (PCNA) with prognosis in anorectal malignant melanoma. Thirty patients with anorectal malignant melanoma were studied. The percentage of tumour cells stained for Ki67 and PCNA in paraffin sections was assessed. Mode of treatment (local excision or abdominoperineal resection), depth of tumour invasion, attempt at cure as defined by complete tumour excision and absence of distant metastases at presentation, tumour blood vessel invasion, and tumour necrosis, as well as Ki67 and PCNA, were all correlated with survival. By univariate analysis, PCNA, Ki67, attempt at cure, local excision (and not abdominoperineal resection), and depth of invasion were all significantly associated with longer survival. By multivariate analysis, only PCNA was significantly associated with survival, while Ki67 showed a significant positive correlation with PCNA. With a cut-off point of 40%, patients with lower Ki67 scores showed survival advantage over those with higher Ki67 scores (P=0.0004). With a cut-off point of 80%, patients with lower PCNA scores showed survival advantage over those with higher PCNA scores (P=0.0001). The staining for proliferation markers was also associated with depth of tumour invasion. Ki67 and PCNA immunostaining in paraffin sections may be useful for the prediction of survival in patients with anorectal malignant melanoma. Larger studies are needed to confirm our results.

Immunohistochemical expression of the transcription factor DP-1 and its heterodimeric partner E2F-1 in non-Hodgkin lymphoma.

DP-1 is a G1 cell cycle-related protein that forms heterodimers with E2F, a family of transcriptional factors regulating the expression of genes important for G1 to S progression. Although the exact role of DP-1 is not well understood, it has been shown to stabilize DNA binding of E2F proteins. By immunohistochemistry, the authors examined the expression of DP-1 in lymphoid tissues, including 8 cases of reactive follicular hyperplasia and 69 cases of B-cell non-Hodgkin lymphoma. The expression of the cell cycle-related proteins E2F-1 and Ki-67 was also assessed. Scoring was based on the proportion of labeled nuclei (1-10%, 11-25%, 26-50%, and > 50%). In reactive follicular hyperplasia, staining for DP-1, E2F-1, and Ki-67 was largely confined to the germinal centers. All 25 cases of follicular lymphoma, regardless of grade, had a high proportion (> 50%) of DP-1-positive cells but a lower proportion of cells marking for E2F-1 and Ki-67 (P < 0.001). The diffuse large B-cell lymphomas (n = 24) had high DP-1 and Ki-67 scores but low E2F-1 scores (P < 0.001). Small lymphocytic (n = 10), marginal zone (n = 3), and mantle cell lymphomas (n = 5) contained relatively low proportions of cells labeled for all three markers. Precursor B-cell lymphoblastic lymphoma (n = 2) displayed high proportions of cells positive for DP-1, Ki-67, and E2F-1 (> 50% in both cases). Except in follicular center cell lesions, DP-1 expression generally correlated with that of Ki-67. However, the expression of DP-1 was discordant with that of E2F-1 in benign and malignant follicular center cells, suggesting that DP-1 may have functions other than facilitating E2F-1-dependent gene regulation and cell cycle progression in these neoplasms.