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Key points During compensated hypertrophy in vivo fractional shortening (FS) remains constant until heart failure (HF) develops, when FS decreases from 70% to 39%.Compensated hypertrophy is accompanied by an increase in I Na,late and a decrease in Na+,K+‐ATPase current. These changes persist as HF develops.SR Ca2+ content increases during compensated hypertrophy then decreases in HF. In healthy cells, increases in SR Ca2+ content and Ca2+ transients can be achieved by the same amount of inhibition of the Na+,K+‐ATPase as measured in the diseased cells.SERCA function remains constant during compensated hypertrophy then decreases in HF, when there is also an increase in spark frequency and spark‐mediated Ca2+ leak.We suggest an increase in I Na,late and a decrease in Na+,K+‐ATPase current and function alters the balance of Ca2+ flux mediated by the Na+/Ca2+ exchange that limits early contractile impairment. Abstract We followed changes in cardiac myocyte Ca2+ and Na+ regulation from the formation of compensated hypertrophy (CH) until signs of heart failure (HF) are apparent using a trans‐aortic pressure overload (TAC) model. In this model, in vivo fractional shortening (FS) remained constant despite HW:BW ratio increasing by 39% (CH) until HF developed 150 days post‐TAC when FS decreased from 70% to 39%. Using live and fixed fluorescence imaging and electrophysiological techniques, we found an increase in I Na,late from –0.34 to –0.59 A F−1 and a decrease in Na+,K+‐ATPase current from 1.09 A F−1 to 0.54 A F−1 during CH. These changes persisted as HF developed (I Na,late increased to –0.82 A F−1 and Na+,K+‐ATPase current decreased to 0.51 A F−1). Sarcoplasmic reticulum (SR) Ca2+ content increased during CH then decreased in HF (from 32 to 15 μm l−1) potentially supporting the maintenance of FS in the whole heart and Ca2+ transients in single myocytes during the former stage. We showed using glycoside blockade in healthy myocytes that increases in SR Ca2+ content and Ca2+ transients can be driven by the same amount of inhibition of the Na+,K+‐ATPase as measured in the diseased cells. SERCA function remains constant in CH but decreases (τ for SERCA‐mediated Ca2+ removal changed from 6.3 to 3.0 s−1) in HF. In HF there was an increase in spark frequency and spark‐mediated Ca2+ leak. We suggest an increase in I Na,late and a decrease in Na+,K+‐ATPase current and function alters the balance of Ca2+ flux mediated by the Na+/Ca2+ exchange that limits early contractile impairment., Key points During compensated hypertrophy in vivo fractional shortening (FS) remains constant until heart failure (HF) develops, when FS decreases from 70% to 39%.Compensated hypertrophy is accompanied by an increase in I Na,late and a decrease in Na+,K+‐ATPase current. These changes persist as HF develops.SR Ca2+ content increases during compensated hypertrophy then decreases in HF. In healthy cells, increases in SR Ca2+ content and Ca2+ transients can be achieved by the same amount of inhibition of the Na+,K+‐ATPase as measured in the diseased cells.SERCA function remains constant during compensated hypertrophy then decreases in HF, when there is also an increase in spark frequency and spark‐mediated Ca2+ leak.We suggest an increase in I Na,late and a decrease in Na+,K+‐ATPase current and function alters the balance of Ca2+ flux mediated by the Na+/Ca2+ exchange that limits early contractile impairment.

We report a flexible light‐sheet fluorescence microscope (LSFM) designed for studying dynamic events in cardiac tissue at high speed in 3D and the correlation of these events to cell microstructure. The system employs two illumination‐detection modes: the first uses angle‐dithering of a Gaussian light sheet combined with remote refocusing of the detection plane for video‐rate volumetric imaging; the second combines digitally‐scanned light‐sheet illumination with an axially‐swept light‐sheet waist and stage‐scanned acquisition for improved axial resolution compared to the first mode. We present a characterisation of the spatial resolution of the system in both modes. The first illumination‐detection mode achieves dual spectral‐channel imaging at 25 volumes per second with 1024 × 200 × 50 voxel volumes and is demonstrated by time‐lapse imaging of calcium dynamics in a live cardiomyocyte. The second illumination‐detection mode is demonstrated through the acquisition of a higher spatial resolution structural map of the t‐tubule network in a fixed cardiomyocyte cell.

BACKGROUND Intraductal carcinoma of the prostate (IDC-P) is a distinct histopathologic feature associated with high-grade, advanced prostate cancer. Although studies have shown that IDC-P is a predictor of progression following surgical or radiation treatment for prostate cancer, there are sparse data regarding IDC-P on diagnostic needle biopsy as a prognosticator of prostate cancer mortality. MATERIALS AND METHODS This was a population-based study of all prostate cancer patients diagnosed using needle biopsy and without evidence of systemic disease between 1991 and 1999 within a defined geographic region of Norway. Patients were identified by cross-referencing the Norwegian Cancer Registry. Of 318 eligible patients, 283 had biopsy specimens available for central pathology review. Clinical data were obtained from medical charts. We examined whether IDC-P on diagnostic needle biopsy was associated with adverse clinicopathological features and prostate cancer mortality. RESULTS Patients with IDC-P on diagnostic needle biopsy had a more advanced stage and a higher Gleason score compared to patients without IDC-P. IDC-P was also associated with an intensively reactive stroma. The 10-year prostate cancer-specific survival was 69% for patients with IDC-P on diagnostic needle biopsy and 89% for patients without IDC-P (Log rank P-value

BACKGROUND Previous studies suggest that lymphovascular invasion (LVI) has a weak and variable effect on prognosis. It is uncertain whether LVI, determined by diagnostic prostate biopsy, predicts prostate cancer death. Data from experimental studies have indicated that carcinoma-associated fibroblasts in the reactive stroma could promote LVI and progression to metastasis. Thus, combining LVI with reactive stromal grade may identify prostate cancer patients at high risk of an unfavorable outcome. The purpose of the present study was to examine if LVI, determined by diagnostic biopsy, alone and in combination with reactive stromal grade could predict prostate cancer death. METHODS This population-based study included 283 patients with prostate cancer diagnosed by needle biopsy in Aust-Agder County (Norway) from 1991 to 1999. Clinical data were obtained by medical charts review. Two uropathologists evaluated LVI and reactive stromal grade. The endpoint was prostate cancer death. RESULTS Patients with LVI had marginally higher risk of prostate cancer death compared to patients without LVI (hazard ratio: 1.8, P-value = 0.04). LVI had a stronger effect on prostate cancer death risk when a high reactive stromal grade was present (hazard ratio: 16.0, P-value

Zinc finger E-box-binding homeobox 1 (ZEB1) is a transcriptional factor known to repress E-cadherin promoter and thus induce EMT. Expression of ZEB-1 has in numerous cancers been associated with aggressive disease and poor clinical outcome. Our aim was to investigate the expression of ZEB1 in esophageal squamous- and small-cell carcinomas. Immunohistochemical staining was performed on tissue sections obtained from 151 patients with esophageal squamous cell carcinoma (ESCC) and 25 patients with primary small-cell carcinoma of the esophagus (PSCCE). Semi-quantitative analysis, and thus statistical analysis, has been accomplished on the samples. Immunohistochemistry revealed ZEB1 expression in the cytoplasm (64.9% of cases), in nuclei (11.3% of cases) and in tumor stroma (80.1% of cases) of ESCC. In PSCCE only nuclear staining (88.0% of cases) was observed. Weak cytoplasmic expression of ZEB1 in ESCC was associated with longer survival. Immunohistochemical evaluation of ZEB1 cytoplasmic expression in ESCC may have clinical prognostic value, but further studies are needed to fully understand the function as well as potential clinical and therapeutic implications of ZEB1 expression in cancers.

BACKGROUND In vitro and in vivo studies have shown that nerves, tumor epithelium, and stroma interact and promote prostate cancer (PC) progression. Perineural invasion (PNI) is established amidst these interactions and may therefore indicate an aggressive PC phenotype. The purpose of the present study was to determine the relationship between PNI, tumor grade, reactive stroma, and PC-specific mortality. METHODS A population-based study on 318 patients, encompassing all cases of PC diagnosed by needle biopsies and without evidence of systemic metastasis at the time of diagnosis in Aust–Agder County in the period of 1991–1999. Patients were identified by cross-referencing the Cancer Registry of Norway. Clinical data were obtained by review of medical charts. Diagnostic prostate needle biopsies were reviewed with respect to presence of PNI, percentage of biopsy cores with PNI, Gleason score (GS), and reactive stromal grade (RSG). The endpoint was PC-specific mortality. RESULTS The presence of PNI was significantly associated with high tumor grade and abundant reactive stroma. The 10-year PC-specific survival for patients with and without PNI was 72% and 91%, respectively (P = 0.001, log rank). PNI predicted PC-specific mortality independently of clinical factors, though the effect of PNI was attenuated when adjusting for GS and RSG. However, a percentage of biopsy cores with PNI >50% was found to predict PC-specific mortality independently of other clinicopathologic parameters. CONCLUSIONS The present population-based study shows that PNI on diagnostic prostate needle biopsy is associated with increased risk of PC-specific mortality. Our findings demonstrate that the prognostic effect of PNI is dependent on an association with high grade carcinoma and reactive stroma. However; the impact of PNI on clinical outcome becomes stronger and independent of other clinicopathologic factors upon increased percentage of PNI positive biopsy cores. Thus, our study highlights the importance of PNI and microenvironmental interactions for the long-term outcome of PC. Prostate 76:207–214, 2016. © 2015 Wiley Periodicals, Inc.

BACKGROUND Reactive tumor stroma has been shown to play an active role in prostatic carcinogenesis. A grading system for reactive stroma in prostate cancer (PC) has recently been established and found to predict biochemical recurrence and prostate cancer-specific mortality (PCSM) in prostatectomized patients. To the best of our knowledge, there has been no study investigating the prognostic value of reactive stromal grading (RSG) with regard to PCSM when evaluated in diagnostic prostate needle biopsies. METHODS A population-based study on 318 patients, encompassing all cases of PC diagnosed by needle biopsies and without evidence of systemic metastasis at the time of diagnosis in Aust–Agder County in the period 1991–1999. Patients were identified by cross-referencing the Cancer Registry of Norway. Clinical data were obtained by review of medical charts. The endpoint was PCSM. RSG was evaluated on haematoxylin and eosin stained sections according to previously described criteria; grade 0, 0–5% reactive stroma; grade 1, 6–15%; grade 2, 16–50%; grade 3, 51–100%. RESULTS RSG could be evaluated in 278 patients. The median follow- up time was 110 months (interquartile range: 51–171). The 10-year PC - specific survival rate for RSGs of 0, 1, 2, and 3 was 96%, 81%, 69%, and 63%, respectively (P

Aims To investigate the aberrant expression of CD117 in oesophageal squamous cell carcinoma (SCC) and its prognostic significance. Methods and results Immunohistochemical staining for CD117 was performed on tissue microarray and routine tissue sections from 157 oesophageal SCC patients and 10 normal oesophageal epithelia adjacent to tumour. The positive rate of CD117 expression was 29.9% in oesophageal SCC tissues, whereas no CD117 expression was detected in the 10 normal oesophageal epithelia. CD117 expression was significantly associated with T stage (P

Detection of pretreatment disseminated cells (pre-DTC) reflecting its homing to bone marrow (BM) in prostate cancer (PCa) might improve the current model to predict recurrence or survival in men with nonmetastatic disease despite of primary treatment. Thereby, pre-DTC may serve as an early prognostic biomarker. Post-treatment DTCs (post-DTC) finding may supply the clinician with additional predictive information about the possible course of PCa. To assess the prognostic impact of DTCs in BM aspirates sampled before initiation of primary therapy (pre-DTC) and at least 2 years after (post-DTC) to established prognostic factors and survival in patients with PCa. Available BM of 129 long-term follow-up patients with T1-3N0M0 PCa was assessed in addition to 100 BM of those in whom a pretreatment BM was sampled. Patients received either combined therapy [n = 81 (63%)], radiotherapy (RT) with different duration of hormone treatment (HT) or monotherapy with RT or HT alone [n = 48 (37%)] adapted to the criteria of the SPCG-7 trial. Mononuclear cells were deposited on slides according to the cytospin methodology and DTCs were identified by immunocytochemistry using the pancytokeratin antibodies AE1/AE3. The median age of men at diagnosis was 64.5 years (range 49.5–73.4 years). The median long-term follow-up from first BM sampling to last observation was 11 years. Categorized clinically relevant factors in PCa showed only pre-DTC status as the statistically independent parameter for survival in the multivariate analysis. Pre-DTCs homing to BM are significantly associated with clinically relevant outcome independent to the patient's treatment at diagnosis with nonmetastatic PCa.

Aims Vascular endothelial growth factor (VEGF), VEGF receptor 1 (VEGFR-1) and VEGF receptor 2 (VEGFR-2) play a role in breast cancer growth and angiogenesis. We examined the expression and relationship with clinical outcome and other prognostic factors. Methods and results Tumour sections from 468 breast cancer patients were immunostained for VEGF, VEGFR-1, and VEGFR-2, and their relationships with tumour vascularity, disseminated tumour cells (DTCs) in bone marrow and other clinicopathological parameters were evaluated. VEGF, VEGFR-1 and VEGFR-2 immunoreactivities were observed in invasive breast carcinoma cells. VEGF expression was significantly associated with VEGFR-1 and VEGFR-2 expression (P Conclusions High-level expression of VEGFR-1 indicates reduced survival. Higher-level expression of VEGFR-1 or VEGFR-2 in primary breast carcinomas combined with the presence of DTC selects a prognostically unfavourable patient group.

Objectives: To assess the expression of the cell surface protein B7-H3 in prostate cancer, and its association to clinically relevant parameters after radical prostatectomy and to the proliferation marker Ki-67. Methods: Radical prostatectomy specimens from a cohort of 130 patients with a median clinical follow up of 8 years were used for the analysis. The expression of B7-H3 and the proliferation marker Ki-67, as well as other standard clinicopathological parameters, were evaluated. Results: A high expression of B7-H3 was associated with pathological stage T3a and T3b, high Gleason score, extraprostatic extension, seminal vesicle invasion and high proliferative activity. Univariable analysis showed that a high expression level of B7-H3 was also correlated with biochemical failure and clinical relapse, and with the expression of Ki-67. A high expression level of Ki-67 was associated with clinical progression and a tendency towards higher rates of prostate-specific antigen relapse in multivariate analyses. Conclusions: Our findings show that a high expression level of B7-H3 in prostate cancer correlates with the expression of the proliferation marker Ki-67, biochemical failure and clinical relapse. Thus, expression of the cell surface molecule B7-H3 adds to the malignant phenotype of prostate cancer cells expressing high levels of Ki-67. The impact of B7-H3 function on prostate cancer and its potential role in immunotherapy should be explored further.

UNLABELLED Several authors have previously reported that transrectal prostate biopsy has a false-negative rate of 20-30%, and that a number of prostate cancers missed on transrectal biopsy can be detected by transperineal biopsy. It has also been shown that most of these tumours are located anteriorly in the prostate gland. The present study showed a high rate of prostate cancer in patients with previous negative transrectal biopsies but elevated PSA levels, and that the cancers were located anteriorly in the prostate gland. Also, most of these cancers were clinically significant in patients that underwent RP, i.e. a high proportion of cancers were high-grade/high-stage tumours. We also showed that the transperineal biopsy technique can be applied successfully to patients with a closed anal orifice after previous surgery for rectal cancer. Transperineal biopsy can be done safely without routine antibiotic prophylaxis. OBJECTIVE To investigate the outcomes of transperineal prostate biopsies in patients with elevated prostate-specific antigen (PSA) levels and negative transrectal biopsies. The aim of this retrospective study was to evaluate the diagnostic yield of the transperineal biopsy approach in these patients, and to evaluate the pathology findings in subsequent radical prostatectomy (RP) specimens in patients undergoing RP. PATIENTS AND METHODS In all, 69 consecutive patients with previous negative transrectal biopsies but elevated PSA levels investigated at urological units in Norway who had been referred to The Norwegian Radium Hospital were included. The patients had undergone a mean (median; range) of 2.42 (2; 0-7) transrectal biopsies. The mean (range) age was 63.1 (42-78) years. The median (range) PSA level was 12 (4.3-229) ng/mL. The patients were examined with transperineal biopsy of the prostate between July 2007 and February 2009. The results of the transperineal biopsies were reviewed for Gleason biopsy score, and these were compared with the histopathology results of the RP specimens, i.e. final Gleason scores. Pathological stage of the prostate specimens and tumour volume were also reviewed. RESULTS Prostate cancer was found in the biopsies of 38 of 69 patients (55%). In all, 20 of 38 patients had a Gleason score estimated at ≥3 + 4 = 7. In all, 26 patients underwent RP. The surgical specimens revealed pathological stage pT2c in 65%, pT3a in 27% and pT3b in 8% of the cases. In all, 23 of the 26 RP specimens showed a final Gleason score of ≥7. The vast majority of cancers detected were situated in the anterior/ventral portion of the prostate. CONCLUSIONS Transperineal biopsy of the prostate in patients with an elevated PSA level after negative transrectal prostate biopsies appears to be a feasible and important option for further investigation to detect prostate cancer. The present study shows that the transperineal biopsy allows good access of the anterior/ventral part of the prostate. Histopathology reports on the RP specimens obtained from the patients that underwent RP revealed significant cancer.

UNLABELLED Study Type - Prognosis (case series) Level of Evidence 4 What's known on the subject? and What does the study add? It is known that a tertiary Gleason grade pattern 4 or 5 found in RP specimens has a negative impact on recurrence rate regarding biochemical relapse after radical prostatectomy. This is the first publication addressing clinical outcome in patients with a tertiary Gleason grade pattern 4 or 5 showing a negative influence on clinical failure rates. OBJECTIVE To investigate the impact of a tertiary Gleason grade (TGG) pattern 4 or 5 on clinical failure, as the presence of a TGG pattern 4 or 5 in radical prostatectomy (RP) specimens has been associated with biochemical failure. PATIENTS AND METHODS In all, 151 consecutive patients undergoing RP between 1985 and 2006 were reviewed, and 148 patients met study inclusion criteria. The RP specimens were pathologically re-examined and the presence of a TGG pattern 4 or 5 was recorded. The endpoint was clinical failure defined as local recurrence and/or development of metastasis at a mean follow-up of 108 months. Univariate analyses were performed using the Kaplan-Meier method. Multivariate analyses were performed using Cox proportional hazards regression. RESULTS Clinical failure was more likely among men with presence of a TGG pattern 4 or 5 than in men without a TGG pattern 4 or 5 (P= 0.006). In the subgroup of patients with Gleason score 7 the presence of a TGG 5 was significantly associated with clinical failure rate (P= 0.002). In patients with Gleason score 7, a TGG pattern 4 or 5 was not associated with increased failure rates. Multivariate Cox regression analyses in patients with Gleason score 7 showed that a TGG pattern 5 was a statistically significant predictor of clinical failure when adjusting for pathological stage, surgical margin status, extraprostatic extension and seminal vesicle invasion (hazard ratio 4.03, 95% confidence interval 1.72-9.46; P= 0.001). Further subgroup analyses showed that a TGG pattern 5 was associated with statistically higher clinical progression rates in patients with Gleason score 3 + 4 (P= 0.03). In patients with Gleason score 4 + 3, a TGG pattern 5 was associated with a trend towards a higher clinical progression rate, although this was not statistically significant (P= 0.189). CONCLUSION A TGG pattern 4 or 5 is associated with decreased clinical recurrence-free survival in Gleason score 7.

B7-H3, an immunoregulatory protein, is known to play a role in tumor progression. In many cancer types, observed correlations between high B7-H3 expression and poor prognosis have been attributed to involvement in antitumor immunity. However, here we demonstrate a nonimmunological alternative function of B7-H3 in cancer metastasis. Since advanced malignant melanoma is a disease with a poor survival rate and a broad pattern of metastasis, we used this disease as a model in our studies. We found that shRNA silencing of B7-H3 reduced the in vitro migratory potential and matrigel invasiveness of MDA-MB-435 and FEMX-I melanoma cells. In an experimental metastasis model in vivo, B7-H3 silencing of MDA-MB-435 cells resulted in reduced metastatic capacity and significantly increased the median symptom-free survival of nude mice (147 vs. 65 days, p < 0.001) and rats (53 vs. 42 days, p = 0.025) injected with MDA-MB-435 cells. Furthermore, a smaller fraction of mice had microscopically detectable metastases compared to control animals, and the pattern of metastases was slightly different between the two groups but with the brain as the predominant organ. Immunohistochemistry on samples from two melanoma patients showed strong B7-H3 staining in both a primary tumor and metastases. Notably, the metastasis-associated proteins, matrix metalloproteinase (MMP)-2, signal transducer and activator of transcription 3 (Stat3), and the level of secreted interleukin-8 (IL-8) were reduced in the B7-H3 knock-down cell variants, whereas tissue inhibitor of metalloproteinase (TIMP)-1 and-2 levels were increased. Taken together, our findings indicate a novel role for B7-H3 in the regulation of the metastatic capacity of melanoma cells and it might be a potential therapeutic target for anti-metastasis therapy.

Y. Yuan, H. P. Dong, D. A. Nymoen, J. M. Nesland, C. Wu and B. Davidson PINCH-2 expression in cancers involving serosal effusions using quantitative PCR Objective: The PINCH-2 gene was previously shown to be overexpressed in malignant mesothelioma compared with ovarian/peritoneal serous carcinoma in Affymetrix array analysis. The objective of the present study was to validate this finding at the mRNA and protein level. Methods: Effusions (n = 91; 71 ovarian and 10 breast carcinomas, 10 malignant mesotheliomas) were assayed for PINCH-2 mRNA expression using quantitative PCR. PINCH-2 protein expression was analysed in 37 effusions using flow cytometry. Results: Quantitative PCR analysis showed significantly higher PINCH-2 mRNA levels in mesotheliomas compared with carcinomas (P = 0.004). Values of

In epoxy resin-embedded bone marrow biopsies from 238 cases of myeloproliferative disease and preleukaemia, crystalline inclusions were found in the bone marrow cells of 24 (10.1%), most often in patients with acute myeloproliferative disease (23%). By light microscopy the crystals were easily recognized after Giemsa staining, lying within the cytoplasm of bone marrow macrophages. By electron microscopy the hexagonal crystals were encountered chiefly within secondary lysosomes of the macrophages, but also within immature myeloid cells in one of the two cases studied. In the latter a tight-fitting membrane around the crystals suggested that they were of lysosomal origin and represented a form of abnormal granulation. We suggest that the presence of crystals in myeloid cells represents a kind of abnormal granula formation in the neoplastic cells. In the macrophages the crystals lodge in secondary lysosomes, probably after phagocytosis of crystal-bearing non-viable myeloid cells. The accumulation of crystals in lysosomes indicates resistance to degradation.

The objective of this study was to analyze large-scale genomic patterns during disease progression from primary tumor to effusion in ovarian carcinoma, and to study the association between DNA ploidy parameters in effusions, proliferation/survival markers, and clinicopathologic characteristics. DNA ploidy status, DNA index (DI), and S-phase fraction (SPF) were compared in 22 matched primary carcinomas (all prechemotherapy specimens) and effusions (14 prechemotherapy and 8 postchemotherapy specimens) using image analysis. The association between these parameters and previously studied cell survival/proliferation biomarkers, previous administration of chemotherapy, chemotherapy response and survival was analyzed in a larger series of 54 effusions. The majority of specimens were aneuploid irrespective of anatomic site, with no significant differences in DI. SPF was significantly higher in effusions compared to matched primary tumors (P = 0.007 for all 22 pairs, P = 0.011 for 14 matched prechemotherapy specimens). Higher SPF was related to higher Ki-67 score (P = 0.045), and both SPF and DI were directly associated with higher level of Survivin (P < 0.001 for both). DI and SPF in effusions showed no association with histological grade, FIGO stage, residual disease volume, previous chemotherapy, response to chemotherapy at primary disease, recurrence or survival. Ovarian carcinoma cells in effusions have increased proliferation compared to corresponding primary tumors, as evidence of disease progression. DNA ploidy parameters in cancer cells in effusions are unaltered by chemotherapy and appear to be unrelated to chemotherapy response and to survival, suggesting that large-scale genomic patterns at this anatomic site are not useful in segregating patients into prognostic groups. Diagn. Cytopathol. 2008;36:637–644. © 2008 Wiley-Liss, Inc.

Expression profiles of primary breast tumors were investigated in relation to disseminated tumor cells (DTCs) in bone marrow (BM) in order to increase our understanding of the dissemination process. Tumors were classified into five pre-defined molecular subtypes, and presence of DTC identified (at median 85 months follow-up) a subgroup of luminal A patients with particular poor outcome ( p ¼ 0.008). This was not apparent for other tumor subtypes. Gene expression profiles associated with DTC and with systemic relapse for luminal A patients were identified. This study suggests that DTC in BM differentially distinguishes clinical outcome in patients with luminal A type tumors and that DTC-associated gene expression analysis may identify genes of potential importance in tumor dissemination. a 2007 Federation of European Biochemical Societies.

Immunocytochemical (ICC) detection of disseminated tumor cells (DTC) in bone marrow (BM) in early breast cancer is an independent prognostic factor. The significance of circulating tumor cells (CTC) in peripheral blood (PB) needs further exploration and comparison to DTC detection. PB and BM were prospectively collected from 341 breast cancer patients median 40 months after operation. PB samples were analyzed for tumor cells by a negative immunomagnetic technique (10x10(6) cells/test). BM aspirates were analyzed by standard ICC (2x10(6) cells/test). CTC were present in 10% of the patients and DTC in 14%. Thirty-seven relapses and 14 breast cancer deaths have occurred at median 66 months after diagnosis. Both CTC-status and DTC-status were significantly associated with disease free survival (DFS) (event rate: CTC-positive 26.5% vs. CTC-negative 9.1%; DTC-positive 29.2% vs. DTC-negative 7.8%) (p

OBJECTIVE To compare the immunohistochemically assessed expression of the epidermal growth factor receptor (EGFR) family in normal and malignant bladder urothelium, and suggest new hypotheses about their function in the development and progression of transitional cell carcinoma (TCC). PATIENTS AND METHODS EGFR, ERBB2, ERBB3 and ERBB4 were evaluated immunohistochemically in normal urothelium (NU, 15), primary non-metastasized invasive TCC (NMC, 19) and in primary invasive TCCs with corresponding metastases (MC, 51, both specimens). RESULTS All NU samples expressed ERBB4, none expressed ERBB2 and two expressed EGFR; all staining was uniform throughout all cell layers. ERBB2 expression increased and ERBB4 decreased from normal samples to carcinomas. There was no difference between NMCs and MCs in ERBB2, ERBB3 and ERBB4, but the NMCs expressed more EGFR than both NU and MC samples. There were no associations with T category, grade or survival. All combinations of expression levels for the four receptors were detected, with no dominant profile. CONCLUSION We hypothesise that: (i) ERBB4 is important for differentiation in NU; (ii) ERBB2 is up-regulated with carcinogenesis in the urinary bladder but does not discriminate between bladder cancer with or without metastases; (iii) EGFR may be a marker of indolent disease. A current hypothesis, that superficial layers of NU do not express EGFR and thus protect the basal cells from the mitogenic effect of urinary EGF, is challenged.

BACKGROUND It was demonstrated previously that the Snail family of transcription factors and Smad-interacting protein 1 (Sip1) regulate E-cadherin and matrix metalloproteinase 2 (MMP-2) expression, cellular morphology, and invasion in carcinoma. For the current study, the authors analyzed the relation between the expression of Snail, Slug, and Sip1; the expression of MMP-2 and E-cadherin; and clinical parameters in patients with metastatic ovarian and breast carcinoma. METHODS One hundred one fresh-frozen, malignant effusions from patients who were diagnosed with gynecologic carcinomas (78 ovarian carcinomas and 23 breast carcinomas) were studied for mRNA expression of Snail, Slug, Sip1, MMP-2, and E-cadherin using reverse transcriptase-polymerase chain reaction analysis. Snail mRNA and E-cadherin protein expression levels also were studied in ovarian carcinoma effusions using in situ hybridization and immunocytochemistry. The results were analyzed for possible correlation with clinicopathologic parameters in both tumor types. RESULTS E-cadherin mRNA expression was lower in breast carcinoma (P = 0.001), whereas Snail expression was higher (P = 0.003). The Snail/E-cadherin ratio (P < 0.001) and the Sip1/E-cadherin ratio (P = 0.002) were higher in breast carcinomas. Sip1 mRNA expression (P < 0.001) and Slug mRNA expression (P < 0.001) were correlated with the expression of MMP-2 in ovarian carcinomas. The Sip1/E-cadherin ratio was higher in primary ovarian carcinomas at the time of diagnosis compared with postchemotherapy ovarian carcinoma effusions (P = 0.003), higher in Stage IV tumors compared with Stage III tumors (P = 0.049), and higher in pleural effusions compared with peritoneal effusions (P = 0.044). In a univariate survival analysis of patients with ovarian carcinoma, a high Sip1/E-cadherin ratio predicted poor overall survival (P = 0.018). High E-cadherin mRNA expression predicted better disease-free survival (P = 0.023), with a similar trend for a low Slug/E-cadherin ratio (P = 0.07). High Snail mRNA expression predicted shorter effusion-free survival (P = 0.008), disease-free survival (P = 0.03), and overall survival (P = 0.008) in patients with breast carcinoma. CONCLUSIONS Transcription factors that regulate E-cadherin were expressed differentially in metastatic ovarian and breast carcinoma. Snail may predict a poor outcome in patients who have breast carcinoma metastatic to effusions. E-cadherin expression generally was conserved in effusions from patients with ovarian carcinoma, but the subset of patients with postulated Sip1-induced repression of this adhesion molecule had a significantly worse outcome. This finding was in agreement with the stronger suppression of E-cadherin by Snail and Sip1 in breast carcinoma effusions, a clinical condition associated with extremely poor survival. Cancer 2005. © 2005 American Cancer Society.

The transcription factor PU.1 has been shown to be crucial for the early stages of B cell development but its function at later stages of B cell development is less well known. We observed previously that PU.1 is expressed uniformly throughout the mature pre-plasma cell B cell population, the only exception being a subpopulation of germinal centre (GC) cells which showed exceptionally high expression of PU.1. This suggested that PU.1 may also have a role in GC B cell biology. To test this hypothesis and to screen for possible genes regulated by PU.1, we first evaluated semi-quantitatively the possible co-expression of PU.1 with proteins known to be upregulated or downregulated during GC B cell development. Normal lymphoid tissues and 255 B cell non-Hodgkin lymphomas of putative GC B cell origin were evaluated. PU.1 expression was positively associated with CD10 (p < 0.0001), CD20 (p = 0.043), CD22 (p = 0.005), CD79a (p = 0.024) and Bcl-6 (p < 0.0001) and negatively associated with cytoplasmic immunoglobulin light-chain expression (p = 0.036) in diffuse large B cell lymphoma. Identical or nearly identical associations were found in follicular lymphoma. Since CD20 is known to be partly regulated by PU.1 and putative PU.1-binding sites have been described in the regulatory regions of the CD22, CD79a and CD10 genes, we looked for putative PU.1 binding sites in the BCL6 promotor. Four such putative PU.1 binding sites were identified. Further analysis by gel-shift electromobility essay showed that PU.1 protein binds to three of the four putative binding sites in the BCL6 promotor. PU.1 and Bcl-6 were also found to be upregulated in centroblasts in the normal GC, but jointly downregulated in a subpopulation of centrocytes. Our findings support the contention that PU.1 may also have an important role in GC B cell development.

Pilomatrix carcinoma is a rare skin tumor with an origin from hair matrix cells. The tumor is locally aggressive with a great tendency for recurrence, but the metastatic potential is limited. A pilomatrix carcinoma in 76-year-old female with lymph node metastases is presented. In addition to classical histopathological criteria and DNA ploidy analysis, a broad panel of antibodies was used for evaluation of the metastatic potential. Both primary tumor and lymph node metastasis revealed extremely high proliferation and apoptotic rates. High constant expressions of CD44v6 and P-cadherin were also observed. In the metastasis, significant reduction of E-cadherin and β-catenin was detected. The best approach for assessment of metastatic potential of pilomatrix carcinoma seems to be the complex evaluation of routine histological criteria like vessel invasion, mitotic index, apoptotic count, and new molecular markers of cell death and adhesion.

We report a tubular adenoma of the esophagus in a 79-year-old man. The tumour had progressed to an intramucosal carcinoma at follow-up 6 months later. The adenoma displayed a peculiar and heterogeneous histological picture with non-dysplastic and cystic areas alternating with those of a typical tubular adenoma. The tumour revealed a strong and diffuse expression of cytokeratin 7 and basal cell cytokeratin, whereas cytokeratin 20 was focally positive only. The most remarkable finding, however, was the demonstration of intracytoplasmic inclusion-like mucoid bodies of the epithelial cells in a small area of the adenomatous component. These inclusions stained PAS positive and alcian blue negative, and electron microscopy revealed a homogeneous structure without viral-like particles.

OBJECTIVES: The role of cyclooxygenase-2 (COX-2) in disease has been extensively studied, (Annu Rev Med (2002) 53:35; N Engl J Med (2001) 345:433) but less information is available with respect to possible physiological functions of COX-2. Information on how and where COX-2 is expressed under physiological conditions may increase our understanding of its physiological role. Previous studies have revealed a COX-2 dependent production of prostanoids under physiological conditions, without entirely determining the source of this production. MATERIALS AND METHODS: To assess COX-2 expression under normal conditions, we analyzed tissue specimens that were removed from 30 healthy study subjects in conjunction with surgical procedure related to insertion of dental implants and from three patients which had muscle tissue from Quadriceps femoris muscle removed as part of surgical treatment of soft tissue sarcomas not directly affecting the muscle tissue. Immunohistochemistry and immunoblotting (Western blotting) was used to assess the presence of COX-2 protein. RESULTS: In 25 of 30 patients (83%), COX-2 protein was expressed in striated muscle, as assessed by immunohistochemistry. All cases had COX-2 expression verified by Western blotting. In none of the 25 subjects with COX-2 expression did we notice concomitant inflammation of the adjacent submucosal tissue. CONCLUSIONS: It is a novel finding that COX-2 is expressed in striated muscle under physiological conditions. COX-2 activity in striated muscle is a possible explanation for the hitherto unknown localization of prostanoids synthesis under physiological conditions.

Aims: The purpose of this study was to examine the occurrence of CD44 isoforms in breast carcinomas and their role in predicting clinical outcome. Methods and results: Shock-frozen tumour tissues from 110 patients with breast carcinoma were examined by immunohistochemistry using antibodies directed against CD44s, v5, v6, v7 and v3–10. In addition, 80 of these tumours were available for quantitative real-time reverse transcriptase-polymerase chain reaction (RT-PCR) analysis of CD44s and CD44v6. Immunohistochemically, the positive tumours showed cytoplasmic and/or membranous staining with all antibodies. Staining results did not correlate with histological subtype, lymph node status, status of steroid receptors, tumour size or age. Neither was any correlation found for overall and disease-free survival. Quantitative real-time RT-PCR of CD44s and CD44v6, however, revealed that expression of CD44v6 mRNA was significantly associated with lower pathological grade (Pearson χ2 test P = 0.009; linear-by-linear association P = 0.003). Linear-by-linear association between CD44s mRNA expression and lower pathological grade was also seen (P = 0.02). Survival analysis with the Kaplan–Meier method demonstrated that increased CD44s mRNA expression was significantly associated with both disease-free survival and overall survival (P = 0.0185 and P = 0.0344, respectively). A similar trend for CD44v6 mRNA expression was seen in these cases, but the difference was not significant. Conclusions: Quantitative real-time RT-PCR revealed clinical correlations of CD44s and CD44v6 mRNA expression in breast carcinomas while immunohistochemistry for the protein expression of CD44s and other CD44 variants did not. This contradictory result merits further studies concerning the clinical impact of CD44 molecules in breast carcinomas.

A large number of experimental studies have linked the S100A4 gene product to the metastatic phenotype of cancer cells and clinical evidence indicates a correlation between S100A4 expression and poor prognosis in several cancer types. The aim of the present study was to analyse the expression of the S100A4 protein in colorectal cancer. Paraffin-embedded samples from 277 colorectal cancer patients were immunostained with anti-S100A4 antibody. Cytoplasmic staining was observed in 178 of 277 samples (64%), whereas, unexpectedly, nuclear expression of S100A4 was found in 88 of 277 of the samples (32%). This novel finding was confirmed by western blot analysis of nuclear fractions isolated from frozen tumour tissue. Statistical analysis revealed a significant correlation between nuclear expression of S100A4 and tumour stage at diagnosis, while there was no such correlation between cytoplasmic staining and tumour stage. The nuclear localization of S100A4 in colorectal cancer and its relationship to tumour stage suggest that this protein may be involved in gene regulatory pathways of relevance to the metastatic phenotype of cancer cells.

The majority of genetic studies of colorectal carcinogenesis have focused on changes found in primary tumors. Despite the fact that liver metastases are a leading cause of colorectal cancer deaths, the molecular genetic basis of the advanced disease stages remains poorly understood. We performed comparative genomic hybridization (CGH) on 17 liver metastases from colorectal carcinomas and compared the quantitative profile with the qualitative profile previously obtained with chromosome banding. An average of 12.6 aberrations per tumor was found by CGH. Chromosome 18 and chromosome arms 4q, 8p, and 17p were most frequently lost, whereas chromosomes 7 and 20 and chromosome arms 6p, 8q, and 13q were most frequently gained. We compared the chromosome banding and CGH data after converting the karyotypes into net copy number gains and losses. Ten tumors showed agreement between the findings of the two techniques, whereas five tumors did not (in two cases, no mitotic cells were obtained for banding analysis). All five discordant cases had a “simple” abnormal or normal karyotype, but revealed multiple changes by CGH. A likely explanation for this discrepancy is that in vitro growth before G-banding selected against the cancer cells. Interestingly, by comparing the CGH profiles of the “complex” vs. the “simple”/normal karyotype groups, deletion of 8p and gain of 16q were seen more frequently in the former group. The liver metastases had the same aberrations as seen in primary colorectal carcinomas, summarized in a literature survey. However, these aberrations were seen more frequently in liver metastases, which may be attributable to increased genetic instability. © 2003 Wiley-Liss, Inc.

Our study evaluates the prognostic significance of the cytokeratin-positive mononuclear cells (CK+ cells) in the bone marrow (BM) and peripheral blood (PB) as detected by immunocytochemistry in patients with locoregionally confined prostate cancer. BM and PB samples were obtained from 66 newly diagnosed patients with T1-4pN0M0 prostate cancer. All samples were analyzed by standardized immunocytochemical methods (anticytokeratin mononuclear antibody; AE1/AE3) applying a negative immunomagnetic cell enrichment technique. A second sampling was obtained in 60 of the 66 patients >or=2 years after definitive radiotherapy. The median follow-up after high-dose radiotherapy of the patients was 65 months. For the analysis of the postradiotherapy clinical progression-free survival (PFS) treatment, failure was defined as pelvic tumor growth or development of distant metastases. At diagnosis CK+ cells were found in BM in 14 of 66 (21%) prostate cancer patients. This was not associated with an increased risk of progression. On the other hand, the presence of CK+ cells in 12 of 60 (20%) patients at the second BM aspiration was significantly related to a shorterPFS (p = 0.02). In the multivariate analysis, the presence of CK+ cells in the posttreatment BM did not remain as an independent variable of PFS assessment if posttreatment PSA was entered into the analysis. CK+ cells in PB were found in 12% of the patients. After therapy, none of the patients had detectable CK+ cells in PB. The presence of CK+ cells in the posttreatment but not in the pretreatment BM was associated with decreased PFS in patients irradiated for pelvis-confined nonmetastatic prostate cancer. Although this association was not retained in multivariate analysis, our observations indicate that the presence of CK+ cells after local therapy define a group of patients that have a high risk of developing distant metastases.

Differential Ki67 and bcl-2 immunoexpression in solid–glandular and spindle cell components of biphasic synovial sarcoma: a double immunostaining assessment with cytokeratin and vimentin Aims: Synovial sarcoma is a malignant soft tissue of uncertain histogenesis that may show a biphasic (spindle and solid/glandular components) or a monophasic histological appearance. In previous studies, we demonstrated that the solid/glandular component possesses higher proliferation rates than the spindle cell component of biphasic synovial sarcomas and that the spindle cell component may exhibit a progressive transition from or to the solid-glandular component in biphasic synovial sarcoma. To evaluate this hypothesis further, we designed a novel approach to correlate immunoexpression of Ki67, bcl-2 and bax in the spindle cell and in the solid-glandular component of biphasic synovial sarcomas. We also performed a double-immunohistochemical assessment of the Ki67 proliferative indices and the immunoexpression of anti-apoptotic protein bcl-2 in neoplastic cells expressing either vimentin or cytokeratin. Methods and results: Immunohistochemistry for vimentin (10 cases), bcl-2 (10 cases), Ki67(10 cases), cytokeratin (10 cases), and bax (eight cases), and double-immunostaining for vimentin/Ki67 (10 cases), vimentin/bcl-2 (nine cases), cytokeratin/Ki67 (10 cases), and cytokeratin/bcl-2 (10 cases) assays were performed in 10 cases of primary biphasic synovial sarcoma. Semiquantitative assessment was adopted for each case in both components. Statistical analysis was performed using Fisher's exact test or χ2 test. On conventional immunohistochemistry, the solid/glandular component revealed more expression of Ki67, bax and cytokeratin than the spindle cell component (P=0.0004, P=0.082, and P

The present study analysed by immunohistochemistry the protein level of cyclin A and Ki-67 in a panel of paraffin-embedded tissue obtained from 172 primary (110 superficial and 62 nodular) and 73 metastatic melanomas, and ten benign naevi. Since cyclin A exists in the same quaternary complex in the S-phase of the cell cycle as the cdk inhibitor p21WAF1/CIP1, the levels of the two proteins were compared. Cyclin A and Ki-67 were heterogeneously expressed in the malignant tumours, whereas in benign naevi, only rare positive cells were detected. In superficial spreading melanomas, the cyclin A level was related to tumour thickness, with less expression in thinner lesions (p

Progression through the mammalian cell cycle is facilitated by cyclin–cyclin-dependent kinase (cdk) complexes, which are activated at specific points during the cell cycle. Alteration in cyclin–cdk complexess may lead to altered cell cycle and tumorigenesis. In this study, we analyzed expression of cyclins A, D1, D3 and E in tumor tissue from 170 patients with primary invasive breast carcinomas. Immunohistochemical methods were used to detect protein expression of these cyclins. We detected positive immunoreactivity in 55 (32%), 22 (13%), 38 (22%) and 37 (21.8%) of the samples for cyclins A, D1, D3 and E, respectively. A highly statistically significant association was observed between expression of cyclin A and early relapse (p = 0.001 univariate analysis, p = 0.006 multivariate analysis) as well as cancer-specific death (p < 0.0001) during the follow-up time. No association was observed between cyclin D1 or cyclin E, respectively, and relapse of disease or survival, while cyclin D3 over-expression was associated with development of metastases during follow-up (p = 0.005 univariate analysis, p = 0.01 multivariate analysis). However, cyclin D3 did not show any statistically significant association when cancer-specific death was examined in a multivariate analysis (Cox regression for survival function). © 2001 Wiley-Liss, Inc.

The use of automated microscopy has reached the maturity necessary for its routine use in the clinical pathology laboratory. In the following study we compared the performance of an automated microscope system (MDS™) with manual method for the detection and analysis of disseminated tumor cells present in bone marrow preparations from breast carcinoma patients. The MDS System detected rare disseminated tumor cells among bone marrow mononuclear cells with higher sensitivity than standard manual microscopy. Automated microscopy also proved to be a method of high reproducibility and precision, the advantage of which was clearly illustrated by problems of variability in manual screening. Accumulated results from two pathologists who had screened 120 clinical slides from breast cancer patients both by manual microscopy and by use of the MDS System revealed only two (3.8%) missed by the automatic procedure, whereas as many as 20 out of 52 positive samples (38%) were missed by manual screening. Cytometry (Comm. Clin. Cytometry) 46:215‐221, 2001. © 2001 Wiley-Liss, Inc.

BACKGROUND Approximately 30% of breast carcinoma patients with negative lymph nodes die of their disease. Biologic markers such p53 protein and c-erb B-2 have been related to tumor progression, but their prognostic value remains controversial. METHODS Two large series of a total of 613 lymph node negative breast carcinoma patients from a single institution were analyzed with respect to tumor size, histologic grade, and immunohistochemical staining for p53, c-erb B-2, estrogen receptor (ER), and progesterone receptor (PgR). Interobserver variation in histologic grading was evaluated by Kappa statistics. The two series had different treatment modalities: 228 patients (SACGS group) were treated surgically with mastectomy and given 1 perioperative chemotherapy course, and 385 patients (HOST group) were treated with mastectomy and ovarian radiation and further randomized to receive postoperative treatment with radiotherapy or no adjuvant treatment. The follow-up ranged from 14–30 years. RESULTS Immunoreactivities for p53, c-erb B-2, ER, and PgR did not differ significantly in the two series. p53 immunostaining was present in 187 of 613 tumors (29%), and c-erb B-2 immunoreactivity was present in 58 of the tumors (10%). Three hundred forty-eight tumors (57%) were positive for ER. Kappa statistics value of interobserver variation in the histologic grading of ductal carcinomas was 0.69, which is considered to be a substantial degree of agreement. No significant differences in survival were found when comparing p53, c-erb B-2, ER, and PgR positive and negative cases. However, both recurrence free survival rates and overall survival rates after 10 years were significantly better in the T1N0M0 group compared with the T2N0M0 group (81% vs. 67% [P 20–50 mm. Approximately 35% of the patients with tumors of histologic Grades 2–3 and measuring > 20 mm were dead after 10 years of follow-up, contrary to 6% of the patients with tumors of histologic Grade 1 measuring ≤ 20 mm. A significantly more favorable prognosis also was observed in patients in the HOST group treated with adjuvant radiotherapy. CONCLUSIONS Histologic grade and tumor size were found to be major prognostic factors for patients after 30 years of follow-up. c-erb B-2 and p53 immunostaining does not appear have any independent prognostic value. Adjuvant radiotherapy may be of value in the treatment of patients with localized tumors. Cancer 2000;88:804–13. © 2000 American Cancer Society.

The aims of this study were firstly, to investigate the expression of E-cadherin complex proteins in ovarian carcinoma cells in serous effusions and in primary and metastatic lesions; and secondly to study the value of these four proteins and calretinin, a mesothelial marker, in the differential diagnosis of ovarian carcinoma cells from reactive mesothelial cells in effusions. Sixty-seven malignant effusions and 97 corresponding primary (n=36) and metastatic (n=61) lesions were immunohistochemically stained for E-cadherin and alpha-, beta-, and gamma-catenin. Staining extent and intensity were scored. Effusion specimens were additionally analysed for calretinin immunoreactivity. Membrane immunoreactivity for E-cadherin and alpha-, beta-, and gamma-catenin was detected on carcinoma cells in the majority of the effusions, but rarely on reactive mesothelial cells (p

In breast cancer, mutations located in the zinc-binding functional domains of the p53 gene have been reported to predict a worse prognosis and a worse response to treatment with doxorubicin, compared with mutations in other parts within exons 5–8 of the gene. Similarly, mutations in residues of p53 that directly contact DNA have been associated with a poor prognosis. To investigate whether these specific p53 mutations are associated with differences in the rate of apoptosis and/or mitosis, or expression of the anti-apoptotic Bcl-2 protein, these parameters were evaluated in 89 invasive breast cancers with a confirmed p53 mutation in exons 5–8 and in 99 tumours without a p53 mutation in exons 5–8. Neither mutations located in the zinc-binding functional domains nor mutations in residues that directly contact DNA were associated with alterations in mitotic or apoptotic activity. However, compared with the wild-type p53 tumours, both apoptotic and mitotic indices showed an approximately two-fold increase in the mutant p53 group ( p< 0·001). The presence of a p53 mutation was also associated with the presence of tumour necrosis ( p< 0·001), high tumour grade ( p< 0·001) and low expression of Bcl-2 ( p< 0·001). Our data support the concept that in invasive breast carcinoma, loss of p53 function is involved in enhanced proliferation rather than decreased apoptosis and that the resulting acceleration of cell turnover may enhance clonal evolution and tumour progression. Copyright © 1999 John Wiley & Sons, Ltd.

In the present study, TP53 alterations have been analysed and compared with the expression of the proteins p21, cyclin D1, cdk4, RB, EGFR, and MDM2 in 53 cancers of the uterine corpus. TP53 gene mutations analysed by CDGE/DGGE and direct sequencing showed a TP53 gene mutation in 18 per cent of the cases. TP53 gene mutations were not significantly related to overexpression or down-regulation of any of the proteins. Immunohistochemically, there was an increased protein level of TP53 in 77 per cent, p21 in 36 per cent, cyclin D1 in 45 per cent, cdk4 in 77 per cent, EGFR in 8 per cent, and MDM2 in 32 per cent of the cases. Expression of RB protein was normal in all cancers. Significant association of protein expression was seen between TP53 and MDM2 (p=0.005) and p21 and MDM2 (p=0.001). Furthermore, there may be an association between TP53 and p21 (p=0. 038) and cyclin D1 and cdk4 (p=0.045). The results revealed increased levels of TP53 protein in all MDM2-positive cases that did not show TP53 mutations, indicating TP53 protein stabilization and inactivation by complex formation with MDM2. In summary, the high number of cases showing an increased level of TP53 and cdk4 proteins suggests that these proteins play an important role in the neoplastic process in cancers of the uterine corpus.

BACKGROUND The objective of the study was to evaluate the pathogenetic and prognostic value of p53 protein expression in squamous cell carcinoma of the vulva. METHODS The clinical data in charts of 167 patients with International Federation of Gynecology and Obstetrics (FIGO) Stages I-III primary tumors who were treated by surgery were reviewed. Samples from the primary tumor were immunostained for p53 protein. p53 overexpression was defined as immunoreactivity in > 5% of nuclei. RESULTS p53 overexpression was observed in 92 tumors (55%). p53 overexpression did not correlate with age at diagnosis, FIGO stage, histologic grade, vessel invasion, tumor thickness, tumor greatest dimension, DNA ploidy, or inguinal lymph node metastasis. In the whole group a significantly reduced 5-year corrected survival was observed in patients with p53 overexpression compared with p53 negative patients (P = 0.04). In the different FIGO stages, disease-related survival was not influenced by p53 overexpression in 37 patients with Stage I disease (P = 0.60) or in 86 patients with Stage II disease (P = 0.96). In 44 patients with Stage III disease, p53 overexpression was significantly associated with poorer prognosis (P = 0.004). Independent prognostic factors for corrected survival in the entire group of 167 patients were: vascular invasion, groin metastasis, tumor greatest dimension, and p53 overexpression. In patients with FIGO Stage III disease p53 overexpression was not an independent prognostic factor. CONCLUSIONS p53 protein overexpression appears to be involved in the pathogenesis of vulvar squamous cell carcinoma. p53 protein overexpression was significantly associated with disease-related survival. p53 prognostic impact was observed only in patients with advanced disease. Cancer 1999;85:1133–8. © 1999 American Cancer Society.