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Urothelial carcinoma of the renal pelvis is a deadly disease with an unclear tumorigenic mechanism. We conducted gene expression profiling on a set of human tumors of this type and identified a phosphatidylinositol 3-kinase (PI3K)/AKT activation expression signature in 76.9% (n = 13) of our samples. Sequence analysis found both activating mutations of PIK3CA (13.6%, n = 22) and loss of heterozygosity at the PTEN locus (25%, n = 8). In contrast, none of the other subtypes of kidney neoplasms (e.g., clear-cell renal cell carcinoma) harbored PIK3CA mutations (n = 87; P < 0.001). Immunohistochemical analysis of urothelial carcinoma samples found loss of PTEN protein expression (36.4%, n = 11) and elevation of phosphorylated mammalian target of rapamycin (mTOR; 63.6%, n = 11). To confirm the role of the PI3K/AKT pathway in urothelial carcinoma, we generated mice containing biallelic inactivation of Pten in the urogenital epithelia. These mice developed typical renal pelvic urothelial carcinomas, with an incidence of 57.1% in mice older than 1 year. Laser capture microdissection followed by PCR confirmed the deletion of Pten exons 4 and 5 in the animal tumor cells. Immunohistochemical analyses showed increased phospho-mTOR and phospho-S6K levels in the animal tumors. Renal lymph node metastases were found in 15.8% of the animals with urothelial carcinoma. In conclusion, we identified and confirmed an important role for the PI3K/AKT pathway in the development of urothelial carcinoma and suggested that inhibitors of this pathway (e.g., mTOR inhibitor) may serve as effective therapeutic agents. [Cancer Res 2009;69(21):8256–64]

Supplementary Figure 1 from Activation of the PI3K/AKT Pathway Induces Urothelial Carcinoma of the Renal Pelvis: Identification in Human Tumors and Confirmation in Animal Models

Supplementary Table 1 from Activation of the PI3K/AKT Pathway Induces Urothelial Carcinoma of the Renal Pelvis: Identification in Human Tumors and Confirmation in Animal Models

Supplementary Figure Legend from Activation of the PI3K/AKT Pathway Induces Urothelial Carcinoma of the Renal Pelvis: Identification in Human Tumors and Confirmation in Animal Models

Supplementary Table 2 from Activation of the PI3K/AKT Pathway Induces Urothelial Carcinoma of the Renal Pelvis: Identification in Human Tumors and Confirmation in Animal Models

Clear cells sarcomas (CCS) are exceptionally rare in the tongue, with, to our knowledge, only three previous reports in anglo-saxon literature. Through our case, we will discuss the differential diagnosis of clear cells tumors of the tongue and bring this tumour closer to the newly described entity of the gastrointestinal tract named "clear cells sarcoma-like gastrointestinal (SCCLGI)", recently renamed "gastrointestinal neuroectodermal tumour (GNET)". SCCLGI/GNET share morphological and molecular characteristics with SCC but had until then been observed only in the digestive tract. Our case could be a lingual localization of a SCCLGI/GNET. SCC and SCCLGI/GNET characteristic molecular profil involves EWSR1-ATF1 [t(12; 22) (q13; q12)] and EWSR1-CREB1 [t(2; 22) (q34; q12)] fusion genes, but it is not specific of these tumours.

Clear cells sarcomas (CCS) are exceptionally rare in the tongue, with, to our knowledge, only three previous reports in anglo-saxon literature. Through our case, we will discuss the differential diagnosis of clear cells tumors of the tongue and bring this tumour closer to the newly described entity of the gastrointestinal tract named "clear cells sarcoma-like gastrointestinal (SCCLGI)", recently renamed "gastrointestinal neuroectodermal tumour (GNET)". SCCLGI/GNET share morphological and molecular characteristics with SCC but had until then been observed only in the digestive tract. Our case could be a lingual localization of a SCCLGI/GNET. SCC and SCCLGI/GNET characteristic molecular profil involves EWSR1-ATF1 [t(12; 22) (q13; q12)] and EWSR1-CREB1 [t(2; 22) (q34; q12)] fusion genes, but it is not specific of these tumours.

Contains fulltext : 171799.pdf (Publisher’s version ) (Closed access) Personalized medicine has gained increasing importance in clinical oncology, and several clinically important biomarkers are implemented in routine practice. In an effort to guarantee high quality of molecular testing in France, three subsequent external quality assessment rounds were organized at the initiative of the National Cancer Institute between 2012 and 2014. The schemes included clinically relevant biomarkers for metastatic colorectal (KRAS, NRAS, BRAF, PIK3CA, microsatellite instability) and non-small cell lung cancer (EGFR, KRAS, BRAF, PIK3CA, ERBB2), and they represent the first multigene/multicancer studies throughout Europe. In total, 56 laboratories coordinated by 28 regional molecular centers participated in the schemes. Laboratories received formalin-fixed, paraffin-embedded samples and were asked to use routine methods for molecular testing to predict patient response to targeted therapies. They were encouraged to return results within 14 calendar days after sample receipt. Both genotyping and reporting were evaluated separately. During the three external quality assessment rounds, mean genotype scores were all above the preset standard of 90% for all biomarkers. Participants were mainly challenged in case of rare insertions or deletions. Assessment of the written reports showed substantial progress between the external quality assessment schemes on multiple criteria. Several essential elements such as the clinical interpretation of test results and the reason for testing still require improvement by continued external quality assessment education.

The gene cyclin-dependent kinase inhibitor 2A (CDKN2A) is frequently inactivated by deletion in bladder carcinoma. However, its role in bladder tumourigenesis remains unclear. We investigated the role of CDKN2A deletion in urothelial carcinogenesis, as a function of FGFR3 mutation status, a marker for one of the two pathways of bladder tumour progression, the Ta pathway. We studied 288 bladder carcinomas: 177 non-muscle-invasive (123 Ta, 54 T1) and 111 muscle-invasive (T2–4) tumours. CDKN2A copy number was determined by multiplex ligation-dependent probe amplification, and FGFR3 mutations by SNaPshot analysis. FGFR3 mutation was detected in 124 tumours (43.1%) and CDKN2A homozygous deletion in 56 tumours (19.4%). CDKN2A homozygous deletion was significantly more frequent in FGFR3-mutated tumours than in wild-type FGFR3 tumours (p = 0.0015). This event was associated with muscle-invasive tumours within the FGFR3-mutated subgroup (p < 0.0001) but not in wild-type FGFR3 tumours. Similar findings were obtained for an independent series of 101 bladder carcinomas. The impact of CDKN2A deletions on recurrence-free and progression-free survival was then analysed in 89 patients with non-muscle-invasive FGFR3-mutated tumours. Kaplan–Meier survival analysis showed that CDKN2A losses (hemizygous and homozygous) were associated with progression (p = 0.0002), but not with recurrence, in these tumours. Multivariate Cox regression analysis identified CDKN2A loss as a predictor of progression independent of stage and grade. These findings highlight the crucial role of CDKN2A loss in the progression of non-muscle-invasive FGFR3-mutated bladder carcinomas and provide a potentially useful clinical marker for adapting the treatment of such tumours, which account for about 50% of cases at initial clinical presentation. Copyright © 2012 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.

Background Lower urinary tract symptoms (LUTS) suggestive of benign prostatic hyperplasia (BPH) and erectile dysfunction (ED) are highly prevalent in aging men and are strongly linked. Alpha 1 -blockers such as alfuzosin are effective monotherapies for LUTS. Phosphodiesterase type 5 (PDE5) inhibitors such as tadalafil are the first-line treatment for ED. Both drugs act by two different mechanisms of action on common urogenital target organs and, thus, may have additive effects. Objectives We evaluated in vitro the effects of alfuzosin, tadalafil, and the combination of both on human detrusor and prostatic smooth muscle. Design, setting, and participants Prostatic and bladder tissue were obtained from patients ( n =20 and n =17, respectively) undergoing cystoprostatectomy for bladder cancer. Measurements In organ baths, isolated prostatic strips and isolated bladder strips were incubated with vehicle, tadalafil (10 −6 M and 10 −5 M), alfuzosin (3×10 −8 M or 10 −6 M and 10 −5 M) or a combination. Concentration-response curves (CRCs) to norepinephrine were generated on prostatic strips and detrusor strips precontracted with carbachol. Strips were also submitted to electrical field stimulation (EFS). Results and limitations When alfuzosin and tadalafil were combined, the maximal relaxation to norepinephrine on carbachol-precontracted detrusor strips was significantly increased compared with tadalafil alone, and EFS-induced detrusor contractions were better inhibited compared with each compound alone. Tadalafil significantly inhibited norepinephrine-induced prostatic strip contractions by reducing the maximal effect, whereas alfuzosin shifted the CRC of norepinephrine to the right. Combining both tadalafil and alfuzosin resulted in a greater relaxant effect. Likewise, the combination was more effective at reducing EFS-induced contractions compared with each compound alone. Conclusions The combination of alfuzosin and tadalafil exerts an additive effect of inhibiting adrenergic smooth muscle tone of prostatic tissue and EFS-induced detrusor contractions and conversely, of enhancing adrenergic relaxation of detrusor precontracted with carbachol. These experiments provide experimental support for the clinical investigation of the combination of α1-blockers and PDE5 inhibitors in the treatment of LUTS.

Purpose The indications of adjuvant chemotherapy for patients with estrogen receptor (ER) –positive breast cancer are controversial. We analyzed the predictive value of Ki67, HER2, and progesterone receptor (PR) expression for the efficacy of docetaxel in patients with ER-positive, node-positive breast cancer. Patients and Methods Expression of Ki67, HER2, and PR was measured by immunohistochemistry in tumor samples from 798 patients with ER-positive breast cancer who participated in PACS01, a randomized trial that evaluated the efficacy of docetaxel. Risk reduction was evaluated using a Cox model adjusted for age, tumor size, nodal involvement, treatment arm, and biomarkers. The predictive value of biomarkers was assessed by an interaction test. Disease-free survival (DFS) was the primary end point. Results Ki67, HER2, and PR were expressed in 21%, 9%, and 62% of samples, respectively. Hazard ratios for relapse associated with docetaxel were 0.51 (95% CI, 0.26 to 1.01) in ER-positive/Ki67-positive tumors and 1.03 (95% CI, 0.69 to 1.55) in ER-positive/Ki67-negative tumors (ratio for interaction: 0.53; 95% CI, 0.24 to 1.16; P = .11). Five-year DFS rates were 81% (95% CI, 76% to 86%) and 84% (95% CI, 75% to 93%) in patients with ER-positive/Ki67-negative and ER-positive/Ki67-positive tumors treated with docetaxel and 81% (95% CI, 76% to 86%) and 62% (95% CI, 52% to 72%) in patients with ER-positive/Ki67-negative and ER-positive/Ki67-positive tumors treated with fluorouracil, epirubicin, and cisplatin. No trend for interaction was observed between docetaxel and HER2 (ratio for interaction: 0.83; 95% CI, 0.35 to 1.94; P = .66), nor between docetaxel and PR (ratio for interaction: 0.89; 95% CI, 0.47 to 1.66; P = .71). Conclusion Ki67 expression identifies a subset of patients with ER-positive breast cancer who could be sensitive to docetaxel treatment in the adjuvant setting.

We report clinicopathologic features of a large series of renal translocation carcinomas from a multicentric study. Diagnosis was performed by cytogenetic examination of fresh material and/or by immunochemistry with antibodies directed against the C-terminal part of transcription factor E3 (TFE3) and native transcription factor EB (TFEB) proteins. Clinical data, follow-up, and histologic features were assessed. Antibodies against CK7, CD10, vimentin, epithelial membrane antigen, AE1-AE3, E-cadherin, alpha-methylacyl-coenzyme A racemase, melan A, and HMB45 were tested on tissue microarrays. Whole-genome microarray expression profiling was performed on 4 tumors. Twenty-nine cases were diagnosed as TFE3 and 2 as TFEB renal translocation carcinomas, including 13 males and 18 females, mean age 24.6 years. Two patients had a previous history of chemotherapy and 1 had a history of renal failure. Mean size of the tumor was 6.9 cm. Thirteen cases were > or = pT3 stage. Twelve cases were N+ or M+. Mean follow-up was 29.5 months. Three patients presented metastases and 5 have died. Mixed papillary and nested patterns with clear and/or eosinophilic cells represented the most consistent histologic appearance, with common foci of calcifications regardless of the type of translocation. Using a 30 mn incubation at room temperature, TFE3 immunostainings were positive in only 82% of our TFE3 translocation carcinomas. Both TFE3 and TFEB renal translocation carcinomas expressed CD10 and alpha-methylacyl-coenzyme A racemase in all cases. An expression of E-cadherin was observed in two-third of cases. Cytokeratins were expressed in less than one-third of cases. Melanocytic markers were expressed at least weakly in all cases except two. Unsupervised clustering on the basis of the gene expression profiling indicated a distinct subgroup of tumors. TRIM 63 glutathione S-transferase A1 and alanyl aminopeptidase are the main differentially expressed genes for this group of tumors. Our results suggest that these differentially expressed genes may serve as novel diagnostic or prognostic markers.

Objective Aschoff's center of proliferation (ACP), poses significant problems of differential diagnosis both in imagery and histology with infiltrating carcinoma. Up to now the criteria of Tabar and Dean (classical criteria) are considered as diagnostically relevant. Material A retrospective study of 113 cases, enabled us to study their clinical, radiological and histological aspects. Results The ACP is a subclinical and seldom palpable entity (12%). The radiological signs of ACP are quite variable. The classical criteria lack specificity and are observed only in 48% of our stellate images. Whereas the frequency of microcalcifications is high (58.5% of the cases), their presence and their type are not predictive of an associated malignancy. The echographic diagnosis of ACP could be made in 55% of the cases but the echographic semiology lacked specificity. We noticed an associated atypical epithelial hyperplasia in 28.5% of the cases, intraductal or lobular in situ carcinoma in 9% and/or a ductal invasive carcinoma in 2% of the cases. Neither tumor size, age of the patients, nor any radiological signs were predictive of such an association. Conclusions The classical criteria are not completely reliable and are observed only in half of our stellate images, whereas microcalcifications are often present but are not predictive of an associated malignancy.

The aim of the study was to investigate the relationships between the expression of alphav, beta1, beta3, beta5, and beta6, integrin subunits and clinical parameters in ovarian cancers. Ovarian surface epithelium (OSE) from five donors and tumour samples from 39 patients with an epithelial ovarian cancer (39 primary tumours and 21 associated peritoneal metastases) were analysed using immunohistochemistry on paraffin-embedded or frozen tissue sections. The alphav and beta5 integrin subunits were always present in normal OSE and in tumours. beta1 and beta3 subunit expression was significantly less frequent in grade 3 than in grade 1-2 tumours. The proportion of stage IV tumours expressing beta3 was significantly lower as compared to other stages. The beta6 subunit was undetectable in OSE but was expressed in about 40% of primary tumours. For all integrin, there was a strong relationship between the expression in primary tumours and in associated peritoneal metastases. Survival analyses restricted to patients receiving platinum-based chemotherapy did not reveal any relationship between integrin subunit expression and 3-year survival rate, in this limited series of patients. In conclusion, the expression of the various beta integrin subunits was differentially altered in ovarian carcinoma, evocative of complementary roles of alphav integrins during tumour development.

Background The aims of the present work were to study the prognostic impact of multiploidy and/or hypoploidy in breast cancers and their relation to other classic clinicopathologic prognostic factors (T, grade, receptors, and lymph node status). Methods From 3 January 1990 to 7 January 1999, 1984 previously untreated, invasive breast carcinoma samples were snap frozen for flow-cytometry. Results Multiploid tumors had the same prognosis as the aneuploid ones, and those with one hypoploid peak had a better prognosis than did the other aneuploid tumors. However, the presence of both multiploid and hypoploid peaks was correlated with a poor outcome, even after multivariate analysis. In this series after quality control, 93.4% of the histograms could be evaluated concerning ploidy; of these 81.6% could be assessed concerning S-phase fraction (SPF) in the entire population and 77.1% in the multiploid population. In the entire population, we performed a multivariate analysis including all relevant prognostic factors remaining after monovariate analysis by using a compound factor (proliferative activity) regrouping SPF and mitotic activity. This analysis showed that lymph node status and proliferative activity correlates with every type of survival, whereas receptor status correlates with all types of survival except recurrence free survival size, correlated with non-metastasis and overall survival. Grade and age correlated only with overall survival and vascular permeations only with disease-free survival. Conclusions SPF is a valuable predictor of survival, can be confidently assessed in multiploid histograms, and thus improves the yield of flow cytometry. When combined with mitotic activity, the prognostic impact of SPF is the same as that of lymph node status. Tumors that are hypoploid and multiploid have a significantly worse prognosis. Cytometry Part B (Clin. Cytometry) 55B:37–45, 2003. © 2003 Wiley-Liss, Inc.

Aims: HER2 protein is over-expressed in 15–30% of breast carcinomas. Immunohistochemistry (IHC) is a common and inexpensive method able to specifically detect HER2 protein. However, lack of standardization of IHC has been considered responsible for discrepancies in HER2 status assessment performed by IHC and fluorescence in-situ hybridization (FISH). This prompted us to perform a multicentric IHC calibration test to achieve a maximum accuracy of HER2-IHC compared with HER2-FISH taken as the reference method. Methods and results: Twelve French laboratories participated in this study, including 119 cases of invasive breast carcinomas for which both fixed and frozen tissues were available. HER2 expression was determined in fixed tissues by individual in-house IHC techniques, using either CB11 (Novocastra, Newcastle, UK) or A0485 (Dako, Glostrup, Denmark) anti-HER2 antibodies. Two cut-off values were used: 10% and 60% of immunostained cells. In 116 of the 119 cases, HER2 gene status could also be determined by FISH on frozen sections, performed in a single laboratory. Results were centralized and compared. When suboptimal concordance between IHC and FISH was observed, IHC was calibrated and a second run was performed. The specificity, sensitivity and accuracy of IHC compared with FISH were noted before and after calibration. Forty-four out of 116 (38%) tumours showed HER2 gene amplification. Accuracy of IHC was complete in the first run for 6/12 laboratories. Calibration, necessary for the six others, relied mainly on the combination of a heat-induced epitope retrieval step with an increase of dilution of the primary antibody. In the second run, HER2 over-expression was found in 46 (40%) and 44 (38%) of the 116 cases, using 10% or 60% of stained cells as cut-offs, respectively. The corresponding accuracy rates were 93% and 95%. Conclusions: This study showed that a high accuracy of IHC could be obtained for the determination of HER2 status in all laboratories using their in-house IHC technique, provided that a calibration process was performed. Antigen retrieval procedure, high dilutions of anti-HER2 antibody and the use of specific controls were crucial for HER2-IHC calibration. A 95% accuracy rate of IHC, using FISH as gold standard, was obtained by considering immunolabelling HER2-IHC results as a continuous variable, and taking 60% invasive stained cells as the cut-off for HER2 over-expression.

Muscle-invasive bladder carcinoma (MIBC) constitutes a heterogeneous group of tumors with a poor outcome. Molecular stratification of MIBC may identify clinically relevant tumor subgroups and help to provide effective targeted therapies. From seven series of large-scale transcriptomic data (383 tumors), we identified an MIBC subgroup accounting for 23.5% of MIBC, associated with shorter survival and displaying a basal-like phenotype, as shown by the expression of epithelial basal cell markers. Basal-like tumors presented an activation of the epidermal growth factor receptor (EGFR) pathway linked to frequent EGFR gains and activation of an EGFR autocrine loop. We used a 40-gene expression classifier derived from human tumors to identify human bladder cancer cell lines and a chemically induced mouse model of bladder cancer corresponding to human basal-like bladder cancer. We showed, in both models, that tumor cells were sensitive to anti-EGFR therapy. Our findings provide preclinical proof of concept that anti-EGFR therapy can be used to target a subset of particularly aggressive MIBC tumors expressing basal cell markers and provide diagnostic tools for identifying these tumors.

BACKGROUND The clinical relevance of DNA image cytometry (ICM) and flow cytometry (FCM) remains under investigation in breast carcinoma. The objective of the current work was to study the prognostic value of DNA ICM and FCM in a series of patients randomized in a control trial. A multivariate analysis has been performed including other factors still under investigation such as Ki-67 index, mitotic count, microvessel density, and P53 and Bcl-2 expression. METHODS Two hundred and eighty-one patients were randomized in the European Organization for Research and Treatment of Cancer 10854 trial comparing surgery followed by one course of perioperative chemotherapy versus surgery alone. Tumor parameters studied were pT, multicentricity, tumor grading according to modified Scarff–Bloom–Richardson, estrogen receptors, mitotic count per 1.7 mm2, MIB-1, and BCL-2 scores, microvessel density, and p53 expression. ICM DNA parameters studied from paraffin embedded specimens, were DNA ploidy, proliferative index, 2c deviation index, malignancy grade, and Auer–Baldetorp typing. FCM DNA parameters analyzed on the same samples were ploidy and S-phase fraction statistics. The influence of tumor parameters, and DNA parameters on overall survival (OS), disease free survival (DFS), and metastasis-free survival (MFS) was evaluated using the Cox model. Median follow-up was 82 months. RESULTS For OS, the prognostic parameters retained were pathologic tumor size (pT) and mitotic index (MI). Overall survival was 94% and 68% for tumors pT1/MI less than 10 and pT2-3 MI greater than or equal to 10, respectively. For DFS, age, multicentricity, and grading according to modified Scarff and Bloom were predicting factors with the same relative risk. Disease free survival was 96%, 78% and 68% respectively, when 1, 2, or 3 of those factors were present. For MFS, the only retained predicting factor was MI. MFS was 97% and 73% when MI was less than 10 and MI was greater than or equal to 10, respectively. CONCLUSIONS Evaluation of proliferative compartment was the most important predicting factor for OS and MFS in the current series of premenopausal lymph node negative patients with breast invasive carcinoma. When working on paraffin embedded tissue, the best way of assessing it was MI count. ICM DNA analysis results were not selected in multivariate analysis. DNA analysis by FCM should be considered as an unsuitable technique when working on paraffin embedded tissue. Cancer 2000;89:1748–57. © 2000 American Cancer Society.

We report the case of a 48-year-old female patient who had a Crohn's disease treated by corticosteroids. The patient developed severe cardiac failure, which was refractory to treatment with inotropic agents. At necropsy, examination of the heart revealed myocardial abscesses. On microscopic study, we diagnosed an aspergillar myocarditis. Aspergillar myocarditis is a rare and fatal localisation in disseminated aspergillosis. Diagnosis is difficult and treatment, usually initiated late, is ineffective.

Background Primary leiomyosarcoma of the seminal vesicle is exceedingly rare. Case Presentation We report a case of a 59-year-old man with tumour detected by rectal symptoms and ultrasonography. Computed tomography and magnetic resonance imaging suggested an origin in the right seminal vesicle. Transperineal biopsy of the tumour revealed leiomyosarcoma. A radical vesiculo-prostactectomy with bilateral pelvic lymphadenectomy was performed. Pathological examination showed a grade 2 leiomyosarcoma of the seminal vesicle. The patient received adjuvant radiotherapy. He developed distant metastases 29 months after diagnosis, and received chemotherapy. Metastatic disease was controlled by second-line gemcitabine-docetaxel combination. Fifty-one months after diagnosis of the primary tumour, and 22 months after the first metastases, the patient is alive with excellent performance status, and multiple asymptomatic stable lung and liver lesions. Conclusions We report the eighth case of primary leiomyosarcoma of the seminal vesicle and the first one with a so long follow-up.

The gene cyclin-dependent kinase inhibitor 2A (CDKN2A) is frequently inactivated by deletion in bladder carcinoma. However, its role in bladder tumourigenesis remains unclear. We investigated the role of CDKN2A deletion in urothelial carcinogenesis, as a function of FGFR3 mutation status, a marker for one of the two pathways of bladder tumour progression, the Ta pathway. We studied 288 bladder carcinomas: 177 non-muscle-invasive (123 Ta, 54 T1) and 111 muscle-invasive (T2-4) tumours. CDKN2A copy number was determined by multiplex ligation-dependent probe amplification, and FGFR3 mutations by SNaPshot analysis. FGFR3 mutation was detected in 124 tumours (43.1%) and CDKN2A homozygous deletion in 56 tumours (19.4%). CDKN2A homozygous deletion was significantly more frequent in FGFR3-mutated tumours than in wild-type FGFR3 tumours (p = 0.0015). This event was associated with muscle-invasive tumours within the FGFR3-mutated subgroup (p0.0001) but not in wild-type FGFR3 tumours. Similar findings were obtained for an independent series of 101 bladder carcinomas. The impact of CDKN2A deletions on recurrence-free and progression-free survival was then analysed in 89 patients with non-muscle-invasive FGFR3-mutated tumours. Kaplan-Meier survival analysis showed that CDKN2A losses (hemizygous and homozygous) were associated with progression (p = 0.0002), but not with recurrence, in these tumours. Multivariate Cox regression analysis identified CDKN2A loss as a predictor of progression independent of stage and grade. These findings highlight the crucial role of CDKN2A loss in the progression of non-muscle-invasive FGFR3-mutated bladder carcinomas and provide a potentially useful clinical marker for adapting the treatment of such tumours, which account for about 50% of cases at initial clinical presentation.

Background Germline mutations in the folliculin (FLCN) gene are associated with the development of Birt-Hogg-Dubé syndrome (BHDS), a disease characterized by papular skin lesions, a high occurrence of spontaneous pneumothorax, and the development of renal neoplasias. The majority of renal tumors that arise in BHDS-affected individuals are histologically similar to sporadic chromophobe renal cell carcinoma (RCC) and sporadic renal oncocytoma. However, most sporadic tumors lack FLCN mutations and the extent to which the BHDS-derived renal tumors share genetic defects associated with the sporadic tumors has not been well studied. Methods BHDS individuals were identified symptomatically and FLCN mutations were confirmed by DNA sequencing. Comparative gene expression profiling analyses were carried out on renal tumors isolated from individuals afflicted with BHDS and a panel of sporadic renal tumors of different subtypes using discriminate and clustering approaches. qRT-PCR was used to confirm selected results of the gene expression analyses. We further analyzed differentially expressed genes using gene set enrichment analysis and pathway analysis approaches. Pathway analysis results were confirmed by generation of independent pathway signatures and application to additional datasets. Results Renal tumors isolated from individuals with BHDS showed distinct gene expression and cytogenetic characteristics from sporadic renal oncocytoma and chromophobe RCC. The most prominent molecular feature of BHDS-derived kidney tumors was high expression of mitochondria-and oxidative phosphorylation (OXPHOS)-associated genes. This mitochondria expression phenotype was associated with deregulation of the PGC-1α-TFAM signaling axis. Loss of FLCN expression across various tumor types is also associated with increased nuclear mitochondrial gene expression. Conclusions Our results support a genetic distinction between BHDS-associated tumors and other renal neoplasias. In addition, deregulation of the PGC-1α-TFAM signaling axis is most pronounced in renal tumors that harbor FLCN mutations and in tumors from other organs that have relatively low expression of FLCN. These results are consistent with the recently discovered interaction between FLCN and AMPK and support a model in which FLCN is a regulator of mitochondrial function.

Urothelial carcinoma of the renal pelvis is a deadly disease with an unclear tumorigenic mechanism. We conducted gene expression profiling on a set of human tumors of this type and identified a phosphatidylinositol 3-kinase (PI3K)/AKT activation expression signature in 76.9% (n = 13) of our samples. Sequence analysis found both activating mutations of PIK3CA (13.6%, n = 22) and loss of heterozygosity at the PTEN locus (25%, n = 8). In contrast, none of the other subtypes of kidney neoplasms (e.g., clear-cell renal cell carcinoma) harbored PIK3CA mutations (n = 87; P < 0.001). Immunohistochemical analysis of urothelial carcinoma samples found loss of PTEN protein expression (36.4%, n = 11) and elevation of phosphorylated mammalian target of rapamycin (mTOR; 63.6%, n = 11). To confirm the role of the PI3K/AKT pathway in urothelial carcinoma, we generated mice containing biallelic inactivation of Pten in the urogenital epithelia. These mice developed typical renal pelvic urothelial carcinomas, with an incidence of 57.1% in mice older than 1 year. Laser capture microdissection followed by PCR confirmed the deletion of Pten exons 4 and 5 in the animal tumor cells. Immunohistochemical analyses showed increased phospho-mTOR and phospho-S6K levels in the animal tumors. Renal lymph node metastases were found in 15.8% of the animals with urothelial carcinoma. In conclusion, we identified and confirmed an important role for the PI3K/AKT pathway in the development of urothelial carcinoma and suggested that inhibitors of this pathway (e.g., mTOR inhibitor) may serve as effective therapeutic agents. [Cancer Res 2009;69(21):8256–64]

We report the case of a 48-year-old female patient who had a Crohn's disease treated by corticosteroids. The patient developed severe cardiac failure, which was refractory to treatment with inotropic agents. At necropsy, examination of the heart revealed myocardial abscesses. On microscopic study, we diagnosed an aspergillar myocarditis. Aspergillar myocarditis is a rare and fatal localisation in disseminated aspergillosis. Diagnosis is difficult and treatment, usually initiated late, is ineffective.

The prognostic significance of microvessel density in ovarian cancer is still a matter of debate. Classically, the degree of vascularisation is assessed in areas of high vascular density (hot spots), considered as regions of increased probability of metastasis. Since ovarian tumours have a particular progression and dissemination behaviour, vascularisation outside hot spots may also contribute to their evolution.In the present study, the degree of tumour vascularisation was estimated both in whole histogical sections and in hot spots, in 235 patients with ovarian carcinoma, using fully automatic image analysis methods. Six parameters were estimated: mean microvessel density (MVD) and mean microvessel surface fraction (MSP) on the whole section, mean and maximum values of MVD and MSP inside hot spots (MVDHS1, MSPHS1 and MVDHS2, MSPHS2). Relationships between vascular parameters and clinicopathologic features were analysed.In stage III-IV patients multivariate analysis showed that stage IV disease (hazards ratio (HR)=1.72, p=0.001), post-surgical residual disease 1cm (HR=2.86, p0.001), upper MVD tercile (HR=1.45, p0.022) and medial MVDHS1 tercile (HR=1.36, p=0.060) retained an independent prognostic value upon overall survival.Our results suggest that quantification of blood vessels, both on the whole histological section and in hot spots might be helpful in evaluating prognosis in advanced ovarian carcinomas.

We evaluated the results, accuracy and clinical incidence of our standard procedure of percutaneous biopsy for solid renal masses.From March 1999 to April 2005, 119 percutaneous core biopsies of renal masses were performed. Biopsies were proposed when there was no formal evidence for a carcinoma diagnosis on computerized tomography.Benign lesions were diagnosed in 24 biopsies (20.1%), including oncocytoma in 13, angiomyolipoma in 5 and chronic pyelonephritis in 5. Malignancy was identified in 70 biopsies (58.8%), including 57 renal carcinomas (conventional renal cell in 41, papillary in 12 and chromophobe in 4), 4 transitional cell carcinomas, 8 metastases and 1 lymphoma. For 25 biopsies (21%) no accurate diagnosis was possible, including 12 that showed inflammatory tissue and 13 with normal or necrotic tissue. These inconclusive biopsies prompted repeat biopsy in 13 patients, in whom a total of 11 malignant lesions were diagnosed. A total of 64 nephrectomies were performed with a biopsy accuracy for histopathological tumor type and Fuhrman nuclear grade of 86% and 46%, respectively. A period of watchful waiting was proposed for 31 patients (34.2%) and no renal malignancies were found. Computerized tomography showed stabilization or disappearance of the initial renal mass.Percutaneous renal tumor biopsies are safe, cost-effective and often conclusive for an acute histological diagnosis. This procedure could be decisive for choosing the optimal treatment, particularly to avoid nephrectomy for benign lesions. Biopsies should not be considered a routine procedure but they could be indicated when there is a lack of radiological evidence in elective patients.

Primary lymphoma of the breast (PBL) is a rare neoplasm, its outcome remains unclear compared to other lymphomas. We performed a retrospective study of 19 cases of high grade PBL. There were 17 Diffuse large B cell lymphoma (DLBCL) and 2 follicular and diffuse grade 3 lymphomas. Four patients were treated with local treatment only, 15 received chemotherapy including 11 treated with CHOP or ACVBP regimens followed by involved field radiotherapy. The actuarial survival for the whole population was 38%. Three of the 4 patients treated only with a local treatment died of their lymphoma. Three patients progressed on therapy and 5 relapsed in the first year of follow-up including 2 central nervous system recurrences. Among the 11 patients treated with chemotherapy, 2 died of their lymphoma. The overall survival of this subgroup was 73% (median follow-up of 57 months). We observed, like others in the literature, a better prognosis for lymphomas co-expressing Bcl6 and CD 10. The treatment should be based on the same modalities, but including a CNS prophylaxis even if poor prognosis factors are lacking. A radical mastectomy increases the risk of treatment failure and has to be avoided.

Medullary breast cancer (MBC) is a rare, diagnostically difficult, pathological subtype. Despite being high grade, it has a good prognosis. MBC patients have an excess of BRCA1 germ-line mutation and reliable identification of MBC could help to identify patients at risk of carrying germline BRCA1 mutations or in whom chemotherapy could be avoided. The aim of this study was therefore to improve diagnosis by establishing an MBC protein expression profile using immunohistochemistry (IHC) on tissue-microarrays (TMA). Using a series of 779 breast carcinomas ('EC' set), diagnosed initially as MBC, a double-reading session was carried out by several pathologists on all of the histological material to establish the diagnosis as firmly as possible using a 'medullary score'. Only MBCs with high scores, i.e. typical MBC (TMBC) (n=44) and non-TMBC grade III with no or low scores (n=160), were included in the IHC study. To validate the results obtained on this first set, a control series of TMBC (n=17) and non-MBC grade III cases (n=140) ('IPC' set) was studied. The expression of 18 proteins was studied in the 61 TMBCs and 300 grade III cases from the two sets. The global intra-observer concordance of the first reading for the diagnosis of TMBC was 94%, with almost perfect kappa (kappa) of 0.815. TMBC was characterized by a high degree of basal/myoepithelial differentiation. In multivariate analysis with logistic regression, TMBC was defined by the association of P-cadherin (R=2.29), MIB1 > 50 (R=3.80), ERBB2 negativity (R=2.24) and p53 positivity (RR=1.45).

The HER2 proto-oncogene encodes a transmembrane protein, which is considered to function as a growth factor receptor. Overexpression of this protein found by immunohistochemistry in about 20% of infiltrating breast carcinomas, has a predictive value of response to treatment by trastuzumab, an anti-HER2 humanized monoclonal antibody. Search for HER2 gene amplification is necessary to adapt the immunohistochemical technique quality and also in the cases of delicate analysis or weak overexpression. It is usually carried out by Fluorescence In Situ Hybridization (FISH). A more recent hybridization technique, named CISH because of its chromogenic revelation is an alternative method, which gives highly correlated results with FISH. We present details of this technique, which may be more familiar for the pathologists than FISH, because reading analysis is similar to that of immunohistochemical staining.

This report concerns a case of diabetic mastopathy, a benign but pseudotumoral clinicopathologic entity which occurs in young women with type 1 diabetes. Positive diagnosis, differential diagnosis and literature data are discussed.

BACKGROUND The goal of the current study was to investigate the prognostic impact of proliferative activity, together with the other classic clinicopathologic prognostic factors (tumor size, tumor grade, receptor status, ploidy, and lymph node status), in breast carcinoma by counting mitoses and evaluating S phase fraction (SPF) in fresh and frozen tumor samples. METHODS From March 1, 1990, to July 1, 1999, a total of 1984 previously untreated invasive breast carcinoma samples were snap-frozen for flow cytometry. RESULTS After multivariate analysis incorporating all classic prognostic factors, SPF combined with mitotic activity (i.e., proliferative activity) remained the sole prognostic factor in the lymph node–negative group; proliferative activity was accompanied by tumor size as a prognostic factor in patients with lymph node–positive disease and by lymph node status, lymphatic invasion, and receptor status in the overall population. The predictive value of proliferative activity was superior to that of the reference standards (classic prognostic predictors according to the guidelines of our institution [common oncology practice] and the St. Gallen classification). A review of the literature, focusing on series in which fresh material was used, allowed us to demonstrate that there is widespread agreement regarding the correlation between SPF and prognosis, even after multivariate analysis. CONCLUSIONS S phase fraction is a valuable predictor of survival and can confidently be assessed in approximately 80% of cases. In conjunction with mitotic activity, SPF should become a prognostic factor that is used in daily practice by oncologists for the management of breast carcinoma. Cancer 2004. © 2003 American Cancer Society.

The aim of this study was to assess the levels of cell cycle regulatory proteins p21waf1 (p21), p53, Cyclin A, Cyclin D1 and Ki-67 to see whether they correlated with recurrence-free survival (RFS). From 1982 to 1996, 50 patients aged less than 51 years underwent lumpectomy followed by radiotherapy for a pure ductal carcinoma in situ (DCIS). For each case, the following immunohistochemical stains were carried out: Ki-67, Cyclin A, Cyclin D1, p53 and p21waf1 (p21). The percentage of positive nuclei was assessed. Multiple combinations of these factors were performed; in particular, we called the sum of Ki-67 and Cyclin A a global proliferation factor (GPF). Correlations with classical clinicopathological data were assessed. After a multivariate analysis, only GPF, Van Nuys Prognostic Index (VNPI) grade and mitotic index were independent predictive factors of recurrence in the whole population. In the population with close surgical margins, when the GPF level was less than the 25th percentile or more than the 75th percentile recurrence was low. In this preliminary study, GPF seems to be of interest to help in the decision process in the post-surgical management of the patient.

BACKGROUND Between January 11, 1991 and January 8, 1992, 104 patients with previously untreated, invasive, primitive breast carcinoma were admitted to the authors' hospital. METHODS For each patient, flow cytometry DNA analyses on frozen samples and on immunohistochemical staining were performed, including Ki-67, cyclin A, p53, and p21waf1 (p21), with assessment of the percentages of positive nuclei were assessed. Correlations with classic clinicopathologic data and survival (overall, metastasis free, or recurrence free) and a multivariate analysis were performed. RESULTS After a multivariate analysis according to a Cox model that was stratified by age, tumor size, tumor grade, lymph node status, and receptor status, among the factors studied, the presence of p21 was the unique remaining prognostic factor for patients with invasive breast carcinoma. Because of the lack of a correlation between p21 and proliferative factors (Ki-67, S-phase, and cyclin A), the authors combined p21 with those markers and found that, for the different combinations, after statistical analysis, only p21 combined with S-phase or with cyclin A and lymph node status were salient survival prognostic factors. CONCLUSIONS Immunohistochemical study of proteins involved in the cell cycle and assessment of proliferative activity using flow cytometric DNA analysis aided the authors in singling out correlations of cyclin A and S-phase, along with p21, with metastasis free survival and overall survival in patients with invasive breast carcinoma. These promising results will require confirmation in a larger series of patients. Cancer 2003;97:1376–86. © 2003 American Cancer Society. DOI 10.1002/cncr.11209

Determination of the HER2/neu (HER2) status in breast carcinoma has become necessary for the selection of breast cancer patients for trastuzumab therapy. Amplification of the gene analysed by fluorescence in situ hybridisation (FISH) or overexpression of the protein determined by immunohistochemistry (IHC) are the two major methods to establish this status. A strong correlation has been previously demonstrated between these two methods. However, FISH is not always feasible in routine practice and weakly positive IHC tumours (2+) do not always correspond to a gene amplification. Our study was performed in order to evaluate the contribution of chromogenic in situ hybridisation (CISH), which enables detection of the gene copies through an immunoperoxidase reaction. CISH was performed in 79 breast carcinomas for which the HER2 status was previously determined by IHC and FISH. The results of IHC, FISH and CISH were compared for each tumour. CISH procedures were successful in 95% of our cases. Whatever the IHC results, we found a very good concordance (96%) between CISH and FISH. Our study confirms that CISH may be an alternative to FISH for the determination of the gene amplification status in 2+ tumours. Our results allow us to think that, in many laboratories, CISH may also be an excellent method to calibrate the IHC procedures or, as a quality control test, to check regularly that the IHC signal is in agreement with the gene status.

The "Standards, Options and Recommendations" (SOR) collaborative project was initiated in 1993 by the Federation of the French Cancer Centres (FNCLCC), with the 20 French Regional Cancer Centres, several French public university and general hospitals, as well as private clinics and medical specialty societies. Its main objective is the development of serviceable clinical practice guidelines in order to improve the quality of health care and the outcome of cancer patients. The methodology is based on a literature review, followed by critical appraisal by a multidisciplinary group of experts. Draft guidelines are produced, then validated by specialists in cancer care delivery.Produce clinical practice guidelines for the management and shipment of histological and cytopathological cancer specimens using the methodology developed by the Standards, Options and Recommendations project.The FNCLCC designated the group of experts. Available data were collected by a search of Medline and lists selected by experts in the group. A first draft of the guidelines was written, they validated by independent reviewers.The main recommendations are: 1) high-quality transmission of information between professionals is essential to the management of cancer specimens in order to assure high-quality diagnosis and evaluation of prognostic factors; 2) written procedures concerning sample shipment, handling, storage, registration, tracking and fixation exist; these procedures, as well as the necessary shipping material, will be sent to all clinical services involved; 3) when possible, fresh, unfractionated, oriented surgical samples will be submitted to the same histological and cytopathological laboratory; 4) samples collected for extemporaneous examination, freezing or cell culture must be shipped immediately under appropriate storage conditions; 5) Once frozen, samples can be stored in a deep freezer at temperatures of 80 C or below, or kept in liquid nitrogen; 6) fixing tissues shortly after sample collection is essential to prevent cell lysis; 7) computerised systems will be used to assure correct specimen registration and tracking in histological and cytopathological laboratories.

During a both retrospective and prospective study of thyroid cancers treated in the Basse Normandie between 1960 and 1999, we have identified 32 patients with thyroid lymphoma. The correct diagnosis was made initially in 69% of all cases. In the other cases, the diagnosis was secondarily corrected after review of the pathological material. According to the REAL classification, 7 (21%) corresponded to low grade MALT lymphomas, 2 to low grade lymphomas, 10 to high grade MALT Lymphomas and 10 (31%) to high grade lymphomas, one plasmocytoma and two unclassified lymphomas. According to the Ann Arbor classification, stage was IE for 56%, IIE for 19%, IIIE for 3% and 9% for IV. Median survival was 28 months with a mean at 61 months. 20 patients died (62%), 12 from the lymphoma and 8 from intercurrent causes. The overall survival at 5 years was 36% (9 5% CI 16 54%). A comparison of our results with those of the literature was performed.

In an extension of a previous in vitro study [Carreiras et al., Int. J. Cancer 63, 530-536 (1995)] and in an effort to understand the adhesive interactions mediated by integrins within epithelial ovarian tumors, the presence of the alpha v and beta 3 subunits and that of vitronectin (Vn) in ovarian carcinomas at various stages of differentiation and in normal ovarian epithelium were comparatively investigated. The study was performed on material from 34 patients. By immunofluorescence, cryostat sections were analyzed for their expression of alpha v (34 cases), beta 3 (19 cases), and Vn (29 cases). alpha v was expressed in normal epithelium and in highly differentiated tumors as well as in a majority of moderately and poorly differentiated carcinomas with identical staining pattern. beta 3 subunit and Vn were also expressed in normal cases and highly differentiated carcinomas. However, they were lacking in most of the less differentiated tumors. The analysis of cases which were simultaneously tested for the presence of alpha v, beta 3, and Vn revealed that a large proportion of normal ovarian epithelium and highly differentiated tumors simultaneously expressed alpha v, beta 3, and Vn; in contrast, in all moderately and poorly differentiated carcinomas either beta 3 or Vn was absent. The potential role of the alpha v beta 3/Vn system in ovarian epithelium functions is discussed. It is also speculated that modifications of this system in ovarian carcinomas might contribute to tumor progression.

543 Background: Several reports suggest an inter-observer variability in Ki67 assessment. Nevertheless, there is no large study that evaluates the rate of discrepancies, together with their impact. Methods: Ki67 expression was assessed on 663 samples from patients with ER-positive breast cancers included in the PACS01 trial (Roche, J Clin Oncol, 2006). Ki67 staining was done using MiB1 antibody (Dako, Copenhagen, Denmark, Dilution 1:250). Prognostic and predictive values have been reported previously (Penault-Llorca, J Clin Oncol, 2009). A second central review was done by a senior breast pathologist from a French Cancer center. A discrepancy was defined as either a false positive or false negative result. Cut-off for positivity was defined at 15% according to data from Cheang et al (JNCI, 2009). Results: The rate of discrepancy was correlated with the percentage of stained tumor cells. A 10% discrepancy rate between the 2 pathologists was observed when the first pathologist reported 30%. We then evaluated the impact of discrepancy in terms of prognosis. Patients presenting a concordant result between the two pathologists showed a better outcome as compared to patients presenting a discrepancy, independently to the percentage of tumor stained (p=0.05, patients with concordant Ki67 ranged between 10-30% versus those with one reader 10%-30%. A Ki67 ranged between 10 and 30% could define a grey zone in which Ki67 should be reported with caution or be double checked by another pathologist. Survival analysis suggested that inter-observer discrepancies could act a prognostic factor. Such finding could reflect underlying intra-tumor heterogeneity.

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