Variant Philadelphia translocations: molecular-cytogenetic characterization and prognostic influence on frontline imatinib therapy, a GIMEMA Working Party on CML analysis.

Variant Philadelphia (Ph) chromosome translocations have been reported in 5%-10% of patients with newly diagnosed chronic myeloid leukemia (CML). Variant translocations may involve one or more chromosomes in addition to 9 and 22, and can be generated by 2 different mechanisms, 1-step and 2-step rearrangements, as revealed by fluorescence in situ hybridization. The prognostic significance of the occurrence of variant translocations has been discussed in previous studies. The European LeukemiaNet recommendations do not provide a "warning" for patients with variant translocations, but there is limited information about their outcome after therapy with tyrosine kinase inhibitors. To identify the role of variant translocations in early chronic phase (CP) CML patients treated with imatinib mesylate, we performed an analysis in a large series of 559 patients enrolled in 3 prospective imatinib trials of the Gruppo Italiano Malattie EMatologiche dell'Adulto (GIMEMA) Working Party on CML. Variant translocations occurred in 30 patients (5%). Our data show that the presence of variant translocations has no impact on the cytogenetic and molecular response or on outcome, regardless of the involvement of different mechanisms, the number of involved chromosomes, or the presence of deletions. Therefore, we suggest that patients with variant translocations do not constitute a "warning" category in the imatinib era. This study is registered at www.clinicaltrials.gov as NCT00514488 and NCT00510926.

Chromosome abnormalities additional to the Philadelphia chromosome at the diagnosis of chronic myelogenous leukemia: pathogenetic and prognostic implications.

Additional chromosome abnormalities (ACAs) occur in less than 10% of cases at diagnosis of Philadelphia chromosome (Ph)-positive chronic myelogenous leukemia (CML). In some cases, on the basis of the persistence of the ACAs in Ph-negative cells after response to imatinib, a secondary origin of the Ph chromosome has been demonstrated. In this study, the possible prognostic value of this phenomenon was evaluated. Thirty-six Ph-positive CML patients were included in the study. In six patients, ACAs persisted after the disappearance of the Ph. A complete cytogenetic response (CCR) was obtained in five of these six patients, and five of six also had a high Sokal score. In all the other cases, ACAs disappeared together (in cases of response to therapy with imatinib) or persisted with the Ph (in cases of no response to imatinib). In the former cases, the primary origin of the Ph was demonstrated. CCR was obtained in 22 cases (17 with low to intermediate Sokal scores), while no response was observed in 8 patients (5 with a high Sokal score). Sokal score seems to maintain its prognostic value for patients in whom the Ph occurs as a primary event, but not in those in whom it occurs as a secondary one.

Endothelial NOS, estrogen receptor beta, and HIFs cooperate in the activation of a prognostic transcriptional pattern in aggressive human prostate cancer.

The identification of biomarkers that distinguish between aggressive and indolent forms of prostate cancer (PCa) is crucial for diagnosis and treatment. In this study, we used cultured cells derived from prostate tissue from patients with PCa to define a molecular mechanism underlying the most aggressive form of PCa that involves the functional activation of eNOS and HIFs in association with estrogen receptor beta (ERbeta). Cells from patients with poor prognosis exhibited a constitutively hypoxic phenotype and increased NO production. Upon estrogen treatment, formation of ERbeta/eNOS, ERbeta/HIF-1alpha, or ERbeta/HIF-2alpha combinatorial complexes led to chromatin remodeling and transcriptional induction of prognostic genes. Tissue microarray analysis, using an independent cohort of patients, established a hierarchical predictive power for these proteins, with expression of eNOS plus ERbeta and nuclear eNOS plus HIF-2alpha being the most relevant indicators of adverse clinical outcome. Genetic or pharmacologic modulation of eNOS expression and activity resulted in reciprocal conversion of the transcriptional signature in cells from patients with bad or good outcome, respectively, highlighting the relevance of eNOS in PCa progression. Our work has considerable clinical relevance, since it may enable the earlier diagnosis of aggressive PCa through routine biopsy assessment of eNOS, ERbeta, and HIF-2alpha expression. Furthermore, proposing eNOS as a therapeutic target fosters innovative therapies for PCa with NO inhibitors, which are employed in preclinical trials in non-oncological diseases.

Prognostic value of HER2 and progesterone receptor expression in endometrial carcinoma with positive peritoneal washing.

BACKGROUND: Although the majority of endometrial cancer (EC) patients can be cured by surgery, unexpected recurrent disease may also occur in early stage patients. In the present study, whether or not the analysis of multiple biopathological parameters might lead to more accurate predictions of the clinical outcome of EC patients with long-term follow-up (FU) was investigated. PATIENTS AND METHODS: Estrogen and progesterone receptor (ER and PgR) positivity and HER2 overexpression by immunohistochemistry were evaluated. The peritoneal washings (PWs) were analyzed by cytology and immunocytochemistry employing AR-3 and B72.3 monoclonal antibodies. RESULTS: The patients with positive PW and HER2 positive tumors showed shorter overall survival compared to those bearing HER2 negative tumors (p =0.004). HER2 overexpression also influenced the patient outcome in the group with tumors lacking PgR (p = 0.004). At multivariate analysis PgR and HER2 overexpression emerged as independent prognostic factors. CONCLUSION: The combined analysis of these biopathological markers could provide useful information for the selection of patients to be enrolled in innovative therapeutic strategies.

Chromogenic in situ hybridization to detect HER-2/neu gene amplification in histological and ThinPrep-processed breast cancer fine-needle aspirates: a sensitive and practical method in the trastuzumab era.

The increasing evidence of trastuzumab efficacy in breast cancer (BC) patients means that an accurate and reproducible evaluation of HER-2 statusis of paramount importance in histological and in cytological samples. Currently, the two main methods used to analyze HER-2 amplification or overexpression are fluorescence in situ hybridization (FISH) and immunohistochemistry (IHC). Although the two methods are strongly correlated for histological tissue, the evaluation of tumor morphology through FISH may be difficult and fluorescence fades quickly. These limitations can be overcome by chromogenic in situ hybridization (CISH), which can visualize the amplification product along with morphological features. In view of this, in the present study, we analyzed the usefulness of CISH on formalin-fixed, paraffin-embedded (FFPE) BC specimens and investigated whether CISH can be a valid technique in the determination of HER-2 status for fine-needle aspirates (FNAs) processed by liquid-based cytology. The results we obtained in a retrospective series of 111 FFPE BC specimens demonstrated good concordance between CISH and IHC and between CISH and FISH. The former concordance was comparable with that observed between FISH and IHC. When CISH was applied to a prospective series of 53 FNAs, from surgically removed BC, our data showed evidence of a higher concordance of results between liquid-based cytology and the companion FFPE tissues using CISH rather than HercepTesttrade mark. Therefore, CISH analysis, which is avaluable and reproducible alternative to FISH for selecting breast cancer patients for trastuzumab therapy, can lower false-positive immunocytochemistry findings in ThinPrep-processed FNAs.

Epithelial-restricted gene profile of primary cultures from human prostate tumors: a molecular approach to predict clinical behavior of prostate cancer.

The histopathologic and molecular heterogeneity of prostate cancer and the limited availability of human tumor tissue make unraveling the mechanisms of prostate carcinogenesis a challenging task. Our goal was to develop an ex vivo model that could be reliably used to define a prognostic signature based on gene expression profiling of cell cultures that maintained the tumor phenotype. To this end, we derived epithelial cultures from tissue explanted from 59 patients undergoing radical prostatectomy or cistoprostatectomy because of prostate benign hyperplasia/prostate cancer or bladder carcinoma. Patient selection criteria were absence of hormonal neoadjuvant treatment before surgery and diagnosis of clinically localized disease. Using this unique experimental material, we analyzed expression of 22,500 transcripts on the Affymetrix Human U133A GeneChip platform (Affymetrix, Inc., High Wycombe, United Kingdom). Cultures from normal/hyperplastic tissues with a prevalent luminal phenotype and from normal prostate epithelial tissue with basal phenotype (PrEC) served as controls. We have established a large number of prostate primary cultures highly enriched in the secretory phenotype. From them, we derived an epithelial-restricted transcriptional signature that (a) differentiated normal from tumor cells and (b) clearly separated cancer-derived lines into two distinct groups, which correlated with indolent or aggressive clinical behavior of the disease. Our findings provide (a) a method to expand human primary prostate carcinoma cells with a luminal phenotype, (b) a powerful experimental model to study primary prostate cancer biology, and (c) a novel means to characterize these tumors from a molecular genetic standpoint for prognostic and/or predictive purposes.

Pegylated liposomal doxorubicin in combination with gemcitabine: a phase II study in anthracycline-naïve and anthracycline pretreated metastatic breast cancer patients.

BACKGROUND: The aim of the study was to assess the toxicity profile, activity in terms of response rate, time to progression, overall survival, and quality of life of pegylated liposomal doxorubicin (PLD) and gemcitabine combination in chemo-naïve and pretreated metastatic breast cancer (MBC) women. METHODS: Patients were eligible if they had disease progression to prior chemotherapy (anthracycline-including or not) for early breast cancer or MBC. Patients received PLD 25 mg/m(2) intravenously on day 1 plus gemcitabine 800 mg/m(2) intravenously on days 1 and 8 of each 21-day cycle. RESULTS: Of 50 patients enrolled, 37 had received prior adjuvant chemotherapy (24 with an anthracycline) and 23 prior chemotherapy for metastatic disease (6 with an anthracycline). Two complete responses and 20 partial responses were achieved in 46 assessable patients (overall response rate: 47.8%). Responses were observed in 14 (46.6%) of 30 patients with previous anthracycline exposure. Median response duration was 7 months, median duration of clinical benefit 8 months, time to progression 7 months. At a median follow-up of 10 months, 79.4% patients were alive at 1 year. No neutropenic complication was observed. Non-hematological toxicities were mild. One patient previously treated with an anthracycline developed a transient decrease (26%) in the left ventricular ejection fraction, with cardiac function recovering within 6 months. CONCLUSION: Because of the non-overlapping toxicity profiles of both PLD and gemcitabine, this combination can be regarded as a reliable therapeutic option for patients who have failed previous treatments, including anthracycline, for MBC.

Cytogenetic profiles as additional markers to pathological features in clinically localized prostate carcinoma.

Fluorescence in situ hybridization analysis for evaluation of 7, 8, X chromosomes and EGFR, LPL, MYC, AR genes in 79 neoplastic foci from 56 patients with clinically localized prostate cancer was performed. We found aneusomy for chromosome 7, 8 and X in 74/77 (96.1%), 56/76 (73.7%), 26/70 (37.1%) of examined foci respectively. No specimen was amplified for EGFR and AR genes, only 2/71 (2.8%) specimens showed MYC gene amplified. LPL deletion was present in 52/76 (68.4%) specimens. Statistically association between Gleason score and both chromosome 7 aneusomy and 8p21 deletion was present. The frequency of chromosome 7 aneusomy was statistically higher in T3-4 cases than T2c and T2a-T2b ones. We considered as unfavorable a genetic set if aneusomy for at least two chromosomes and one altered gene were present. The percentage of tumors, with unfavorable genetic pattern, increased from 36.4 to 75.0% in those with Gleason >7 and from 40.0 to 73.7% in those with stage T3 or more. These alterations could be considered potent genetic markers adjunctive to conventional prognostic parameters. Our objective was to establish specific genetic profiles which may discriminate favorable and unfavorable genetic prognosis tumors.

Complex variant Philadelphia translocation involving the short arm of chromosome 9 in a case of chronic myeloid leukemia.

Here we describe how we detected the BCR/ABL fusion gene on the short arm of der(9) combining classical GTG banding and Fluorescence In Situ Hybridization (FISH) in a case of chronic myeloid leukemia (CML). To our knowledge, variant translocations involving the short arm of chromosome 9, in literature, are almost rare in chronic myeloid leukemia. It is not clear if this complex genetic translocation represents clonal evolution or a unique, initial presentation variant of the Philadelphia chromosome (Ph).

Genetic instability in superficial bladder cancer and adjacent mucosa: an interphase cytogenetic study.

A systematic analysis of both tumors and the surrounding urothelium to help identify what lies behind the mechanism of multifocal tumor development has not yet been performed. In this study we investigated chromosome 1, 7, 9, and 17 aneusomy in 25 superficial papillary carcinomas and in 51 tissue samples taken from sites of macroscopically uninvolved urothelium surrounding the tumors, using the fluorescence in situ hybridization method. Our data demonstrated a close genetic relationship between all examined tumors and normal-appearing mucosa. Numeric aberrations of chromosomes 1, 7, 9, and 17 were found to exhibit similar patterns in all analyzed specimens, although with different frequencies.