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Silvana Pilotti

Publications and source records attributed to Silvana Pilotti.

At least 19 recordsLinked to original sources
PubMed2006

High-risk human papillomavirus affects prognosis in patients with surgically treated oropharyngeal squamous cell carcinoma.

PURPOSE: Human papillomavirus (HPV) DNA tumors actively integrating the E6 and E7 oncogenes have a distinct biologic behavior resulting in a more favorable prognosis. To which extent the viral integration by itself, and/or the associated wild-type (wt) TP53 status, and/or a functional p16 contribute to prognosis is unclear. PATIENTS AND METHODS: To clarify how the presence of high-risk (HR) -HPV, TP53, and p16INK4a status interact with clinical outcome, we considered a retrospective series of 90 consecutive oropharyngeal cancer patients treated primarily with surgery. RESULTS: Seventeen (19%) patients showed integrated HPV 16 DNA (HPV positive), wt TP53 in all but two patients, normal p16INK4a in 15 assessable patients, and p16 expression in all 17 patients. Thirty-five patients (39%), two of whom were HPV positive, harbored TP53 mutations. p16INK4a deletion and p16 null immunophenotype occurred in 28 and 58 patients, respectively, and was similarly distributed in both patients with mutated TP53 (48% and 82%, respectively) and in patients with wt TP53 (46% and 77%, respectively). Statistical analysis showed that HPV-positive status significantly affects all investigated end points: overall survival (P = .0018), incidence of tumor relapse (P = .0371), and second tumor (P = .0152), whereas TP53 and p16INK4a status and p16 expression were not prognostic by themselves. CONCLUSION: Our molecular and clinical results are in agreement with previous findings but provide additional information into the biologic mechanisms involved in HR-HPV oropharyngeal cancer in comparison to HPV-negative tumors. According to the reduced risk of relapse and second tumors associated with HR-HPV positivity of oropharyngeal cancer, the therapeutic strategy and follow-up procedures should be reviewed.

Adult↗
PubMed2006

Molecular and biochemical analyses of platelet-derived growth factor receptor (PDGFR) B, PDGFRA, and KIT receptors in chordomas.

PURPOSE: We have previously shown the presence of an activated platelet-derived growth factor (PDGF) receptor (PDGFR) B and its ligand PDGFB in a limited number of patients with clinical and radiological responses to imatinib mesylate treatment. This article describes the results of comprehensive molecular/biochemical analyses of the three receptors targeted by the drug (PDGFRB, PDGFRA, and KIT) in a series of 31 chordoma patients. EXPERIMENTAL DESIGN: The presence and activation status of PDGFRB, PDGFRA, and KIT receptors were investigated by means of immunoprecipitation and Western blot analyses complemented by immunohistochemistry, their expression level was analyzed by means of real-time PCR, and the occurrence of activating point mutations was investigated by means of cDNA sequencing. The PDGFB, PDGFA, and stem cell factor cognate ligands were investigated by reverse transcription-PCR, and gene status was assessed by fluorescence in situ hybridization. RESULTS: The results show that PDGFRB was highly expressed and phosphorylated, whereas PDGFRA and KIT were less expressed but phosphorylated and thus activated. These findings, together with the absence of gain-of-function mutations and the presence of the cognate ligands, strongly support the hypothesis that the activation mechanism is the autocrine/paracrine loop. No role seems to be played by gene amplification. CONCLUSIONS: In the light of our findings, the clinical benefit observed in chordoma patients treated with imatinib seems to be attributable to the switching off of all three receptors.

Adult↗
PubMed2006

Molecular and cytogenetic subgroups of oropharyngeal squamous cell carcinoma.

PURPOSE: The aim of this study was to acquire further insights into the pathogenetic pathways of head and neck squamous cell carcinomas (HNSCC) that may be useful for identifying new biomarkers instrumental in developing more specific treatment approaches. EXPERIMENTAL DESIGN: Cell cycle regulators and epidermal growth factor receptor (EGFR) and BRAF genes were analyzed in a series of 90 oropharyngeal SCCs of a cohort of surgically treated patients from a single institution, and the results were matched with the presence of high-risk human papillomavirus (HR-HPV) DNA and the TP53 status. RESULTS: At least four distinct groups of tumors were identified sharing a common histology but displaying different molecular/cytogenetic patterns: (a) 19% were HPV-positive SCCs whose lack of alterations of the investigated genes could explain their particular natural history, which requires less aggressive treatment; (b) 37% were HPV-negative SCCs carrying TP53 mutations, which may be more effectively treated by drugs acting through p53-independent apoptosis; (c) 34% were HPV-negative SCCs carrying wild-type TP53 and loss of 9p21 (p16INK4a and p15INK4b) and/or cyclin D1 overexpression that justify treatment with DNA-damaging drugs followed by cell cycle inhibitors; and (d) 10% were HPV-negative lacking tumor suppressor genes and cell cycle alterations. The second, third, and fourth groups also showed an increased copy number of EGFR and chromosome 7 (43%) that might justify the additional or alternative use of EGFR inhibitors. CONCLUSIONS: Our findings suggest that assessing HPV, TP53, 9p21, and EGFR status may be crucial to finding more tailored and beneficial treatments for oropharyngeal SCCs.

Biomarkers, Tumor↗
PubMed2006

Malignant peripheral nerve sheath tumors: prognostic factors and survival in a series of patients treated at a single institution.

BACKGROUND: The authors explored the prognostic factors and clinical outcomes of patients who had malignant peripheral nerve sheath tumors (MPNST) with and without neurofibromatosis type 1 (NF-1). METHODS: Two hundred five patients with localized MPNST who underwent surgery at the Istituto Nazionale per lo Studio e la Cura dei Tumori (Milan, Italy) over 25 years were reviewed. Forty-six patients had concomitant NF-1 syndrome, and 159 patients did not. Local recurrence, distant metastases, and survival rates were studied. RESULTS: One hundred thirty patients presented with primary disease, and 75 patients had locally recurrent tumors. The disease-specific mortality rate was 43% at 10 years, with a continuously disease-free survival rate of no greater than 40%. Presentation with either primary or recurrent disease, tumor size, and tumor site (trunk vs. extremity) were the strongest independent predictors of survival. Margin status and radiation therapy also played a role, mostly related to their effect on local outcome. Pathologic grade influenced distant metastases, but only a trend for survival could be observed. No significant independent differences between patients with and without NF-1 were observed. CONCLUSIONS: To the authors' knowledge, this was among the largest single-institution series to date. The results confirmed that patients with MPNST share similar prognostic factors with patients who have other soft tissue sarcomas and have some of the worst clinical outcomes. The presence of NF-1 syndrome per se did not affect survival, but patients with NF-1 were more likely to have larger tumors. Therefore, such patients should be followed carefully to detect disease as early as possible.

Adolescent↗
PubMed2006

Telomere maintenance mechanisms in liposarcomas: association with histologic subtypes and disease progression.

Human cancer cells maintain telomeres by telomerase activity (TA) or by alternative lengthening of telomeres (ALT). We proposed to define the prevalence of the two telomere maintenance mechanisms (TMM), to assess their association with histology, and to compare their prognostic relevance in a series of 93 patients with liposarcoma. ALT was detected by assaying ALT-associated promyelocytic leukemia nuclear bodies and TA was assayed using the telomeric repeat amplification protocol. ALT or TA was found in 25.9% or 26.6% of 139 tested liposarcoma lesions, respectively. Three lesions were ALT+/TA+ whereas approximately 50% of lesions did not show any known TMM. TMM phenotype was consistent during disease progression. ALT was prevalent in dedifferentiated and in grade 3 liposarcomas whereas TA prevailed in most round-cell myxoid and in grade 2 liposarcomas. ALT and TA incidence was similar in primary and recurrent lesions whereas metastases were more frequently TA+ than ALT+ (59% versus 18%; P = 0.04). TMM presence negatively affected patient prognosis (P = 0.001): increased mortality was associated with positivity for TA (P = 0.038) or ALT (P < 0.0001) compared with TMM absence. ALT proved to be a stronger prognostic discriminant of increased mortality than TA even when adjusted for tumor location, grade, and histology (hazard ratio for cause-specific death, 3.58 versus 1.15). Our results indicate that ALT can support fully malignant liposarcomas and is associated with unfavorable disease outcome.

Adult↗
PubMed2006

Phenotype-genotype correlation: challenge of intestinal-type adenocarcinoma of the nasal cavity and paranasal sinuses.

BACKGROUND: Intestinal-type adenocarcinoma (ITAC) of the nasal cavity and paranasal sinuses shows microscopic features indistinguishable from colorectal cancer. Our aim was to verify whether the morphologic resemblances mirror genetic profile similarities. METHODS: Twenty consecutive surgically treated ITAC cases, previously investigated for p16(INK4a) and TP53, were investigated for hMLH1, hMSH2, and beta-catenin immunoreactivity, and for adenomatous polyposis coli (APC), K-ras, and BRAF gene mutations. RESULTS: One case was immunonegative for both hMLH1 and hMSH2, and 12 tumors (40%) revealed a strong beta-catenin overexpression. No BRAF and APC truncating mutations were identified, whereas K-ras mutations were detected in 9 ITACs (50%). CONCLUSIONS: Our data confirm the phenotypic similarities at the genetic level between colorectal cancer and ITACs showing deregulation of K-Ras/BRAF and loss of heterozygosity (LOH) of chromosome 18q. By contrast, both frequency rate and type of inactivation of the APC-beta-catenin pathway differ in the 2 tumors, suggesting different gatekeeper events in the early development of ITAC (p16(INK4a) and TP53) and colorectal cancer (APC).

Adaptor Proteins, Signal Transducing↗
PubMed2006

Functional analysis and molecular modeling show a preserved wild-type activity of p53(C238Y).

In human tumors, p53 is often disabled by mutations in its DNA-binding domain and is thus inactive as a transcription factor. Alternatively, MDM2 gene amplification or up-regulation represents a mechanism of p53 wild-type inactivation, mainly reported in soft tissue sarcomas. In a previous TP53 analysis carried out on sporadic and NF1-related malignant peripheral nerve sheath tumors, in two cases, we observed the occurrence of C238Y missense mutation, leading to p53 stabilization unexpectedly coupled with immunophenotypic MDM2 overexpression. To investigate this TP53 missense mutation not yet functionally characterized in mammalian cell, we did MDM2 Southern blot and p53(C238Y)/MDM2 biochemical and functional analyses followed by molecular modeling. The results showed a lack of MDM2 gene amplification, evidence of p53-MDM2 protein complexes, and presence of a p53 that retains the ability to become phosphorylated on Ser15 and to induce the transcription of p21(waf1). Additional molecular modeling data highlighted the structural similarities between p53(C238Y) and wild-type p53, further supporting that the p53(C238Y) mutant still retains functional wild-type p53 properties.

Adult↗
PubMed2006

Quantitative analysis of plasma DNA in colorectal cancer patients: a novel prognostic tool.

Extracellular DNA in the plasma or serum of cancer patients has been recently proposed as a source of analyzable cancer-related gene sequences (qualitative approach). Furthermore, patients with different tumor types show high levels of cell-free circulating DNA both in plasma and serum (quantitative approach) at the time of surgery. Our aim was to verify whether the level of cell-free DNA in plasma might help in detecting recurrences during follow-up of colorectal cancer (CRC) patients. We studied 70 patients undergoing surgery for primary CRC. Plasma samples were obtained at the time of surgery and during follow-up. The cell-free circulating DNA in plasma was quantified by the Dipstick Kit method. At the time of surgery, in all patients, cell-free DNA levels in plasma were about 25 times higher in comparison with 20 healthy donors. In contrast, the carcinoembryonic antigen (CEA) value of this cohort of patients was altered in only about 37% of cases. During follow-up, cell-free DNA levels decreased progressively in tumor-free patients, while it increased in those developing recurrences or metastases. The results were further supported by qualitative analysis of circulating tumor-specific DNA, such as K-Ras mutations and p16(INK4a) promoter hypermethylation. These preliminary data confirm that plasma tumor DNA levels (i) are significantly higher in patients with CRC, (ii) decrease progressively in the follow-up period in tumor-free patients, and (iii) increase in patients with recurrence or metastasis. We suggest, therefore, that the quantification of plasma cell-free DNA might represent a useful tool for monitoring of CRC and, prospectively, for identifying high-risk individuals.

Colorectal Neoplasms↗
PubMed2005

Induction of a proinflammatory program in normal human thyrocytes by the RET/PTC1 oncogene.

Rearrangements of the RET receptor tyrosine kinase gene generating RET/PTC oncogenes are specific to papillary thyroid carcinoma (PTC), the most frequent thyroid tumor. Here, we show that the RET/PTC1 oncogene, when exogenously expressed in primary normal human thyrocytes, induces the expression of a large set of genes involved in inflammation and tumor invasion, including those encoding chemokines (CCL2, CCL20, CXCL8, and CXCL12), chemokine receptors (CXCR4), cytokines (IL1B, CSF-1, GM-CSF, and G-CSF), matrix-degrading enzymes (metalloproteases and urokinase-type plasminogen activator and its receptor), and adhesion molecules (L-selectin). This effect is strictly dependent on the presence of the RET/PTC1 Tyr-451 (corresponding to RET Tyr-1062 multidocking site). Selected relevant genes (CCL20, CCL2, CXCL8, CXCR4, L-selectin, GM-CSF, IL1B, MMP9, UPA, and SPP1/OPN) were found up-regulated also in clinical samples of PTC, particularly those characterized by RET/PTC activation, local extrathyroid spread, and lymph node metastases, when compared with normal thyroid tissue or follicular thyroid carcinoma. These results, demonstrating that the RET/PTC1 oncogene activates a proinflammatory program, provide a direct link between a transforming human oncogene, inflammation, and malignant behavior.

Blotting, Western↗
PubMed2005

Prediction of BRCA1 status in patients with breast cancer using estrogen receptor and basal phenotype.

PURPOSE: To investigate the proportion of breast cancers arising in patients with germ line BRCA1 and BRCA2 mutations expressing basal markers and developing predictive tests for identification of high-risk patients. EXPERIMENTAL DESIGN: Histopathologic material from 182 tumors in BRCA1 mutation carriers, 63 BRCA2 carriers, and 109 controls, collected as part of the international Breast Cancer Linkage Consortium were immunohistochemically stained for CK14, CK5/6, CK17, epidermal growth factor receptor (EGFR), and osteonectin. RESULTS: All five basal markers were commoner in BRCA1 tumors than in control tumors (CK14: 61% versus 12%; CK5/6: 58% versus 7%; CK17: 53% versus 10%; osteonectin: 43% versus 19%; EGFR: 67% versus 21%; P < 0.0001 in each case). In a multivariate analysis, CK14, CK5/6, and estrogen receptor (ER) remained significant predictors of BRCA1 carrier status. In contrast, the frequency of basal markers in BRCA2 tumors did not differ significant from controls. CONCLUSION: The use of cytokeratin staining in combination with ER and morphology provides a more accurate predictor of BRCA1 mutation status than previously available, that may be useful in selecting patients for BRCA1 mutation testing. The high percentage of BRCA1 cases positive for EGFR suggests that specific anti-tyrosine kinase therapy may be of potential benefit in these patients.

Biomarkers, Tumor↗
PubMed2005

9p21 locus analysis in high-risk gastrointestinal stromal tumors characterized for c-kit and platelet-derived growth factor receptor alpha gene alterations.

BACKGROUND: Gastrointestinal stromal tumors (GISTs) are noncomplex sarcomas that often are due to c-kit-activating and platelet-derived growth factor receptor alpha gene (PDGFRalpha)-activating mutations and perturbations of their related signaling pathways. Molecular and cytogenetic findings have indicated correlations between tumor progression and high-risk GISTs with c-kit mutations, the overexpression of genes such as ezrin, and losses at 9p. In particular, it was reported recently that malignant GISTs showed alterations in the p16INK4a gene located at the 9p21 locus. METHODS: To assess the involvement of p14ARF and p15INK4b in addition to p16INK4a in GISTs, the authors undertook a molecular and cytogenetic study of the 9p21 locus. A series of 22 pre-Gleevec era, cryopreserved, high-risk GISTs that were characterized well in terms of KIT and PDGFRalpha receptors were investigated for mRNA expression, homozygous deletions, mutations, and promoter methylation of locus 9p21, in some instances complemented by fluorescent in situ hybridization studies. RESULTS: The results indicated the loss of p16INK4a mRNA expression in 41% of the GISTs, mainly due to the homozygous deletion of both the p16INK4a gene and the p14ARF gene (24%). No mutations were found, and promoter methylation (detected by means of methylation-specific polymerase chain reaction analysis in 27% of tumors) was restricted mainly to the p15INK4b gene (20%). It is noteworthy that, in all of the methylated GISTs, the epigenetic promoter alteration was coupled with mRNA expression. CONCLUSIONS: Alterations in the 9p21 locus were found cumulatively in 54% of the tumors in the current series and were represented mainly by the loss of tumor suppressor gene expression. The p16INK4a deletion, which always was coupled with p14ARF gene loss, seemed to be the most common 9p21 inactivation mechanism.

Adult↗
PubMed2005

Isolation and in vitro propagation of tumorigenic breast cancer cells with stem/progenitor cell properties.

Breast cancer-initiating cells have been recently identified in breast carcinoma as CD44+/CD24(-/low) cells, which exclusively retain tumorigenic activity and display stem cell-like properties. However, at present, direct evidence that breast cancer-initiating cells can be propagated in vitro is still lacking. We report here the isolation and in vitro propagation of breast cancer-initiating cells from three breast cancer lesions and from an established breast carcinoma cell line. Our breast carcinoma-derived cultures encompassed undifferentiated cells capable of self-renewal, extensive proliferation as clonal nonadherent spherical clusters, and differentiation along different mammary epithelial lineages (ductal and myoepithelial). Interestingly, cultured cells were CD44+/CD24- and Cx43-, overexpressed neoangiogenic and cytoprotective factors, expressed the putative stem cell marker Oct-4, and gave rise to new tumors when as few as 10(3) cells were injected into the mammary fat pad of SCID mice. Long-term cultures of breast tumorigenic cells with stem/progenitor cell properties represent a suitable in vitro model to study breast cancer-initiating cells and to develop therapeutic strategies aimed at eradicating the tumorigenic subpopulation within breast cancer.

Animals↗
PubMed2005

SMARCB1/INI1 tumor suppressor gene is frequently inactivated in epithelioid sarcomas.

Epithelioid sarcoma is a rare soft tissue neoplasm of uncertain lineage that usually arises in the distal extremities of adults, presents a high rate of recurrences and metastases and frequently poses diagnostic dilemmas. The recently reported large-cell "proximal-type" variant is characterized by increased aggressiveness, deep location, preferential occurrence in proximal/axial regions of older patients, and rhabdoid features. Previous cytogenetic studies indicated that the most frequent alterations associated with this tumor entity affect chromosome 22. In this study, combined spectral karyotyping, fluorescence in situ hybridization, and array-based comparative genomic hybridization analyses of two proximal-type cases harboring a rearrangement involving 10q26 and 22q11 revealed that the 22q11 breakpoints were located in a 150-kb region containing the SMARCB1/INI1 gene, and that homozygous deletion of the gene was present in the tumor tissue. The SMARCB1/INI1 gene encodes for an invariant subunit of SWI/SNF chromatin remodeling complex and has been previously reported to act as a tumor suppressor gene frequently inactivated in infantile malignant rhabdoid tumors. We analyzed SMARCB1/INI1 gene status in nine additional epithelioid sarcoma cases (four proximal types and five conventional types) and altogether we identified deletions of SMARCB1/INI1 gene in 5 of 11 cases, all proximal types. We confirmed and further extended the number of cases with SMARCB1/INI1 inactivation to 6 of 11 cases, by real-time quantitative PCR analysis of mRNA expression and by SMARCB1/INI1 immunohistochemistry. Overall, these results point to SMARCB1/INI1 gene involvement in the genesis and/or progression of epithelioid sarcomas. Analysis of larger series of epithelioid sarcomas will be necessary to highlight putative clinically relevant features related to SMARCB1/INI1 inactivation.

Adult↗
PubMed2005

Lack of telomerase activity in lung carcinoids is dependent on human telomerase reverse transcriptase transcription and alternative splicing and is associated with long telomeres.

PURPOSE: Preliminary evidence indicates that telomerase activity is significantly less expressed in typical carcinoids than in large cell neuroendocrine carcinomas or in small cell lung cancers. Knowledge of the mechanisms by which telomerase is differentially regulated in neuroendocrine lung tumors is important for a better understanding of the pathogenesis of these malignancies. EXPERIMENTAL DESIGN: We investigated telomerase activity in 86 neuroendocrine lung tumors and correlated the enzyme activity with the expression of the enzyme subunits [human RNA component (hTR), human telomerase reverse transcriptase (hTERT), and alternatively spliced hTERT variants], with the telomere-associated protein human protection of telomere-1, and with the telomere length pattern. RESULTS: A significantly (P = 0.0001) lower frequency of telomerase-positive cases was found in typical carcinoids (14%) than in large cell neuroendocrine carcinomas (87%) and small cell lung cancers (92%). hTR was constitutively expressed in all carcinoids. Telomerase-negative carcinoids were characterized by the absence of any hTERT transcript, only displayed the beta(-) alternatively spliced variant, or concomitantly expressed the alpha(+)beta(+) full-length message with different combinations of alternatively spliced variants. However, in these tumors, a more abundant level of alternatively spliced transcripts than that of the alpha(+)beta(+) full-length transcript was generally found. No significant difference was observed in human protection of telomere-1 expression between telomerase-negative and telomerase-positive carcinoids. Telomeres were significantly (P < 0.05) longer in telomerase-negative carcinoids than in telomerase-positive carcinoids (median value, 9.15 versus 4.47 kb). However, alternative lengthening of telomeres, as shown by associated promyelocytic leukemia bodies, was not observed in these tumors. CONCLUSIONS: Our results indicate that telomerase is repressed in most lung carcinoids and that hTERT transcription and alternative splicing play a role in such a negative regulation. Moreover, the absence of any telomerase maintenance mechanism may contribute to the favorable prognosis of this malignancy.

Alternative Splicing↗