TRK-A, HER-2/neu, and KIT Expression/Activation Profiles in Salivary Gland Carcinoma.

Salivary duct carcinomas (SDCs) and adenoid cystic carcinomas (ACCs) are the most aggressive and the most frequent carcinomas of the salivary glands, respectively. Little is known about them in terms of molecular/biochemical characterization and conventional treatments are ineffective. On cryopreserved material, we analyzed the expression/activation status of TRK-A, HER-2/neu, and KIT receptors by means of immunoprecipitation and Western blot analysis experiments, and the presence of their cognate ligands by means of Western blot analysis and/or reverse transcription-polymerase chain reaction in 9 SDCs, 12 ACCs, and 8 normal glands. The amplification status of HER-2/neu was also investigated by means of fluorescent in situ hybridization analysis on fixed material. The receptor tyrosine kinase (RTK)-deregulated profile of the SDCs was characterized by the overexpression of activated TRK-A in the presence of its ligand, and the overexpression of HER-2/neu sustained by gene amplification. The RTK signature of the ACCs was represented by the overexpression of activated KIT and TRK-A and their cognate ligands, and the overexpression of activated HER-2/neu, in the absence of gene amplification, possibly sustained by epidermal growth factor receptor heterodimerization. In conclusion, SDCs and ACCs, although sharing TRK-A autocrine loop activation, have different pathologically activated RTK-deregulated profiles that may be potential targets for pharmacological RTK inhibitors.

Subcellular localization of activated leukocyte cell adhesion molecule is a molecular predictor of survival in ovarian carcinoma patients.

PURPOSE: Currently available clinicopathologic prognostic factors are imperfect predictors of clinical course in advanced-stage epithelial ovarian cancer patients. New molecular predictors are needed to identify patients with higher risk of relapse or death from disease. In a retrospective study, we investigated the prognostic impact of activated leukocyte cell adhesion molecule (ALCAM) expression in epithelial ovarian cancer. EXPERIMENTAL DESIGN: We analyzed the effect of cell-anchorage loss on ALCAM cellular localization in vitro and assessed ALCAM expression by immunohistochemistry in a series of 109 well-characterized epithelial ovarian cancer patient samples. Chi-square test, Kaplan-Meier method, and Cox proportional hazard analyses were used to relate ALCAM cellular localization to clinical-pathologic parameters and to overall survival (OS) rate. RESULTS: Loss of epithelial ovarian cancer cell anchorage was associated both in vitro and in vivo with decreased ALCAM membrane expression. In vivo, ALCAM was localized to cell membrane in normal surface ovarian epithelium, whereas in 67% of the epithelial ovarian cancer samples, membrane localization was decreased or even lost, and the molecule was mainly expressed in cytoplasm. Median OS in this group of patients was 58 months, whereas a median OS was not yet reached in patients with ALCAM membrane localization (P = 0.036, hazard ratio [HR] = 2.0, 95% confidence interval [CI] 1.1 to 3.5). In a multivariate Cox regression model including all the available clinicopathologic variables, loss of ALCAM membrane expression was an independent factor of unfavorable prognosis (P = 0.042, HR = 2.15, 95% CI: 1.0 to 4.5). CONCLUSIONS: Decreased/lost ALCAM membrane expression is a marker of poorer outcome in epithelial ovarian cancer patients and might help to identify patients who could benefit from more frequent follow-up or alternative therapeutic modalities.

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.

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.

Detection of overexpressed and phosphorylated wild-type kit receptor in surgical specimens of small cell lung cancer.

PURPOSE: The combinations of various chemotherapeutic drugs currently used to treat advanced small cell lung cancer (SCLC) led to similarly poor survival outcomes, which is why new molecular biology approaches are needed to design and select targeted therapies. EXPERIMENTAL DESIGN: Thirteen stage I SCLC surgical specimens were screened for c-Kit gene mutations by sequencing whole cDNA and for KIT receptor expression/activation by immunoprecipitation and Western blotting. Both the paraffin-embedded and frozen materials were analyzed by immunocytochemistry, and the stem cell factor cognate ligand was assessed by retrotranscription PCR. RESULTS: In all cases, we showed the presence of wild-type KIT receptors by analyzing the entire coding sequence, which together with the detection of the cognate ligand stem cell factor, supports the establishment of an autocrine loop. In addition, the KIT receptor was activated/phosphorylated. The immunoprecipitation/Western blotting data fit the observed immunophenotype. Interestingly, comparison of the level of KIT expression was at least 10 times higher in the tumoral specimens than the normal reference lungs. CONCLUSIONS: The KIT molecular profile derived from the analysis of SCLC surgical specimens shows that wild-type KIT is overexpressed and phosphorylated in the presence of stem cell factor. This finding, which is consistent with pathological KIT activation driven by an autocrine loop, is particularly interesting in the light of the recent development of new tyrosine kinase inhibitory drugs, which are highly effective in blocking wild-type KIT receptors.