Expression of FK506-binding protein 51 (FKBP51) in Mycosis fungoides.

BACKGROUND: Mycosis fungoides (MF) is the major subtype of cutaneous T-cell lymphomas (CTCL). It usually has a prolonged indolent clinical course with a minority of cases acquiring a more aggressive biological profile and resistance to conventional therapies, partially attributed to the persistent activation of nuclear factor-kappa B (NF-κB) pathway. In the last decade, several papers suggested an important role for the FK506-binding protein 51 (FKBP51), an immunophilin initially cloned in lymphocytes, in the control of NF-κB pathway in different types of human malignancies. OBJECTIVES: We aimed to investigate the possible value of FKBP51 expression as a new reliable marker of outcome in patients with MF. METHODS: We assessed by immunohistochemistry (IHC) FKBP51 expression in 44 patients with MF, representative of different stages of the disease. Immunohistochemical results were subsequently confirmed at mRNA level with quantitative PCR (qPCR) in a subset of enrolled patients. In addition, IHC and qPCR served to study the expression of some NF-κB-target genes, including the tumour necrosis factor receptor-associated factor 2 (TRAF2). RESULTS: Our results show that FKBP51 was expressed in all evaluated cases, with the highest level of expression characterizing MFs with the worst prognosis. Moreover, a significant correlation subsisted between FKBP51 and TRAF2 IHC expression scores. CONCLUSIONS: We hypothesize a role for FKBP51 as a prognostic marker for MF and suggest an involvement of this immunophilin in deregulated NF-κB pathway of this CTCL.

Overexpression of chromatin assembly factor-1/p60 predicts biological behaviour of laryngeal carcinomas.

This study analysed the immunohistochemical expression of the CAF-1/p60 protein in laryngeal cancers. CAF-1/p60 assumes an independent discriminative and prognostic value in laryngeal neoplasms; the presence of this protein in carcinoma in situ compared with laryngeal precancerous and larynx infiltrating tumours. We assessed the immunohistochemical expression of CAF-1/p60 in 30 cases of moderate and/or severe dysplasia, 30 cases of carcinoma in situ and 30 cases of laryngeal squamous cell carcinoma (LSCCs). CAF-1/p60 expression increased significantly according to the high index of neoplastic cellular replication; therefore, CAF-1/p60 was overexpressed in neoplastic cells and its moderate-severe expression is correlated with poorer prognosis compared to less expression. In conclusion, overexpression of the CAF-1/p60 protein is related to a risk of higher morbidity and mortality and is a reliable independent prognostic index of laryngeal carcinoma. CAF1-p60 protein overexpression can be used in cancer management as an indicator of malignant evolution, especially in carcinoma in situ.

An "ex vivo model" contributing to the diagnosis and evaluation of new drugs in cystic fibrosis.

Cystic fibrosis (CF) is an autosomal recessive disease caused by mutations in the cystic fibrosis transmembrane regulator (CFTR) gene. About 2000 mutations have been described so far. We setup an ex vivo model of human nasal epithelial cells (HNECs) to study CF patients testing the effect of novel mutations and molecular therapies. We performed sampling (by brushing), followed by culture and analysis of HNECs using a series of molecular techniques. We performed 50 brushings from CF patients and controls. Using cultured cells, we: i) demonstrated the widely heterogeneous CFTR expression in patients and in controls; ii) defined the splicing effect of a CFTR mutation; iii) assessed the CFTR gating activity in patients bearing different mutations; iv) demonstrated that butyrate significantly enhances CFTR expression. Based on our data, we can conclude: 1) HNEC brushing is performed without anaesthesia and is well tolerated in all CF patients (children and adults); 2) HNECs can be preserved for up to 48 hours before culture allowings multicentre studies; 3) HNECs culture can be considered a suitable model to study the molecular effects of new CFTR gene mutations and/or uncertain meaning specific mutations of carriers; 4) an ex vivo model of HNECs may be used to evaluate, before human use, the effect of new drugs on patients' cells bearing specific CFTR mutations; 5) the methodology is adequate for a quantitative measurement, by fluorescence, of the CFTR gating activity of the HNECs from patients with different genotypes identifying: a) CF patients bearing two severe mutations with an activity < 10% (compared to controls - 100%); b) CF patients bearing at least a mild mutation with an activity of 10-20%; c) CF carriers (heterozygous subjects) with an activity between 40-70%.

Histopathological determinants of tumor resistance: a special look to the immunohistochemical expression of carbonic anhydrase IX in human cancers.

Intrinsic and acquired drug resistance of tumor cells still causes the failure of treatment regimens in advanced human cancers. It may be driven by intrinsic tumor cells features, or may also arise from micro environmental influences. Hypoxia is a microenvironment feature associated with the aggressiveness and metastasizing ability of human solid cancers. Hypoxic cancer cells overexpress Carbonic Anhydrase IX (CA IX). CA IX ensures a favorable tumor intracellular pH, while contributing to stromal acidosis, which facilitates tumor invasion and metastasis. The overexpression of CA IX is considered an epiphenomenon of the presence of hypoxic, aggressive tumor cells. Recently, a relationship between CA IX overexpression and the cancer stem cells (CSCs) population has been hypothesized. CSCs are strictly regulated by tumor hypoxia and drive a major non-mutational mechanism of cancer drug-resistance. We reviewed the current data concerning the role of CA IX overexpression in human malignancies, extending such information to the expression of the stem cells markers CD44 and nestin in solid cancers, to explore their relationship with the biological behavior of tumors. CA IX is heavily expressed in advanced tumors. A positive trend of correlation between CA IX overexpression, tumor stage/grade and poor outcome emerged. Moreover, stromal CA IX expression was associated with adverse events occurrence, maybe signaling the direct action of CA IX in directing the mesenchymal changes that favor tumor invasion; in addition, membranous/cytoplasmic co-overexpression of CA IX and stem cells markers were found in several aggressive tumors. This suggests that CA IX targeting could indirectly deplete CSCs and counteract resistance of solid cancers in the clinical setting.

FK506 binding protein 51 positively regulates melanoma stemness and metastatic potential.

Melanoma is the most aggressive skin cancer; there is no cure in advanced stages. Identifying molecular participants in melanoma progression may provide useful diagnostic and therapeutic tools. FK506 binding protein 51 (FKBP51), an immunophilin with a relevant role in developmental stages, is highly expressed in melanoma and correlates with aggressiveness and therapy resistance. We hypothesized a role for FKBP51 in melanoma invasive behaviour. FKBP51 promoted activation of epithelial-to-mesenchymal transition (EMT) genes and improved melanoma cell migration and invasion. In addition, FKBP51 induced some melanoma stem cell (MCSC) genes. Purified MCSCs expressed high EMT genes levels, suggesting that genetic programs of EMT and MCSCs overlap. Immunohistochemistry of samples from patients showed intense FKBP51 nuclear signal and cytoplasmic positivity for the stem cell marker nestin in extravasating melanoma cells and metastatic brains. In addition, FKBP51 targeting by small interfering RNA (siRNA) prevented the massive metastatic substitution of liver and lung in a mouse model of experimental metastasis. The present study provides evidence that the genetic programs of cancer stemness and invasiveness overlap in melanoma, and that FKBP51 plays a pivotal role in sustaining such a program.

Mononuclear blood cell magnesium content and serum magnesium concentration in critically ill hypomagnesemic patients after replacement therapy.

Magnesium (Mg) deficiency, commonly diagnosed as hypomagnesemia based upon low serum Mg concentrations, is a frequent electrolyte abnormality in critically ill patients. Intravenous replacement therapy is empiric and serum Mg concentrations have traditionally been used as guidelines for measuring efficacy. Recent studies have shown that the Mg content of mononuclear blood cells (MBCs) may provide a better index for Mg status than serum concentrations. The purpose of this study was to evaluate the effects of intravenous Mg replacement therapy on MBC Mg content and serum Mg concentrations in critically ill hypomagnesemic patients. Adult patients admitted to the trauma intensive-care unit (ICU) with serum Mg concentration < or = 0.6 mmol/L (< or = 1.5 mg/dL) were considered for study entry. Patients with severe renal disease (Scr > 133 mumol/L), pregnancy, or those who were seropositive for HIV were excluded. Ten patients with moderate (> 0.4-0.6 mmol/L [> 1.0-1.5 mg/dL]) and severe (< or = 0.4 mmol/L [< or = 1.0 mg/dL]) hypomagnesemia received 0.5 and 0.75 mmol/kg of intravenous MgSO4, respectively, over 24 h. MBC Mg content and serum concentrations of magnesium, phosphorus, calcium, sodium, potassium, blood urea nitrogen, creatinine, glucose, and albumin were measured at baseline (0 h), end of infusion (24 h), 36 h, and 48 h. Data were analyzed using ANOVA with repeated measures and a P value < 0.05 was considered significant. Serum Mg concentrations increased significantly from baseline to 48 h (0.5 +/- 0.1 to 0.8 +/- 0.2 mmol/L, P < 0.001). MBC Mg content did not change significantly within the study period (2.6 +/- 1.0 to 3.0 +/- 1.3 fmol/cell, P > 0.7). The doses of MgSO4 (0.5-0.75 mmol/kg) used in this study increased serum Mg concentrations, but did not result in a statistically significant change of MBC Mg content in this group of trauma ICU patients.