Epidermal growth factor receptor regulates fibrinolytic pathway elements in cervical cancer: functional and prognostic implications

Epidermal growth factor receptor (EGFR) signaling and components of the fibrinolytic system, including urokinase-type plasminogen activator (uPA) and thrombomodulin (TM), have been implicated in tumor progression. In the present study, we employed cBioPortal platform (http://www.cbioportal.org/), cancer cell lines, and an in vivo model of immunocompromised mice to evaluate a possible cooperation between EGFR signaling, uPA, and TM expression/function in the context of cervical cancer. cBioPortal analysis revealed that EGFR, uPA, and TM are positively correlated in tumor samples of cervical cancer patients, showing a negative prognostic impact. Aggressive human cervical cancer cells (CASKI) presented higher gene expression levels of EGFR, uPA, and TM compared to its less aggressive counterpart (C-33A cells). EGFR induces uPA expression in CASKI cells through both PI3K-Akt and MEK1/2-ERK1/2 downstream effectors, whereas TM expression induced by EGFR was dependent on PI3K/Akt signaling alone. uPA induced cell-morphology modifications and cell migration in an EGFR-dependent and -independent manner, respectively. Finally, treatment with cetuximab reduced in vivo CASKI xenografted-tumor growth in nude mice, and decreased intratumoral uPA expression, while TM expression was unaltered. In conclusion, we showed that EGFR signaling regulated expression of the fibrinolytic system component uPA in both in vitro and in vivo settings, while uPA also participated in cell-morphology modifications and migration in a human cervical cancer model.

Radiotherapy modulates expression of EGFR, ERCC1 and p53 in cervical cancer

Cervical cancer is a public health problem and the molecular mechanisms underlying radioresistance are still poorly understood. Here, we evaluated the modulation of key molecules involved in cell proliferation, cell cycle and DNA repair in cervical cancer cell lines (CASKI and C33A) and in malignant tissues biopsied from 10 patients before and after radiotherapy. The expression patterns of epidermal growth factor receptor (EGFR), excision repair cross-complementation group 1 (ERCC1) and p53 were evaluated in cancer cell lines by quantitative PCR and western blotting, and in human malignant tissues by immunohistochemistry. The mutation status of TP53 gene was evaluated by direct sequencing. Among cell lines, absent or weak modulations of EGFR, ERCC1 and p53 were observed after exposure to 1.8 Gy. Conversely, increased expressions of p53 (5/10 patients; P=0.0239), ERCC1 (5/10 patients; P=0.0294) and EGFR (4/10 patients; P=0.1773) were observed in malignant tissues after radiotherapy with the same radiation dose. TP53 mutations were found only in one patient. Here we show that a single dose of radiotherapy induced EGFR, ERCC1 and p53 expression in malignant tissues from cervical cancer patients but not in cancer cell lines, highlighting the gap between in vitro and in vivo experimental models. Studies on larger patient cohorts are needed to allow an interpretation that an upregulation of p53, EGFR and ERCC1 may be part of a radioresistance mechanism.

Increased expression of keratinase and other peptidases by Candida parapsilosis mutants

Keratinases are enzymes of great importance involved in pathogenic processes of some fungi. They also have a widespread ecological role since they are responsible for the degradation and recycling of keratin. On the one hand, studying them furthers our knowledge of pathogenicity mechanisms, which has important implications for human health, and on the other hand, understanding their ecological role in keratin recycling has biotechnological potential. Here, a wild-type keratinolytic Candida parapsilosis strain isolated from a poultry farm was treated with ethyl methanesulfonate in order to generate mutants with increased keratinase activity. Mutants were then cultured on media with keratin extracted from chicken feathers as the sole source of nitrogen and carbon. Approximately 500 mutants were screened and compared with the described keratinolytic wild type. Three strains, H36, I7 and J5, showed enhanced keratinase activity. The wild-type strain produced 80 U/mL of keratinolytic activity, strain H36 produced 110 U/mL, strain I7, 130 U/mL, and strain J5, 140 U/mL. A 70 percent increase in enzyme activity was recorded for strain J5. Enzymatic activity was evaluated by zymograms with proteic substrates. A peptidase migrating at 100 kDa was detected with keratin, bovine serum albumin and casein. In addition, a peptidase with a molecular mass of 50 kDa was observed with casein in the wild-type strain and in mutants H36 and J5. Gelatinase activity was detected at 60 kDa. A single band of 35 kDa was found in wild-type C. parapsilosis and in mutants with hemoglobin substrate.

A distributed computation of Interpro Pfam, PROSITE and ProDom for protein annotation

Interpro is a widely used tool for protein annotation in genome sequencing projects, demanding a large amount of computation and representing a huge time-consuming step. We present a strategy to execute programs using databases Pfam, PROSITE and ProDom of Interpro in a distributed environment using a Java-based messaging system. We developed a two-layer scheduling architecture of the distributed infrastructure. Then, we made experiments and analyzed the results. Our distributed system gave much better results than Interpro Pfam, PROSITE and ProDom running in a centralized platform. This approach seems to be appropriate and promising for highly demanding computational tools used for biological applications.