Novel comprehensive approach for accessible biomarker identification and absolute quantification from precious human tissues

in Journal of Proteome Research (2011), 10(7), 3160-82

The identification of specific biomarkers obtained directly from human pathological lesions remains a major challenge, because the amount of tissue available is often very limited. We have developed a ... [more ▼]

The identification of specific biomarkers obtained directly from human pathological lesions remains a major challenge, because the amount of tissue available is often very limited. We have developed a novel, comprehensive, and efficient method permitting the identification and absolute quantification of potentially accessible proteins in such precious samples. This protein subclass comprises cell membrane associated and extracellular proteins, which are reachable by systemically deliverable substances and hence especially suitable for diagnosis and targeted therapy applications. To isolate such proteins, we exploited the ability of chemically modified biotin to label ex vivo accessible proteins and the fact that most of these proteins are glycosylated. This approach consists of three successive steps involving first the linkage of potentially accessible proteins to biotin molecules followed by their purification. The remaining proteins are then subjected to glycopeptide isolation. Finally, the analysis of the nonglycosylated peptides and their involvement in an in silico method increased the confident identification of glycoproteins. The value of the technique was demonstrated on human breast cancer tissue samples originating from 5 individuals. Altogether, the method delivered quantitative data on more than 400 potentially accessible proteins (per sample and replicate). In comparison to biotinylation or glycoprotein analysis alone, the sequential method significantly increased the number (≥30% and ≥50% respectively) of potentially therapeutically and diagnostically valuable proteins. The sequential method led to the identification of 93 differentially modulated proteins, among which several were not reported to be associated with the breast cancer. One of these novel potential biomarkers was CD276, a cell membrane-associated glycoprotein. The immunohistochemistry analysis showed that CD276 is significantly differentially expressed in a series of breast cancer lesions. Due to the fact that our technology is applicable to any type of tissue biopsy, it bears the ability to accelerate the discovery of new relevant biomarkers in a broad spectrum of pathologies. [less ▲]

Cancer du sein: importance de la nomenclature anatomo-pathologique

in Revue Médicale de Liège (2011), 66(5-6), 261-4

The breast pathology includes a large array of entities for which macroscopic and microscopic analysis remains fundamental. Tissue and cell morphology allows in most cases the distinction between benign ... [more ▼]

The breast pathology includes a large array of entities for which macroscopic and microscopic analysis remains fundamental. Tissue and cell morphology allows in most cases the distinction between benign or malignant tumours and therefore provides the clinicians with essential information for the therapeutic strategy. In the Pathology laboratory, immunohistochemistry and molecular biology have improved the specificity of the diagnosis and have introduced new prognostic and predictive markers for tumour management. The last edition of the WHO classification, released in 2003, distinguishes 21 varieties of invasive carcinoma and 2 categories of intraepithelial neoplasia based on the morphology and immunohistochemical profile. Other diseases can affect the breast, although much less frequently, such as Paget’s disease of the nipple, phyllode tumours, sarcomas, lymphomas... These diseases will not be reviewed here. [less ▲]

Mucosal junctions: open doors to HPV and HIV infections?

in Trends in microbiology (2011), 19(3), 114-120

Throughout adult life, new developmental commitment of adult stem cells causes reversible epithelial replacements in various mucosal surfaces, including the uterine cervix and the anal canal. Located at ... [more ▼]

Throughout adult life, new developmental commitment of adult stem cells causes reversible epithelial replacements in various mucosal surfaces, including the uterine cervix and the anal canal. Located at the squamocolumnar junctions, these metaplastic conversions are associated with chronic inflammation and deregulated expression of soluble and cell-membrane factors important for antiviral immune response. In this paper, we propose that these histological and immunological features increase the susceptibility of these metaplastic microenvironments to human papillomavirus and human immunodeficiency virus infections. Identification of the anatomical sites and cell populations within the anogenital tract, which is the site primary infected by these viruses, is crucial for the understanding of the pathogenesis of viral disease and development of antiviral strategies. [less ▲]

IFN-gamma and TNF-alpha potentiate prostaglandin D2-induced human eosinophil chemotaxis through up-regulation of CRTH2 surface receptor.

in International immunopharmacology (2011), 11(11), 1864-70

Prostaglandin D2 (PGD2) receptor CRTH2, is a pro-inflammatory molecule involved in eosinophil recruitment to the allergic airway. We investigated the expression of CRTH2 in eosinophil from allergic ... [more ▼]

Prostaglandin D2 (PGD2) receptor CRTH2, is a pro-inflammatory molecule involved in eosinophil recruitment to the allergic airway. We investigated the expression of CRTH2 in eosinophil from allergic rhinitis patients (AR) and tested the modulatory role of several TH1 and TH2 cytokines closely related to the allergic immunological response, on the expression of CRTH2 receptor, utilizing human eosinophil cell line (Eol-1).The expression of CRTH2 was tested by immunohistochemistry and flow cytometry (FACS). Chemotaxis was performed in micro-chemotaxis chambers. It is shown that the expression of CRTH2 by eosinophils was significantly higher in the nasal tissue and peripheral blood of AR patients, when compared to control subjects. PGD2 exhibited a typical bell shape dose response in attracting eosinophil from AR patients with optimal activity at 10(-7)M. Eol-1 cell surface expression of CRTH2 was significantly up-regulated by 10ng/ml IFN-gamma and TNF-alpha. The percentage of Eol-1 cells expressing the receptor increased by IFN-gamma and TNF-alpha from 12.74%+/-2.66 to 55%+/-8 and 33.8%+/-9.4, respectively. PGD2-induced Eol-1 chemotaxis was not blocked by SB203580, H-89 Dihydrochloride, Bisindo-lylmaleimide, or Genistein. PGD2-induced Eol-1 chemotaxis was potentiated by IFN-gamma and TNF-alpha without changing the signal transduction pathway. Correlation of our results to peripheral blood eosinophils from allergic rhinitis patients confirmed that 3hour pretreatment of eosinophils by 10ng/ml IFN-gamma and TNF-alpha, increased the mean fluorescence intensity (MFI) of CRTH2 from 8.23 to 9.68 and 9.38, respectively, and potentiated PGD2-induced eosinophil chemotaxis. Our results demonstrate a novel synergism between PGD2, IFN-gamma and TNF-alpha, in eosinophil chemotaxis. [less ▲]

Traitement par anti-TNF-alpha et cancers de la peau

in ONCO-HEMATO (2011), 5

High prevalence of high-risk human papillomavirus in palatine tonsils from healthy children and adults.

in Otolaryngology - Head & Neck Surgery (2011), 145(2), 230-5

OBJECTIVE: The aim of this study was to determine the prevalence of human papillomavirus (HPV) in 80 tumor-free tonsils from healthy children and adults using a sensitive E6/E7 type-specific polymerase ... [more ▼]

OBJECTIVE: The aim of this study was to determine the prevalence of human papillomavirus (HPV) in 80 tumor-free tonsils from healthy children and adults using a sensitive E6/E7 type-specific polymerase chain reaction (PCR). STUDY DESIGN: Cross-sectional study. SETTING: Ear, nose, and throat department, university hospital. SUBJECTS AND METHODS: Paraffin-embedded tissues from tumor-free tonsils (TFTs) were evaluated for HPV DNA using GP5+/6+ consensus PCR and subsequent genotyping using E6/E7 type-specific PCR for HPV types 6, 11, 16, 18, 31, 33, 35, 39, 45, 51, 52, 53, 56, 58, 59, 66, and 68. The immunohistochemical expression of p16 and p53 was also assessed. RESULTS: In 80 TFTs, the authors identified 10 (12.5%) that tested positive for the following high-risk HPV types: HPV 16 (8 cases), 18 (1 case), and 31 (1 case). Twelve patients (15%) tested positive for HPV infection using the GP5+/GP6+ consensus primers but were negative using quantitative PCR. These patients were considered infected with low-risk HPV types. Fifty-eight TFTs (72.5%) tested negative for both GP5+/GP6+ and type-specific HPV PCR analysis (HPV negative). Among patients infected with HPV, the authors observed a slight increase in frequency with age. CONCLUSION: In TFTs, oncogenic and nononcogenic HPVs were present at a relatively high frequency in children and adults. The presence of high-risk HPV DNA in young children supports the horizontal transmission hypothesis and argues in favor of HPV vaccination at birth. [less ▲]

Unimpeded skin carcinogenesis in K14-HPV16 transgenic mice deficient for plasminogen activator inhibitor

in International Journal of Cancer = Journal International du Cancer (2011), 128(2), 283-93

Angiogenesis, extracellular matrix remodeling and cell migration are associated with cancer progression and involve at least, the plasminogen activating system and its main physiological inhibitor, the ... [more ▼]

Angiogenesis, extracellular matrix remodeling and cell migration are associated with cancer progression and involve at least, the plasminogen activating system and its main physiological inhibitor, the plasminogen activator inhibitor-1 (PAI-1). Considering the recognized importance of PAI-1 in the regulation of tumor angiogenesis and invasion in murine models of skin tumor transplantation, we explored the functional significance of PAI-1 during early stages of neoplastic progression in the transgenic mouse model of multistage epithelial carcinogenesis (K14-HPV16 mice). We have studied the effect of genetic deletion of PAI-1 on inflammation, angiogenesis, lymphangiogenesis, as well as tumor progression. In this model, PAI-1 deficiency neither impaired keratinocyte hyperproliferation or tumor development, nor affected the infiltration of inflammatory cells and development of angiogenic or lymphangiogenic vasculature. We are reporting evidence for concomitant lymphangiogenic and angiogenic switches independent to PAI-1 status. Taken together, these data indicate that PAI-1 is not rate limiting for neoplastic progression and vascularization during premalignant progression, or that there is a functional redundancy between PAI-1 and other tumor regulators, masking the effect of PAI-1 deficiency in this long-term model of multi-stage epithelial carcinogenesis. [less ▲]

Implication of plasmacytoid dendritic cells in the malignant transformation of cervical epithelial metaplasia

Poster (2011)

Natalizumab to kill two birds with one stone: A case of celiac disease and multiple sclerosis.

in Inflammatory Bowel Diseases (2011), 17(6), 62-63

High incidence of high-risk HPV in benign and malignant lesions of the larynx.

in International Journal of Oncology (2011), 39(1), 51-9

The aim of this study was to determine the prevalence of human papillomavirus (HPV) in patients with laryngeal benign lesions (LBLs) and laryngeal squamous cell carcinomas (LSCCs) using a sensitive E6/E7 ... [more ▼]

The aim of this study was to determine the prevalence of human papillomavirus (HPV) in patients with laryngeal benign lesions (LBLs) and laryngeal squamous cell carcinomas (LSCCs) using a sensitive E6/E7 type-specific PCR. Paraffin-embedded samples from LBL (n=39) and LSCC patients (n=67) were evaluated for the presence of HPV DNA by GP5+/GP6+ consensus PCR and E6/E7 type-specific PCR for HPV types 6, 11, 16, 18, 31, 33, 35, 39, 45, 51, 52, 53, 56, 58, 59, 66 and 68. In LSCCs, immunohistochemical staining of p16, p53 and EGFR was also assessed. The E6/E7 type-specific PCR showed that 44 out of 59 LSCC patients (i.e., 75%) had high-risk (hr) HPV types and that 27 out of 35 LBL patients (i.e., 77%) had hrHPV types. HPV-16 viral load was significantly higher in LSCC than in LBL patients (p<10-6). The presence of hrHPV DNA did not correlate with the proportion of disease-free patients. Comparable levels of p16, p53 and EGFR expression were observed in the hrHPV+ tumor group (100% p16+, 56% p53+ and 97% EGFR+) and in the HPV- or low-risk (lr) HPV+ tumor group (92% p16+, 66% p53+ and 100% EGFR+). A very high prevalence of oncogenic HPV-16 was found in a series of benign and malignant laryngeal lesions. LSCC appears to be characterized by an active hrHPV infection. In LSCCs, the hrHPV+ subgroup had a similar prognosis (in terms of risk of recurrence) as the HPV- subgroup. [less ▲]