Introduction: Breast cancer stem cells are suspected to be responsible for tumour recurrence, metastasis formation as well as chemoresistance. Consequently, great efforts have been made to understand the molecular mechanisms underlying cancer stem cell maintenance. In order to study these rare cells in-vitro, they are typically enriched via mammosphere culture. Here we developed a mammosphere-based negative selection shRNAi screening system suitable to analyse the involvement of thousands of genes in the survival of cells with cancer stem cell properties. Methods: We describe a sub-population expressing the stem-like marker CD44 + /CD24 -/low in SUM149 that were enriched in mammospheres. To identify genes functionally involved in the maintenance of the sub-population with cancer stem cell properties, we targeted over 5000 genes by RNAi and tested their ability to grow as mammospheres. The identified candidate ATG4A was validated in mammosphere and soft agar colony formation assays. Further, we evaluated the influence of ATG4A expression on the sub-population expressing the stem-like marker CD44

L1 cell adhesion molecule (L1CAM) overexpression is often associated with bad prognosis in various human carcinomas. Recent studies also suggest a role of L1CAM in pancreatic ductal adenocarcinomas (PDAC). To further address its contribution, we expressed functional domains of L1CAM in PT45-P1 PDAC cells. We found that L1CAM that is full length (L1-FL), but neither the soluble ectodomain (L1ecto) nor the cytoplasmic part (L1cyt), could enhance cell proliferation or tumour growth in mice. Expression of L1-FL resulted in constitutive activation of NF-jB, which was abolished by L1CAM knockdown. We showed that the expression of IL-1b was selectively upregulated by L1-FL, and increased IL-1b levels were instrumental for sustained NF-jB activation. IL-1b production and NF-jB activation were abolished by knockdown of a5-integrin and integrin-linked kinase, but insensitive to depletion of L1CAM cleavage proteinases. Supporting these data, PT45-P1 cells transduced with an L1CAM mutant deficient in integrin binding (L1-RGE) did not support the described L1-FL functions. Our results suggest that membranous L1CAM interacts with RGD-binding integrins, leading to sustained NF-jB activation by IL-1b production and autocrine/paracrine signalling. The unravelling of this novel mechanism sheds new light on the important role of L1CAM expression in PDAC cells.

The gold standard for determining the tumorigenic potential of human cancer cells is a xenotransplantation into immunodeficient mice. Higher tumorigenicity of cells is associated with earlier tumor onset. Here, we used xenotransplantation to assess the tumorigenic potential of human breast cancer cells following RNA interference-mediated inhibition of over 5000 genes. We identify 16 candidate tumor suppressors, one of which is the zinc-finger transcription factor SALL1. Analyzing this particular molecule in more detail, we show that inhibition of SALL1 correlates with reduced levels of CDH1, an important contributor to epithelial-tomesenchymal transition. Furthermore, SALL1 expression led to an increased migration and more than twice as many cells expressing a cancer stem cell signature. Also, SALL1 expression correlates with the survival of breast cancer patients. These findings cast new light on a gene that has previously been described to be relevant during embryogenesis, but not carcinogenesis.

BACKGROUND Hepatitis B virus (HBV) infection is a major public health problem in Brazil. Several risk factors are involved in HBV infection and their identification by a rational and essential approach is required to prevent the transmission of this infection in Brazil.OBJECTIVES To evaluate risk factors associated with HBV infection in South Brazil.METHODS A total of 260 patients with HBV and 260 controls from Caxias do Sul (state of Rio Grande do Sul, Brazil) participated in this study. All participants were given a standard questionnaire to yield the sociodemographic information and to identify HBV risk factors. HBV infection was detected by HBsAg test in all participants.FINDINGSHBV infection in these cases was strongly associated with history of a family member HBV-infected, mainly mother [odds ratio (OR) = 4.86; 95% confidence intervals (CI): 1.69–13.91], father (OR = 5.28; 95% CI: 1.58–17.71), and/or siblings (OR = 22.16; 95% CI: 9.39–52.25); sharing personal objects (OR = 1.40; 95% CI: 1.37–2.38); and having history of blood transfusion (OR = 2.05; 95% CI: 1.10–2.84).CONCLUSIONS HBV infection was strongly associated with having a family member infected with hepatitis B, sharing personal objects, and having history of blood transfusion.

SummaryChemotherapy of advanced pancreatic cancer has mainly been gemcitabine-based for the past 15 years, with only limited effect. Recently, combination therapy that also targets checkpoint kinase 1 (CHK1) has become an attractive option. The central role of CHK1 in many DNA-damage response pathways, however, may result in undesired cytotoxicity in normal cells, causing side effects. We were searching for other target molecules of similar function that may be more specific and thus better suited for combination therapy. To this end a negative selection RNAi screen was performed in cell lines with small hairpin RNA molecules targeting over 10,000 genes. Genes that were found to be synthetically lethal with gemcitabine and whose proteins act upstream of CHK1 were characterised in more detail. In particular, the inhibition of RAD17 potentiated gemcitabine cytotoxicity in the pancreatic cancer cell lines BxPC-3 and MiaPaca-2 and in the primary cell line JoPaca-1 that closely resembles primary tumour tissue. Further analysis showed that the synergistic effect of RAD17 knockdown and gemcitabine leads to forced mitotic entry of cells arrested in S phase by gemcitabine treatment, resulting in asymmetric DNA distribution during anaphase followed by DNA fragmentation and finally cell death by mitotic catastrophe. Our data suggest RAD17 as a novel target protein for gemcitabine combination therapy supplementing or complementing inhibition of CHK1. In contrast to CHK1, RAD17 knockdown by itself does not lead to abnormal DNA segregation, suggesting a more specific action.

Electrifying the chemical industry is a key step towards the generation of green and sustainable chemical products. Electrochemical hydrogenations have gained a central role in this effort, generating important synthons...

Human papillomavirus (HPV) infection is common in sexually active women and viral persistence may cause intraepithelial lesions and eventually progress to cervical cancer (CC). The present study aimed to investigate epidemiological factors related to HPV infection and to evaluate viral persistence and CC precursor lesions frequencies in women from a city in the countryside of South Brazil. Three hundred women were recruited from a primary public health care clinic. The patients were interviewed and underwent sampling with cervical brushes for HPV-DNA detection/typing by a PCR-based assay and cytological analysis by Pap smear test. HPV was detected in 47 (15.7%) women. HPV infection was significantly associated with young age (<30 years) and low socio-economic status. Seventeen (5.7%) women presented cytological abnormalities, three of them with precursor CC intraepithelial lesions. A subgroup of 79 women had been previously analyzed and thirteen (16.4%) were persistently infected, two with precursor CC intraepithelial lesions and high-risk HPV types infection (both of them without cervical abnormalities in the first exam). In conclusion, HPV infection was associated with young age (<30 years) and low family income; viral persistence was low (16.4%) but related to CC precursor lesions; and HPV-DNA high risk types detection would help to screen CC in the population.

These findings indicate that hMPV is in circulation in southern Brazil and highlight the importance of diagnosing hMPV for influenza-like illness in the population.