Potency analysis of cellular therapies: the emerging role of molecular assays

Potency testing is an important part of the evaluation of cellular therapy products. Potency assays are quantitative measures of a product-specific biological activity that is linked to a relevant biological property and, ideally, a product's in vivo mechanism of action. Both in vivo and in vitro assays can be used for potency testing. Since there is often a limited period of time between the completion of production and the release from the laboratory for administration to the patient, in vitro assays such are flow cytometry, ELISA, and cytotoxicity are typically used. Better potency assays are needed to assess the complex and multiple functions of cellular therapy products, some of which are not well understood. Gene expression profiling using microarray technology has been widely and effectively used to assess changes of cells in response to stimuli and to classify cancers. Preliminary studies have shown that the expression of noncoding microRNA which play an important role in cellular development, differentiation, metabolism and signal transduction can distinguish different types of stem cells and leukocytes. Both gene and microRNA expression profiling have the potential to be important tools for testing the potency of cellular therapies. Potency testing, the complexities associated with potency testing of cellular therapies, and the potential role of gene and microRNA expression microarrays in potency testing of cellular therapies is discussed

Primary malignant melanoma of the stomach: report of a case

We report a case of primary malignant melanoma (MM) of the stomach. The patient, a 73-year-old man, was referred to our hospital for investigation of an elevated lesion in the stomach, detected by gastroscopy. On admission, physical examinations and laboratory data were unremarkable. Gastroscopy revealed a pigmented, elevated tumor, approximately 2 cm in diameter, in the posterior wall of the stomach. A biopsy was taken, which resulted in a diagnosis of MM, based on the presence of melanin in tumor cells. F-18 fluorodeoxyglucose positron emission tomography showed no accumulation of tracer except for the tumor in the stomach, indicating that it was a primary MM of the stomach. The patient underwent distal gastrectomy, but died of recurrence 1 year later. Very few cases of primary MM of the stomach have been reported. Thus, we report this case, followed by a review of the literature

Interleukins, laminin and epstein - barr virus latent membrane protein 1 (EBV LMP1) Promote metastatic phenotype in nasopharyngeal carcinoma

<p>Abstract</p> <p>Background</p> <p>Nasopharyngeal carcinoma (NPC) is a type of neoplasm that is highly prevalent in East Asia and Africa with Epstein-Barr virus (EBV), genetic, and dietary factors implicated as possible aetiologic factors. Previous studies suggested the association of certain cytokines with the invasion and metastatic properties of NPC. The present study examined the roles of EBV latent membrane protein-1 (LMP1), interleukin-6 (IL-6), interleukin-10 (IL-10), transforming growth factor-beta 1 (TGF-β1) and laminin in the regulation of matrix-metalloproteinases (MMPs) and vascular endothelial growth factor (VEGF) in NPC. The effects of these factors on <it>bmi-1</it>, an oncogene, and <it>ngx6</it>, a tumour suppressor gene, were also investigated.</p> <p>Methods</p> <p>TW01 cells expressing LMP1 (TW01-LMP1) were established via transfection with the B95.8 EBV LMP1 gene. Both TW01 and TW01-LMP1 cells were treated with 100 pg/ml IL-6, 1000 pg/ml IL-10 and 100 pg/ml TGF-β1, separately and also in combination at their respective concentration for 48 hours. Treated cells were subjected to laminin adherence assay. The cells were also cultured with and without laminin and assayed for MMP-3, MMP-9 and VEGF production using enzyme-linked immunosorbent assay (ELISA). The cellular apoptotic property was analysed using caspase-3 apoptosis assay. The expression of <it>bmi-1 </it>and <it>ngx6 </it>gene was investigated using real time reverse transcriptase polymerase chain reaction.</p> <p>Results</p> <p>LMP1 was found to reduce the adherence of NPC cells towards laminin (p < 0.05) as compared to control. Treatment with IL-6 at 100 pg/ml enhanced the production of MMP-9 in both TW01 and TW01-LMP1 cells (p < 0.05). When cultured on laminin, the levels of MMP-3 and VEGF were significantly increased (p < 0.05) in TW01-LMP1 cells. TW01-LMP1 cells had relatively greater resistance to apoptosis as compared to TW01 cells (p < 0.05). Laminin, IL-6 and LMP1 were found to up-regulate the expression of <it>bmi-1 </it>and suppressed the expression of <it>ngx6</it>.</p> <p>Conclusions</p> <p>We conclude that IL-6 reduced cell adherence towards laminin and increased MMP-9 production in NPC cells. Our data suggested that EBV LMP1 was able to confer resistance of apoptosis and increased MMP-9 production in NPC cells. When cultured on laminin, TW01 cells expressing the EBV LMP1 (TW0-LMP1) that were treated with IL-6 at 100 pg/ml displayed increased MMP-9 production, up-regulation of <it>bmi-1 </it>oncogene expression and down-regulation of <it>ngx6 </it>tumour suppressor gene expression. These findings implicate the roles of EBV LMP1, laminin and IL-6 in the promotion of invasion and metastasis in NPC.</p

MUC1 alters oncogenic events and transcription in human breast cancer cells

INTRODUCTION: MUC1 is an oncoprotein whose overexpression correlates with aggressiveness of tumors and poor survival of cancer patients. Many of the oncogenic effects of MUC1 are believed to occur through interaction of its cytoplasmic tail with signaling molecules. As expected for a protein with oncogenic functions, MUC1 is linked to regulation of proliferation, apoptosis, invasion, and transcription. METHODS: To clarify the role of MUC1 in cancer, we transfected two breast cancer cell lines (MDA-MB-468 and BT-20) with small interfering (si)RNA directed against MUC1 and analyzed transcriptional responses and oncogenic events (proliferation, apoptosis and invasion). RESULTS: Transcription of several genes was altered after transfection of MUC1 siRNA, including decreased MAP2K1 (MEK1), JUN, PDGFA, CDC25A, VEGF and ITGAV (integrin α(v)), and increased TNF, RAF1, and MMP2. Additional changes were seen at the protein level, such as increased expression of c-Myc, heightened phosphorylation of AKT, and decreased activation of MEK1/2 and ERK1/2. These were correlated with cellular events, as MUC1 siRNA in the MDA-MB-468 line decreased proliferation and invasion, and increased stress-induced apoptosis. Intriguingly, BT-20 cells displayed similar levels of apoptosis regardless of siRNA, and actually increased proliferation after MUC1 siRNA. CONCLUSION: These results further the growing knowledge of the role of MUC1 in transcription, and suggest that the regulation of MUC1 in breast cancer may be more complex than previously appreciated. The differences between these two cell lines emphasize the importance of understanding the context of cell-specific signaling events when analyzing the oncogenic functions of MUC1, and caution against generalizing the results of individual cell lines without adequate confirmation in intact biological systems

Emerging concepts in biomarker discovery; The US-Japan workshop on immunological molecular markers in oncology

Supported by the Office of International Affairs, National Cancer Institute (NCI), the "US-Japan Workshop on Immunological Biomarkers in Oncology" was held in March 2009. The workshop was related to a task force launched by the International Society for the Biological Therapy of Cancer (iSBTc) and the United States Food and Drug Administration (FDA) to identify strategies for biomarker discovery and validation in the field of biotherapy. The effort will culminate on October 28th 2009 in the "iSBTc-FDA-NCI Workshop on Prognostic and Predictive Immunologic Biomarkers in Cancer", which will be held in Washington DC in association with the Annual Meeting. The purposes of the US-Japan workshop were a) to discuss novel approaches to enhance the discovery of predictive and/or prognostic markers in cancer immunotherapy; b) to define the state of the science in biomarker discovery and validation. The participation of Japanese and US scientists provided the opportunity to identify shared or discordant themes across the distinct immune genetic background and the diverse prevalence of disease between the two Nations

Breast cancer stem cells: implications for therapy of breast cancer

The concept of cancer stem cells responsible for tumour origin, maintenance, and resistance to treatment has gained prominence in the field of breast cancer research. The therapeutic targeting of these cells has the potential to eliminate residual disease and may become an important component of a multimodality treatment. Recent improvements in immunotherapy targeting of tumour-associated antigens have advanced the prospect of targeting breast cancer stem cells, an approach that might lead to more meaningful clinical remissions. Here, we review the role of stem cells in the healthy breast, the role of breast cancer stem cells in disease, and the potential to target these cells

Consensus guidelines for the use and interpretation of angiogenesis assays

The formation of new blood vessels, or angiogenesis, is a complex process that plays important roles in growth and development, tissue and organ regeneration, as well as numerous pathological conditions. Angiogenesis undergoes multiple discrete steps that can be individually evaluated and quantified by a large number of bioassays. These independent assessments hold advantages but also have limitations. This article describes in vivo, ex vivo, and in vitro bioassays that are available for the evaluation of angiogenesis and highlights critical aspects that are relevant for their execution and proper interpretation. As such, this collaborative work is the first edition of consensus guidelines on angiogenesis bioassays to serve for current and future reference

Search for dark matter produced in association with a Higgs boson decaying to a pair of bottom quarks in proton-proton collisions at root s=13TeV

A search for dark matter produced in association with a Higgs boson decaying to a pair of bottom quarks is performed in proton-proton collisions at a center-of-mass energy of 13 TeV collected with the CMS detector at the LHC. The analyzed data sample corresponds to an integrated luminosity of 35.9 fb(-1). The signal is characterized by a large missing transverse momentum recoiling against a bottom quark-antiquark system that has a large Lorentz boost. The number of events observed in the data is consistent with the standard model background prediction. Results are interpreted in terms of limits both on parameters of the type-2 two-Higgs doublet model extended by an additional light pseudoscalar boson a (2HDM+a) and on parameters of a baryonic Z simplified model. The 2HDM+a model is tested experimentally for the first time. For the baryonic Z model, the presented results constitute the most stringent constraints to date.Peer reviewe

Search for strongly interacting massive particles generating trackless jets in proton-proton collisions at s = 13 TeV

A search for dark matter in the form of strongly interacting massive particles (SIMPs) using the CMS detector at the LHC is presented. The SIMPs would be produced in pairs that manifest themselves as pairs of jets without tracks. The energy fraction of jets carried by charged particles is used as a key discriminator to suppress efficiently the large multijet background, and the remaining background is estimated directly from data. The search is performed using proton-proton collision data corresponding to an integrated luminosity of 16.1 fb - 1 , collected with the CMS detector in 2016. No significant excess of events is observed above the expected background. For the simplified dark matter model under consideration, SIMPs with masses up to 100 GeV are excluded and further sensitivity is explored towards higher masses