Cost-effectiveness of CYP2C19 genotyping to guide antiplatelet therapy for acute minor stroke and high-risk transient ischemic attack.

Dual antiplatelet therapy (DAPT) with clopidogrel plus aspirin within 48 h of acute minor strokes and transient ischemic attacks (TIAs) has been indicated to effectively reduce the rate of recurrent strokes. However, the efficacy of clopidogrel has been shown to be affected by cytochrome P450 2C19 (CYP2C19) polymorphisms. Patients carrying loss-of-function alleles (LoFAs) at a low risk of recurrence (ESRS < 3) cannot benefit from clopidogrel plus aspirin at all and may have an increased bleeding risk. In order to optimize antiplatelet therapy for these patients and avoid the waste of medical resources, it is important to identify the subgroups that genuinely benefit from DAPT with clopidogrel plus aspirin through CYP2C19 genotyping. This study sought to assess the cost-effectiveness of CYP2C19 genotyping to guide drug therapy for acute minor strokes or high-risk TIAs in China. A decision tree and Markov model were constructed to evaluate the cost-effectiveness of CYP2C19 genotyping. We used a healthcare payer perspective, and the primary outcomes included quality-adjusted life years (QALYs), costs and the incremental cost-effectiveness ratio (ICER). Sensitivity analyses were performed to evaluate the robustness of the results. CYP2C19 genotyping resulted in a lifetime gain of 0.031 QALYs at an additional cost of CNY 420.13 (US$ 59.85), yielding an ICER of CNY 13,552.74 (US$ 1930.59) per QALY gained. Probabilistic sensitivity analysis showed that genetic testing was more cost-effective in 95.7% of the simulations at the willingness-to-pay threshold of CNY 72,100 (GDP per capita, US$ 10,300) per QALY. Therefore, CYP2C19 genotyping to guide antiplatelet therapy for acute minor strokes and high-risk TIAs is highly cost-effective in China.

Screening and identification of key biomarkers in alimentary tract cancers: A bioinformatic analysis.

BACKGROUND: Alimentary tract cancers (ATCs) are the most malignant cancers in the world. Numerous studies have revealed the tumorigenesis, diagnosis and treatment of ATCs, but many mechanisms remain to be explored. METHODS: To identify the key genes of ATCs, microarray datasets of oesophageal cancer, gastric cancer and colorectal cancer were obtained from the Gene Expression Omnibus (GEO) database. In total, 207 differentially expressed genes (DEGs) were screened. KEGG and GO function enrichment analyses were conducted, and a protein-protein interaction (PPI) network was generated and gene modules analysis was performed using STRING and Cytoscape. RESULTS: Five hub genes were screened, and the associated biological processes indicated that these genes were mainly enriched in cellular processes, protein binding and metabolic processes. Clinical survival analysis showed that COL10A1 and KIF14 may be significantly associated with the tumorigenesis or pathology grade of ATCs. In addition, relative human ATC cell lines along with blood samples and tumour tissues of ATC patients were obtained. The data proved that high expression of COL10A1 and KIF14 was associated with tumorigenesis and could be detected in blood. CONCLUSION: In conclusion, the identification of hub genes in the present study helped us to elucidate the molecular mechanisms of tumorigenesis and identify potential diagnostic indicators and targeted treatment for ATCs.

Generation of IgE-specific cytotoxic T lymphocytes as a novel immunotherapeutic approach for the treatment of allergic asthma.

INTRODUCTION: Overproduction of immunoglobulin E (IgE) by a subset of B cells plays a key role in the pathogenesis of allergic asthma. Anti-IgE monoclonal antibodies have been successfully used to treat the disease, but long-term application is required. METHODS: For this study, cytotoxic T lymphocytes (CTLs) against IgE-producing B cells were generated ex vivo by stimulating naive CD8 T cells with IgE-derived peptides presented by Drosophila-derived artificial antigen-presenting cells. Based on the treatment of allergic asthma in mice, the inhibitive effect of this CTL on IgE responses and airway inflammation was determined with the enzyme-linked immunosorbent assay and histochemical method. RESULTS: The IgE-specific CTLs effectively lysed target cells in vitro, while the adoptively transferred CTLs specifically inhibited IgE responses and airway inflammation in an asthmatic mouse model. The effect of IgE-specific CTLs is MHC (major histocompatibility complex) Class I-restricted and requires the expression of perforin. CONCLUSION: IgE-specific CTLs generated ex vivo may provide a novel treatment for allergic asthma and lead to a new therapy for other immunological disorders.

Photochemical Treatment of Drosophila APCs Can Eliminate Associated Viruses and Maintain the APC Function for Generating Antigen-Specific CTLs Ex Vivo.

Drosophila cells transfected with MHC class I and a number of costimulation molecules including B7.1, ICAM, LFA-3, and CD70 are potent antigen-presenting cells (APCs) for the generation of antigen-specific cytotoxic T cells (CTLs) in vitro. Using Drosophila APCs, CTLs specific for melanoma antigens have been generated in vitro and adoptively transferred to melanoma patients. However, the recent discovery that Drosophila cells can carry insect viruses raises the potential risk of Drosophila APCs transmitting xenogenic viruses to patient CTLs. In this study, we have investigated photoreactive methods to inactivate insect viruses in APC. A clinical grade psoralen compound, 8-MOP (UVADEX) in combination with UVA treatment (5 joules/cm2) can be used to inactivate Drosophila cell viruses. UVADEX treatment is sufficient to inactivate insect viruses but does not affect the expression of MHC class I molecules and costimulation molecules on Drosophila APCs. In fact, UVADEX treatment prevents Drosophila APC growth while maintaining APC function. Furthermore, UVADEX-treated Drosophila APCs maintain or have enhanced APC function as determined by enhanced T cell activation, proliferation, and CTL generation. Thus, the use of UVADEX-treated Drosophila APCs may provide a valuable tool for immunotherapy to generate tumor antigen-specific CTLs.

Girdin regulates the proliferation and apoptosis of pancreatic cancer cells via the PI3K/Akt signalling pathway.

Girdin functions as an Akt phosphorylation enhancer (APE), which expedites the proliferation and survival of many types of tumours. However, the influence of Girdin on pancreatic cancer and the underlying molecular mechanisms have yet to be uncovered. Hence, in the present study, we sought to elucidate the function of Girdin in pancreatic cancer malignancy, particularly its role in pancreatic cancer cell proliferation, migration and apoptosis. Immunohistochemistry (IHC) was used to evaluate Girdin expression in pancreatic cancer tissues and to analyse its correlation with pathological grade. Girdin expression was further validated in pancreatic cancer cell lines (AsPC­1, BxPC­3 and PANC­1), and human pancreatic ductal epithelial (HPNE) cells were used as a control. Recombinant adenovirus vectors containing Girdin­siRNA were constructed to inhibit Girdin expression and were used in subsequent experiments to determine the effects of Girdin silencing on pancreatic cancer cells. Girdin silencing suppressed pancreatic cancer cell proliferation and induced pancreatic cancer cell apoptosis in vitro and in vivo. According to the results of further mechanistic investigations, Girdin may regulate cell processes through the phosphatidylinositol­3­kinase/protein kinase B (PI3K/Akt) signalling pathway to exert additive effects on pancreatic cancer.

Combination of PI3K/Akt Pathway Inhibition and Plk1 Depletion Can Enhance Chemosensitivity to Gemcitabine in Pancreatic Carcinoma.

The prognosis of pancreatic cancer (PC) remains pessimistic because of the difficulty in early diagnosis as well as the little advance in chemotherapy. Although being the first-line chemotherapy drug for PC at present, gemcitabine still has some disadvantages, such as low drug sensitivity and significant side effects. Thus, how to further improve the sensitivity of PC cells to gemcitabine is still a difficult subject in the field of pancreatic cancer-treatment. Polo-like kinase 1 (Plk1) is closely related to poor outcome in many malignant tumors and its high expression is linked to chemoresistance in PC. As a downstream gene activated by PI3K/Akt signal pathway, we assumed that the targeted depletion of Plk1 could contribute to the chemosensitization induced by synergistic drug interaction of PI3K inhibitor LY294002 together with gemcitabine. To analyze effect of Plk1 in chemotherapy, we constructed two recombinant adenoviral vectors which carry enhanced green fluorescent protein (rAd-EGFP) and Plk1-shRNA (rAd-shPlk1), respectively. Both inhibition of PI3K/Akt signal pathway through PI3K inhibitor LY294002 and targeted depletion of Plk1 via recombinant adenoviral shRNA can cause chemosensitization, and the targeted depletion of Plk1 can enhance the chemosensitization of LY294002. Thus, the gene therapy like targeted depletion of Plk1 may create new perspectives for chemosensitization of PC.

The pseudogene derived from long non-coding RNA DUXAP10 promotes colorectal cancer cell growth through epigenetically silencing of p21 and PTEN.

Recently, substantial evidence has demonstrated that pseudogene derived lncRNAs are crucial regulators of cancer development and progression. DUXAP10,a pseudogene derived long non-coding RNA(lncRNA), is overexpression in colorectal cancer (CRC), but its expression pattern, biological function and underlying mechanism in CRC is still undetermined. In this study, we observed that DUXAP10 was up-regulated in CRC tissues which was positively correlated with advanced pathological stages, larger tumor sizes and lymph node metastasis. Additionally, knockdown of DUXAP10 inhibited cell proliferation, induced cell apoptosis and increase the number of G0/G1 cells significantly in the HCT116 and SW480 cell lines. Moreover, DUXAP10 silencing inhibited tumor growth in vivo. Further mechanism study showed that, by binding to histone demethylase lysine-specific demethylase 1 (LSD1), DUXAP10 promote CRC cell growth and reduced cell apoptosis through silencing the expression of p21 and phosphatase and tensin homolog (PTEN) tumor suppressor. Our findings suggested that the pseudogene-derived from lncRNA DUXAP10 promotes the biological progression of CRC and is likely to be a potential therapeutic target for CRC intervention.

Long non-coding RNA FOXP4-AS1 is an unfavourable prognostic factor and regulates proliferation and apoptosis in colorectal cancer.

OBJECTIVES: Despite improvements in diagnosis and treatment, colorectal cancer (CRC) remains the third most common malignancy, and fourth-leading cause of cancer-related death worldwide, and has a particularly high incidence in Western countries. Recent studies have suggested that long non-coding RNAs (lncRNAs) compose a novel class of regulators of cancer biological processes, such as proliferation, apoptosis and metastasis. Here, we report that lncRNA FOXP4-AS1 acts as a functional oncogene in CRC pathogenesis. Moreover, we have attempted to investigate the effects of FOXP4-AS1 on tumour progression, both in vitro and in vivo. MATERIALS AND METHODS: In this study, bioinformatic analyses and qPCR were performed to investigate FOXP4-AS1 expression in CRC tissue samples and CRC cell lines. We inhibited FOXP4-AS1 expression via FOXP4-AS1-specific siRNA transfection. Cell proliferation was assessed using cell viability and colony formation assays, as well as by flow cytometry and ethynyl deoxyuridine (Edu) analyses. Apoptosis was assessed using flow cytometry. Animal tumour xenografts were generated, and immunohistochemistry (IHC) was performed to evaluate effects of FOXP4-AS1 on CRC tumour growth in vivo. RESULTS: We found that FOXP4-AS1 was up-regulated in CRC tissues and cell lines and that its overexpression positively correlated with advanced pathological stages and larger tumour size. Additionally, we found that FOXP4-AS1 knockdown inhibited cell proliferation and induced apoptosis. Furthermore, FOXP4-AS1 knockdown induced marked increase in number of cells in G0/G1 phase and reduction in number of cells in S phase, in DLD-1, HT-29 and HCT116 cell lines. Consistent with these findings, FOXP4-AS1 silencing inhibited tumour growth in vivo. CONCLUSION: These findings suggest that FOXP4-AS1 plays a crucial role in CRC progression and may be a new biomarker in patients with CRC.

HOTTIP: a critical oncogenic long non-coding RNA in human cancers.

Long non-coding RNAs (lncRNAs), which represent a novel group of non-protein-coding RNAs and are commonly defined as RNA molecules larger than 200 nucleotides in length, have been shown to get involved in diverse biological processes, such as cell growth, apoptosis, migration and invasion. In addition, aberrant expression of lncRNAs has been discovered in human tumors, where they function as either oncogenes or tumor suppressor genes. Recently tumorigenic effects of one specific lncRNA, termed as 'HOXA transcript at the distal tip' (HOTTIP), on the initiation and progression of human cancer has been widely reported. An increasing amount of data has shown that dysregulation of HOTTIP is associated with various malignancies including hepatocellular carcinoma, pancreatic cancer, gastric cancer and colorectal cancer, and affects the survival and prognosis of cancer patients. Here, we focus on the current knowledge of HOTTIP in various cancers and illustrate the corresponding mechanism and biological function of HOTTIP during tumor development.

ACVR1-Fc suppresses BMP signaling and chondro-osseous differentiation in an in vitro model of Fibrodysplasia ossificans progressiva.

Fibrodysplasia ossificans progressiva (FOP) is a rare and devastating genetic disease of heterotopic endochondral ossification (HEO), and currently no effective therapies are available for this disease. A recurrent causative heterozygous mutation (c.617 G>A; R206H) for FOP was identified in activin receptor type IA (ACVR1), a bone morphogenetic protein (BMP) type I receptor. This mutation aberrantly activates the BMP-Smad1/5/8 signaling pathway and leads to HEO in FOP patients. Here we report development of a soluble recombinant ACVR1-Fc fusion protein by combining the extracellular domain of human wild type ACVR1 and the Fc portion of human immunoglobulin gamma 1 (IgG1). The ACVR1-Fc fusion protein significantly down-regulated the dysregulated BMP signaling caused by the FOP ACVR1 mutation and effectively suppressed chondro-osseous differentiation in a previously described cellular FOP model, human umbilical vein endothelial cells (HUVECs) that were infected with adenovirus-ACVR1R206H (HUVECR206H). This ACVR1-Fc fusion protein holds great promise for prevention and treatment of HEO in FOP and related diseases.

Regulation of cell apoptosis and proliferation in pancreatic cancer through PI3K/Akt pathway via Polo-like kinase 1.

Pancreatic cancer has a poor prognosis. It is reported that the PI3K/Akt pathway is activated in many cancers, and inhibition of the PI3K/Akt pathway can induce cell apoptosis in most cancers. Polo-like kinase 1 (Plk1) is also overexpressed in most malignancies, and it controls multiple aspects of mitosis and apoptosis. Previous studies identified that PI3K/Akt-dependent phosphorylation of Plk1-Ser99 is required for metaphase-anaphase transition. In this study, we aimed to investigate the molecular mechanism of PI3K/Akt pathway regulating cell proliferation and apoptosis in pancreatic cancer cell lines (AsPC-1, BxPC-3, PANC-1). Immunohistochemistry (IHC) was used to assess Akt levels in human pancreatic tissues and pancreatic cancer tissues. MTT assay was used to detect cell proliferation. The mRNA was quantified by quantitative reverse transcription-PCR. Western blot analysis was used to detect the protein levels of p-Akt, Akt, Plk1, BAX, Bcl-2, XIAP, cleaved caspase-3 and caspase-3. Recombinant adenovirus vector containing Plk1-shRNA was constructed to inhibit Plk1 expression. Cell apoptosis was detected by flow cytometry and the apoptosis of tumor xenograft was assessed by TUNEL assay. The study showed that inhibition of PI3K/Akt pathway can induce cell apoptosis and reduce cell proliferation by downregulating Plk1 in vitro and in vivo. Additionally, Plk1 inhibition can lead to cancer cell apoptosis through inactivating XIAP, activating caspase-3, upregulating BAX and downregulating Bcl-2. Therefore, this study provided the molecular mechanism of PI3K/Akt pathway and Plk1 in the pancreatic cancer cell proliferation and apoptosis, which may benefit for the therapy of pancreatic cancer.

CD8 T cells and aging.

Aging of the immune system, or "immunosenescence," is associated with both a marked reduction in responsiveness as well as functional dysregulation. These changes have been implicated in the increased morbidity and mortality of the elderly population from infectious disease, and may also play a role in autoimmunity and cancer. Though marginal alterations in B lymphocytes are apparent, the dramatic decline in humoral and cell-mediated responses is predominantly the consequence of alterations in the T-cell compartment. The effect of aging on CD4 cell function has been extensively summarized elsewhere. This review, therefore, focuses on the CD8 T-cell subset. Age-related changes in thymic function and involution, cellular homeostasis and lifespan, population shifts, T-cell activation, the process of replicative senescence, and oligoclonal expansions are discussed in terms of their effect on CD8 T cells. Age-associated alterations in CD4 T cells and antigen-presenting cells are mentioned insofar as these cells affect CD8 T-cell activation and function. Distinct patterns of immunosenescence in humans and mice are also noted.

Early antigen-specific response by naive CD8 T cells is not altered with aging.

Both a dramatic decline in CD8 responses and a switch to memory T cell predominance occur with aging. The extent to which the loss of responsiveness is the consequence of the accumulation of more differentiated vs intrinsically defective T cells (or both) has been unclear. Using similar conditions of Ag stimulation, we have examined the responses generated by CD8(+) cells isolated from aged TCR transgenic mice. We found that the naive transgene(+) CD8(+) cells from aged 2C mice expressed activation markers, produced IL-2, proliferated, and differentiated into cytotoxic T cells as efficiently as their young counterparts. The extent of responsiveness and the level of the responses were comparable in both age groups regardless of the stimulatory conditions used, i.e., partial costimulation/adhesion molecule expression on APCs, or presentation of lower affinity peptide or diminished peptide concentrations. By day 4 after Ag stimulation, no significant age-related differences were observed in the number of effector cells generated nor in the levels of secreted IL-2 or IFN-gamma. Upon restimulation of effector cells, IL-2 secretion and to a lesser extent TNF-alpha expression, but not IFN-gamma secretion, were diminished with age. These findings suggest that age-associated alterations in naive CD8 cell function are not found after primary stimulation, but may become apparent upon restimulation.