Tumor-infiltrating programmed death receptor-1+ dendritic cells mediate immune suppression in ovarian cancer

Within the ovarian cancer microenvironment, there are several mechanisms that suppress the actions of antitumor immune effectors. Delineating the complex immune microenvironment is an important goal toward developing effective immune-based therapies. A dominant pathway of immune suppression in ovarian cancer involves tumor-associated and dendritic cell (DC)-associated B7-H1. The interaction of B7-H1 with PD-1 on tumor-infiltrating T cells is a widely cited theory of immune suppression involving B7-H1 in ovarian cancer. Recent studies suggest that the B7-H1 ligand, programmed death receptor-1 (PD-1), is also expressed on myeloid cells, complicating interpretations of how B7-H1 regulates DC function in the tumor. In this study, we found that ovarian cancer-infiltrating DCs progressively expressed increased levels of PD-1 over time in addition to B7-H1. These dual-positive PD-1(+) B7-H1(+) DCs have a classical DC phenotype (i.e., CD11c(+)CD11b(+)CD8(-)), but are immature, suppressive, and respond poorly to danger signals. Accumulation of PD-1(+)B7-H1(+) DCs in the tumor was associated with suppression of T cell activity and decreased infiltrating T cells in advancing tumors. T cell suppressor function of these DCs appeared to be mediated by T cell-associated PD-1. In contrast, ligation of PD-1 expressed on the tumor-associated DCs suppressed NF-κB activation, release of immune regulatory cytokines, and upregulation of costimulatory molecules. PD-1 blockade in mice bearing ovarian cancer substantially reduced tumor burden and increased effector Ag-specific T cell responses. Our results reveal a novel role of tumor infiltrating PD-1(+)B7-H1(+) DCs in mediating immune suppression in ovarian cancer.     

Effects of selected fertilizers on the life history of Bemisia tabaci (Hemiptera: Aleyrodidae) biotype B

We tested the effects among a purportedly sustainable water-soluble fertilizer, a conventional water-soluble fertilizer, an alternation of these, a controlled-release fertilizer, and a clear water control on the life-history traits of sweetpotato whitefly, Bemisia tabaci (Gennadius) (Hemiptera: Aleyrodidae; =Bemisia argentifolii Bellows & Perring) biotype B reared on poinsettia (Euphorbia pulcherrima Willdenow ex Klotzch). Free amino acids in petioles were measured to estimate plant nutrient assimilation and phloem nutritional quality for B. tabaci biotype B. The sustainable fertilizer produced plants with the highest concentration of amino acids. In contrast, fecundity of whiteflies was lowest in plants treated with the sustainable fertilizer and the water control. The relationship between total amino acids in phloem and survival was significantly quadratic, with the highest survival at intermediate levels. Fecundity, however, was negatively correlated with total amino acid content of the maternal host plant. Variation in total amino acid concentration in petioles of plants treated within fertilizer treatments makes it difficult to predict whether a particular fertilizer will produce plants with enough amino acids to deleteriously affect both survivorship and fecundity and yet yield a plant of good quality. Despite this limitation, we can conclude that the use of this sustainable fertilizer will not cause increases in whitefly populations relative to plants fertilized with water-soluble and slow-release fertilizers that deliver the same level of nitrogen to the plant.     

Evacetrapib is a novel, potent, and selective inhibitor of cholesteryl ester transfer protein that elevates HDL cholesterol without inducing aldosterone or increasing blood pressure

Cholesteryl ester transfer protein (CETP) catalyses the exchange of cholesteryl ester and triglyceride between HDL and apoB containing lipoprotein particles. The role of CETP in modulating plasma HDL cholesterol levels in humans is well established and there have been significant efforts to develop CETP inhibitors to increase HDL cholesterol for the treatment of coronary artery disease. These efforts, however, have been hampered by the fact that most CETP inhibitors either have low potency or have undesirable side effects. In this study, we describe a novel benzazepine compound evacetrapib (LY2484595), which is a potent and selective inhibitor of CETP both in vitro and in vivo. Evacetrapib inhibited human recombinant CETP protein (5.5 nM IC(50)) and CETP activity in human plasma (36 nM IC(50)) in vitro. In double transgenic mice expressing human CETP and apoAI, evacetrapib exhibited an ex vivo CETP inhibition ED(50) of less than 5 mg/kg at 8 h post oral dose and significantly elevated HDL cholesterol. Importantly, no blood pressure elevation was observed in rats dosed with evacetrapib at high exposure multiples compared with the positive control, torcetrapib. In addition, in a human adrenal cortical carcinoma cell line (H295R cells), evacetrapib did not induce aldosterone or cortisol biosynthesis whereas torcetrapib dramatically induced aldosterone and cortisol biosynthesis. Our data indicate that evacetrapib is a potent and selective CETP inhibitor without torcetrapib-like off-target liabilities. Evacetrapib is currently in phase II clinical development.     

Canonical NF-kappaB activation is essential for Epstein-Barr virus latent membrane protein 1 TES2/CTAR2 gene regulation

Epstein-Barr virus (EBV) latent membrane protein 1 (LMP1) transforms rodent fibroblasts and is expressed in most EBV-associated malignancies. LMP1 (transformation effector site 2 [TES2]/C-terminal activation region 2 [CTAR2]) activates NF-κB, p38, Jun N-terminal protein kinase (JNK), extracellular signal-regulated kinase (ERK), and interferon regulatory factor 7 (IRF7) pathways. We have investigated LMP1 TES2 genome-wide RNA effects at 4 time points after LMP1 TES2 expression in HEK-293 cells. By using a false discovery rate (FDR) of <0.001 after correction for multiple hypotheses, LMP1 TES2 caused >2-fold changes in 1,916 mRNAs; 1,479 RNAs were upregulated and 437 were downregulated. In contrast to tumor necrosis factor alpha (TNF-α) stimulation, which transiently upregulates many target genes, LMP1 TES2 maintained most RNA effects through the time course, despite robust and sustained induction of negative feedback regulators, such as IκBα and A20. LMP1 TES2-regulated RNAs encode many NF-κB signaling proteins and secondary interacting proteins. Consequently, many LMP1 TES2-regulated RNAs encode proteins that form an extensive interactome. Gene set enrichment analyses found LMP1 TES2-upregulated genes to be significantly enriched for pathways in cancer, B- and T-cell receptor signaling, and Toll-like receptor signaling. Surprisingly, LMP1 TES2 and IκBα superrepressor coexpression decreased LMP1 TES2 RNA effects to only 5 RNAs, with FDRs of <0.001-fold and >2-fold changes. Thus, canonical NF-κB activation is critical for almost all LMP1 TES2 RNA effects in HEK-293 cells and a more significant therapeutic target than previously appreciated.     

Aberrant epigenetic and genetic marks are seen in myelodysplastic leukocytes and reveal Dock4 as a candidate pathogenic gene on chromosome 7q

Myelodysplastic syndromes (MDS) are characterized by abnormal and dysplastic maturation of all blood lineages. Even though epigenetic alterations have been seen in MDS marrow progenitors, very little is known about the molecular alterations in dysplastic peripheral blood cells. We analyzed the methylome of MDS leukocytes by the HELP assay and determined that it was globally distinct from age-matched controls and was characterized by numerous novel, aberrant hypermethylated marks that were located mainly outside of CpG islands and preferentially affected GTPase regulators and other cancer-related pathways. Additionally, array comparative genomic hybridization revealed that novel as well as previously characterized deletions and amplifications could also be visualized in peripheral blood leukocytes, thus potentially reducing the need for bone marrow samples for future studies. Using integrative analysis, potentially pathogenic genes silenced by genetic deletions and aberrant hypermethylation in different patients were identified. DOCK4, a GTPase regulator located in the commonly deleted 7q31 region, was identified by this unbiased approach. Significant hypermethylation and reduced expression of DOCK4 in MDS bone marrow stem cells was observed in two large independent datasets, providing further validation of our findings. Finally, DOCK4 knockdown in primary marrow CD34(+) stem cells led to decreased erythroid colony formation and increased apoptosis, thus recapitulating the bone marrow failure seen in MDS. These findings reveal widespread novel epigenetic alterations in myelodysplastic leukocytes and implicate DOCK4 as a pathogenic gene located on the 7q chromosomal region.     

Altered contribution of RhoA/Rho kinase signaling in contractile activity of myometrium in leptin receptor-deficient mice

In late gestation, enhanced myometrial contractility is mediated in part through increased Rho/Rho kinase. Since leptin, which is elevated in pregnancy and obesity, can directly depress myometrial function, we hypothesized that in leptin receptor-deficient mice, myometrial contractility would be greater in late pregnancy due to increased Rho/Rho kinase activity. To test this, we correlated RhoA and Rho kinase expression to contractility in myometrium from nonpregnant (NP) and late-pregnant (P18) heterozygous leptin receptor-deficient mice (db/+) vs. wild-type (WT) mice. In NP mice, KCl-induced contractions were similar between WT and db/+ myometrium. However, the Rho kinase-dependent component of the contractions was greater in db/+ mice, along with an increased expression of Rho kinase. KCl-induced contractions increased in strength in myometrium from P18 WT and db/+ compared with NP. Although the contribution of Rho kinase to contractions was unchanged in P18 WT mice, it was decreased in P18 db/+ mice. The decrease in Rho kinase-dependent contractions in P18 db/+ mice coincided with reduced RhoA and Rho kinase expression relative to NP db/+. Addition of high-fat-induced abnormal glucose utilization prevented changes in Rho kinase function. We conclude that abnormal leptin signaling increases expression and function of Rho kinase to maintain contractile function in NP myometrium and that during pregnancy the contribution of RhoA and Rho kinase expression to myometrial function is reduced despite an increase in myometrial contractility. Thus, other signaling mechanisms appear to compensate when leptin signaling is reduced to maintain contractile function during pregnancy.     

The scarless heart

Over the past several years many mechanisms by which myocardial replacement could be achieved have been described. These include resident cardiac stem cells or circulating stem cells that can either differentiate into, or fuse to cardiomyocytes, or mature cells that can transdifferentiate into cardiomyocytes. However, the fact remains that after injury to the heart, the overriding response is scar formation with little myocardial replacement. One exception to this response is the MRL mouse, which heals with little scarring and shows nearly full myocardial replacement after injury. Results obtained with this model will be discussed.     

Interrelationships among plasma progesterone concentrations, luteal anatomy and function, and placental ontogeny during gestation in a viviparous lizard (Niveoscincus metallicus: Scincidae)

Plasma progesterone concentrations were measured at six stages of gestation in the viviparous lizard Niveoscincus metallicus. Anatomical and functional parameters of luteal activity were also investigated. The diameter of the corpus luteum (CL) decreased gradually though gestation, as did the diameter of the luteal cells. Major degenerative changes were observed in CLs post-partum. Plasma progesterone concentrations were basal both prior to, and just after, ovulation; a rapid increase occurred in early gestation. Plasma progesterone concentrations remained elevated until late gestation, but fell some 2 weeks before parturition. In vitro production of progesterone was greater in CLs in mid- than in late-gestation, and the addition of prostaglandin F(2alpha) to the incubation medium had no effect on progesterone production. Non-luteal ovarian tissue and adrenals produced progesterone, but at approximately one-tenth the rate of production by CLs. Temporal correlations between the plasma progesterone profile and stages of placental development were also assessed. The rise in plasma progesterone concentrations occurs before differentiation of the chorioallantoic placenta, but progesterone is still high when it degenerates. We conclude that the CLs are the major source of gestational progesterone in N. metallicus.