Ex vivo stimulation

In a Phase la study in healthy volunteers, ARRY-438162 was well tolerated up to 20mg/kg q.d. oral dose. There was a dose proportional increase in plasma concentration and decrease in the production of IL-1 ft, TNFa, IL-6 and pERK in the ex-vivo-stimulated whole blood from drug treated volunteers. Array has initiated a Phase lb study of ARRY-438162 in combination with methotrexate in rheumatoid arthritis patients with the goals of assessing safety, tolerability, pharmacokinetics (PK), biomarkers and initial signs of efficacy. 

Inhibition of COX can be quantified in recombinant or natural enzyme preparations, cellular systems, isolated human cell populations such as platelets (COX-1) and white blood cells (COX-2), or in ex vivo stimulated whole blood samples The closer the experimental system is to the physiological state, the lower the selectivity for most COX-2-inhibitors. The standard test for comparison is considered to be a whole blood assay which mimics in vivo conditions like plasma binding (e.g. Patrignani et al., 1996). It is commonly accepted that reasonable variations occur between different laboratories (see data for Piroxicam). Therefore, whenever possible, data of several compounds generated with a given test system should be compared with each other. 

Freshly isolated spleens and draining lymph nodes can also be used for ex vivo stimulation with collagen type II or other stimuli of interest. These procedures are described in several research articles (7-9). 

Fleck C, Bachner B, Gockeritz S, et al. Ex vivo stimulation of renal tubular PAH transport by dexamethasone and triiodothyronine in human renal cell carcinoma. Urol Res 2000 28 383-390. 

Figure 6 Proliferation of T cells from mice immunized with nanoparticulate of soluble formulations in response to ex vivo stimulation with the recall antigen (tetanus toxoid). The T cells were derived from the mice immunized with TT and various doses of CpG ODN. The number of T cells and APCs incubated in the assay plate wells were 5x10 and 1x10 respectively. The recall proliferation response is shown on the y-axis as (A) CPM for clarity SI values are also given on the top of each bar (B) stimulation index for nanoparticulate (solid circles) and soluble (hollow circles) mode of vaccine delivery. Error bars indicate standard deviation. (From Ref. 135.)...

We modified polyanionic polymers by use of a grafting reaction of hydrophobic groups onto the polymers. After an extensive evaluation for the affinity of the hy-drophobically modified (hydrophobized) polymers to cell membrane, the immuno-stimulating activity of polymers was investigated by in vitro or ex vivo experiments. Consequently, the increased biological activity was found in the hydrophobized polymer, indicating that... 

For additional evaluation of the effect of hydrophobization and the molecular weight of the polymers on the biological immuno-stimulating activity, we investigated the ex vivo cytokine (interIeukin-6 [IL-6], and tumor necrosis factor [TNFj-inducing activity from human peripheral whole blood cells of hydrophobized polymers by use of fractionated poly(M A-CDA) with narrow poly-dispersity. Since this assay uses the intact human cells, it shows more accurate results than in vitro assay using cultured cell line [25]. 

In 1995, HlV-1 latency was first documented in HIV-1-infected patients when ex vivo T cell cultures were found to contain a subpopulation of cells that produced infectious virions when stimulated with T cell activators (Chun et al. 1995 Finzi et al. 1997). Latently infected T cells are rare, to the order of one in a million resting... 

Several of the postulated roles for nematode-secreted AChEs assume that they gain access to the intestinal mucosa. Several possibilities exist for transport of parasite AChE across the epithelial cell barrier, such as (i) utilization of existing pathways for receptor-mediated transcytosis (ii) a paracellular route facilitated by parasite-secreted proteases as observed for a bacterial elastase (Azghani et al., 1993) and (iii) increased paracellular permeability resulting from inflammatory events in the mucosa. We consider the latter suggestion most likely, as this has been duplicated by ex vivo perfusion with rat mast cell protease II (Scudamore et al., 1995). Moreover, cholinergic stimulation attenuates epithelial barrier properties to macromolecules in rat ileal crypts (Phillips et al., 1987). 

LOX-dependent superoxide production was also registered under ex vivo conditions [55]. It has been shown that the intravenous administration of lipopolysaccharide to rats stimulated superoxide production by alveolar and peritoneal macrophages. O Donnell and Azzi [56] proposed that a relatively high rate of superoxide production by cultured human fibroblasts in the presence of NADH was relevant to 15-LOX-catalyzed oxidation of unsaturated acids and was independent of NADPH oxidase, prostaglandin H synthase, xanthine oxidase, and cytochrome P-450 activation or mitochondrial respiration. LOX might also be involved in the superoxide production by epidermal growth factor-stimulated pheochromo-cytoma cells [57]. 

Li+, at therapeutically relevant concentrations, is a potent inhibitor of norepinephrine-stimulated adenylate cyclase activity ex vivo in both rat [133] and human brain [134], and it inhibits norepinephrine-stimulated cAMP accumulation in Li+-treated patients. Li+ also inhibits dopamine-stimulated cAMP accumulation in rat brain [135]. These inhibitory effects of Li+ have been shown to be region specific within rat brain, a fact that has obvious significance for a therapeutic mechanism of action. It is interesting that other antimanic drugs may also have dampening effects on dopaminergic neurotransmission. 

Ex vivo studies have revealed that trichothecenes can both inhibit and stimulate leukocyte function.12 For example, trichothecenes are toxic to alveolar macrophages,13 but drive differentiation of human myeloid leukemic cells.14 Dose-dependent decreases or increases in B- and T-cell mitogen responses are observable in lymphocytes from animals exposed to T-2 toxin, DON, or various macrocyclic trichothecenes these toxins similarly impair or enhance mitogen-induced lymphocyte proliferation in vitro.12 Rank order of inhibitor potency in rodent and human lymphocyte proliferation assays is Type D > Type A group > Type B group and is dependent on degree of acylation as well as of uptake and metabolism. 

Intracellular messengers A biphasic effect of ginkgo extract is seen on cAMP phosphodiesterase under in vitro and ex vivo conditions (Saponara and Bosisio 1998 Macovschi et al. 1987). Whereas low concentrations (0.25-4.0 mg/L) activate the enzyme, higher concentrations (5-250 mg/L), dose-dependently inhibit it. However, tolerance develops to this effect because it is undetectable after daily administration for 4 days. Thus, ginkgo may initially produce effects by inhibiting enzymatic breakdown of cAMP. This mechanism is similar to the stimulant caffeine, but it is not likely to explain any long-term effects of ginkgo because it disappears after chronic daily treatment. The responsible constituent for this effect has not been identified. 

Amri H, Drieu K, Papadopoulos V. (1997). Ex vivo regulation of adrenal cortical cell steroid and protein synthesis, in response to adrenocorticotropic hormone stimulation, by the Ginkgo biloba extract EGb 761 and isolated ginkgolide B. Endocrinology. 138(12) 5415-26. 

In ex vivo studies in which transmitter release is measured in synaptosome or slice preparations from animals treated with nicotine in vivo, no change (Grilli et al. 2005), decreased (Grady et al. 1997) and increased (Yu and Wecker 1994) striatal [ HJdopamine release has been reported. A similar picture has been observed for nAChR-evoked hippocampal [ H]noradrenaline release, with unchanged (Barik and Wonnacott 2006), decreased (Grilli et al. 2005) and increased (Jacobs et al. 2002) responses. As tissue is extensively washed prior to nAChR stimulation, these discrepancies are unlikely to reflect differences in nAChR desensitisation due... 

Although this study provided insight into the clonal composition of human HSC hierarchy, it was unknown if the ex vivo manipulations required for retroviral marking had quantitative or qualitative effects on SRC fates. The developments in lentivector transduction protocols enabled efficient transduction of SRC in the absence of cytokine and serum stimulation . Using minimal ex vivo manipulation to mark hematopoietic cells with lentiviral integration, the fates of SRC were evaluated . As reported previously using retroviral-marking, lentivector transduced short-term SRC contributed rapidly... 

Obtained results suggest that stromal cells play important role in long term maintenance of hematopoiesis ex vivo. They induce proliferation and partially inhibit terminal differentiation and prolong hematopoiesis in long term in vitro cultures. Feeder layer derived from human embryos was adequate model for AC133+ cultures and had stimulating effect on the proliferation of the progenitor cells in vitro. 

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