Prediction of clinical pharmacokinetics of AMG 181, a human anti-α 4 β 7 monoclonal antibody for treating inflammatory bowel diseases.

The purpose of this study was to predict a safe starting dose of AMG 181, a human anti-α 4 β 7 antibody for treating inflammatory bowel diseases, based on cynomolgus monkey pharmacokinetic (PK) and pharmacodynamic (PD) data. A two-compartment model with parallel linear and target-mediated drug disposition for AMG 181 PK in cynomolgus monkey was developed. The estimated parameters were allometrically scaled to predict human PK.

Janus kinase 3 inhibition with CP-690,550 prevents allograft vasculopathy.

Janus kinase 3 (JAK3) mediates signal transduction from cytokine receptors using the common gamma chain. The rationally designed inhibitor of JAK3, CP-690,550, prevents acute allograft rejection in rodents and in nonhuman primates. Here we investigated the ability of CP-690,550, to prevent allograft vasculopathy in a rodent model of aorta transplantation.

Similarities in the immunoglobulin response and VH gene usage in rhesus monkeys and humans exposed to porcine hepatocytes.

Background: The use of porcine cells and organs as a source of xenografts for human patients would vastly increase the donor pool; however, both humans and Old World primates vigorously reject pig tissues due to xenoantibodies that react with the polysaccharide galactose alpha (1,3) galactose (alphaGal) present on the surface of many porcine cells. We previously examined the xenoantibody response in patients exposed to porcine hepatocytes via treatment(s) with bioartficial liver devices (BALs), composed of porcine cells in a support matrix. We determined that xenoantibodies in BAL-treated patients are predominantly directed at porcine alphaGal carbohydrate epitopes, and are encoded by a small number of germline heavy chain variable region (VH) immunoglobulin genes.

Combined use of the JAK3 inhibitor CP-690,550 with mycophenolate mofetil to prevent kidney allograft rejection in nonhuman primates.

Background: Immunosuppression via Janus kinase (JAK) 3 inhibition affords significant prolongation of allograft survival. We investigated the effects of an immunosuppressive regimen combining the JAK3 inhibitor CP-690,550 with mycophenolate mofetil (MMF) in nonhuman primates (NHPs).

Methods: Life-supporting kidney transplantations were performed between ABO-compatible, MLR-mismatched NHPs.

Effects of JAK3 inhibition with CP-690,550 on immune cell populations and their functions in nonhuman primate recipients of kidney allografts.

Background: Janus Kinase (JAK) 3 is a tyrosine kinase essential for proper signal transduction downstream of selected cytokine receptors and for robust T-cell and natural killer cells activation and function. JAK3 inhibition with CP-690,550 prevents acute allograft rejection. To provide further insight into the mechanisms of efficacy, we investigated the immunomodulatory effects of CP-690,550 in vitro and in vivo in nonhuman primates.

Quantitative analysis of the immunosuppressant CP-690,550 in whole blood by column-switching high-performance liquid chromatography and mass spectrometry detection.

A fast and accurate method to quantify the new immunosuppressive JAK3 inhibitor CP-690,550 in whole blood using a dual-pump liquid chromatography-liquid chromatography-mass spectrometry (LC/LC-MS) system was developed and validated in nonhuman primate blood. Before injection, blood samples were prepared by precipitation with a reagent that included methanol and acetonitrile (30:70, vol/vol) along with the internal standard (CP-istd). Column-switching LC/LC-MS analysis used online extraction followed by separation on a C8 analytic column and MS detection of the [M + H] CP-690,550 (m/z = 313.

Immunosuppressive effects of surgery assessed by flow cytometry in nonhuman primates after nephrectomy.

Despite previous studies suggesting that surgery cause immune suppression, the underlying biologic mechanisms have not been studied using advanced immune function assays. Unilateral nephrectomy was performed in nonhuman primates. Blood was collected before surgery and at different time-points through 14 days after surgery.

Identification of the V genes encoding xenoantibodies in non-immunosuppressed rhesus monkeys.

The major immunological barrier that prevents the use of wild-type pig xenografts as an alternative source of organs for human xenotransplantation is antibody-mediated rejection. In this study, we identify the immunoglobulin variable region heavy (IgV(H)) chain genes encoding xenoantibodies to porcine heart and fetal porcine islet xenografts in non-immunosuppressed rhesus monkeys. We sought to compare the IgV(H) genes encoding xenoantibodies to porcine islets and solid organ xenografts.

Appraisal of the extent of chronic allograft vasculopathy in animal models: proposition of a standardized micromorphometric method.

Arterial obstruction due to intimal myproliferation is an outcome used for investigating chronic allograft vasculopathy in animal models. Since harvested tissue may be distorted while processing, discrepancy may happen in the measure of the obstruction. We propose a standardized micromorphometric method for calculating the percentage of obstruction with eliminating the variability in the shape of the vessel cross-section.

Effects of the kinase inhibitor CGP41251 (PKC 412) on lymphocyte activation and TNF-alpha production.

CGP41251 is a serine/threonine and tyrosine kinase inhibitor that is a novel anticancer agent. Because the kinases that CGP41251 inhibits play important roles in T lymphocyte activation, we hypothesized that this compound may have useful immunomodulatory properties. Here we characterized the in vitro immunomodulatory effects of CGP41251.

A technique of bone marrow collection from vertebral bodies of cynomolgus macaques for transplant studies.

Background: Strategies to induce donor-specific allograft tolerance are best tested in preclinical models developed in nonhuman primates (NHPs). Most protocols prepare the recipient by infusing hematopoietic cells from the donor. We report here a procedure to isolate and characterize large numbers of bone marrow cells (BMCs) from cynomolgus monkeys (cynos) that can then successfully be transplanted into conditioned recipients.

Immunosuppression by the JAK3 inhibitor CP-690,550 delays rejection and significantly prolongs kidney allograft survival in nonhuman primates.

Background: Janus kinase 3 (JAK3) mediates signal transduction from cytokine receptors using the common chain (gammac). Because mutations in genes encoding gammac or JAK3 result in immunodeficiency, we investigated the potential of a rationally designed inhibitor of JAK3, CP-690,550, to prevent renal allograft rejection in nonhuman primates.

Methods: Life-supporting kidney transplantations were performed between mixed leukocyte reaction-mismatched, ABO blood group-matched cynomolgus monkeys.

Improved assessment of graft function by echocardiography in cynomolgus monkey recipients of hDAF-transgenic pig cardiac xenografts.

Background: The current practice of evaluating heterotopic heart xenografts by palpation allows only detection of severe graft dysfunction, which indicates terminal graft failure. Therefore, we evaluated whether echocardiography is a better method of detecting early graft dysfunction as a marker of rejection in abdominal pig heart xenografts in cynomolgus monkeys.

Methods: Six cynomolgus monkeys received heterotopic heart transplants from pig donors transgenic for human decay-accelerating factor (hDAF).

Outbreak of Mycobacterium bovis in a conditioned colony of rhesus (Macaca mulatta) and cynomolgus (Macaca fascicularis) macaques.

We describe a tuberculosis outbreak caused by Mycobacterium bovis in a conditioned colony of rhesus (Macaca mulatta) and cynomolgus (Macaca fascicularis) macaques. Animals in five rooms were exposed, but most (16/27) infections were confined to the room that housed a mixed population of cynomolgus and rhesus macaques. In this room, rhesus (8/8) and cynomolgus (10/11) macaques naturally exposed to M.

Inhibition of lymphocyte activation and function by the prenylation inhibitor L-778,123.

Prenylated Ras GTPases transduce signals from the T cell receptor, CD28 costimulatory receptor and IL-2 receptor. Since signals from these receptors mediate T cell activation, proliferation and survival, we hypothesized that the prenylation inhibitor L-778,123 would impart immunomodulation. The effect of L-778,123 on T cell activation (CD71 or CD25 surface expression) was determined by flow cytometry.

JAK3 inhibition, a viable new modality of immunosuppression for solid organ transplants.

The field of organ transplantation has had tremendous success because of the availability of immunosuppressive drugs that efficiently prevent acute organ rejection. Numerous and severe side effects are, however, associated with all current immunosuppressive therapies and justify a search for drugs with better efficacy and safety profiles. Janus kinase (JAK) 3, a tyrosine kinase that is crucial for mediating signals from the common gamma-chain of cytokine receptors, is peculiar in that its expression, contrarily to the targets of most current immunosuppressive drugs, is limited to cells that actively participate to the immune response to allografts.

Compared with cyclosporine, ISATX247 significantly prolongs renal-allograft survival in a nonhuman primate model.

Background: ISATX247 is a novel calcineurin inhibitor that has shown more potency than cyclosporine in vitro. This is the first study to compare the survival times of renal allografts in nonhuman primates treated with either ISATX247 or cyclosporine.

Methods: Adult, male cynomolgus monkeys were divided into blood-group compatible and mixed-lymphocyte, stimulation-mismatched, donor-recipient pairs.

In vitro evaluation of the effects of candidate immunosuppressive drugs: flow cytometry and quantitative real-time PCR as two independent and correlated read-outs.

Background: Immune monitoring may use flow cytometry or molecular biology techniques. Flow cytometry assays cells that are phenotypically characterized, whereas TaqMan RT-PCR starts with RNA extraction from unfractionated heterogeneous cell populations. We therefore wondered how the effects of immunosuppressive drugs on cytokine production in stimulated whole blood, as determined by flow cytometry, would correlate with those obtained with quantitative real-time PCR (TaqMan RT-PCR).

JAK3 inhibition as a new concept for immune suppression.

Although current immunosuppressive drugs are effective, they have numerous severe side effects that mandate the search for new agents. Mutations in the gene for janus kinase (JAK)3 result in severe combined immune deficiency with severely impaired humoral and cellular immunity, an observation that has prompted the development of JAK3 inhibitors. Due to its central role in lymphocyte activation, proliferation and homeostasis, targeting the JAK/signal transducer and activator of transcription (STAT) pathway may provide the required efficacy, without the toxicities associated with current therapies.

Prevention of organ allograft rejection by a specific Janus kinase 3 inhibitor.

Because of its requirement for signaling by multiple cytokines, Janus kinase 3 (JAK3) is an excellent target for clinical immunosuppression. We report the development of a specific, orally active inhibitor of JAK3, CP-690,550, that significantly prolonged survival in a murine model of heart transplantation and in cynomolgus monkeys receiving kidney transplants. CP-690,550 treatment was not associated with hypertension, hyperlipidemia, or lymphoproliferative disease.

Immunomodulatory effects of docetaxel on human lymphocytes.

Docetaxel is an antineoplastic taxoid that interferes with microtubule polymerization dynamics and is used clinically to treat advanced cancers. Because microtubules play significant roles in T lymphocyte activation and function we characterized the in vitro immunomodulatory properties of docetaxel. Effects of docetaxel on lectin-induced peripheral blood mononuclear cell (PBMC) proliferation were measured by the 3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium (MTS) assay and proliferating cell nuclear antigen (PCNA) staining.