Lipidomics Screening of Celecoxib in ex Vivo Human Whole Blood Assay - Part B
Condition: Healthy
Study type: Interventional

Summary: 
      Cardiovascular complications of NSAIDs, selective for inhibition of COX-2, stimulated
      interest in microsomal prostaglandin E synthase-1 (mPGES-1) as an alternative drug target.
      Global deletion of mPGES-1 in mice suppresses prostaglandin (PG) E2 and augments PGI2 by PGH2
      substrate rediversion. Unlike COX-2 inhibition or gene deletion, mPGES-1 deletion does not
      cause a predisposition to thrombogenesis and hypertension. However, cell-specific deletion of
      mPGES-1 reveals that the predominant substrate rediversion product among the prostaglandins
      varies by cell type, complicating drug development. The research team has developed an ultra
      performance liquid chromatography/ tandem mass spectrometry (UPLC-MS/MS) technique that
      allows the quantification of a wide range of lipids beyond the prostaglandin pathway
      (leukotrienes, anandamide and the 2-arachidonylglycerol cascades).

      This study is designed to examine different pathway interventions from the arachidonic acid
      cascade by anti-inflammatory compounds (with a focus on mPGES-1 inhibition) in whole human
      blood in vitro (Part A) and ex vivo (Part B). In Part B, healthy volunteers will be asked to
      take a single, therapeutic dose of celecoxib and blood and urine samples will be collected
      before and after drug administration. Collected blood will be stimulated ex vivo, and lipids
      and their metabolites will be measured in blood and urine, respectively. The investigators
      expect that lipid profile from ex vivo hWBA done on celecoxib-treated subjects will
      recapitulate findings from the in vitro hWBA received with celecoxib-treated human blood
      (Part A).
    

Detailed description: 
      Nonsteroidal anti-inflammatory drugs (NSAIDs), selective for inhibition of cyclooxygenase
      (COX)-2, alleviate pain and inflammation by suppressing COX-2-derived prostacyclin (PGI2) and
      prostaglandin (PG) E2 (1). However, eight placebo-controlled clinical trials have revealed
      that NSAIDs, designed to inhibit specifically COX-2, predispose patients to increased
      cardiovascular risks including myocardial infarction, stroke, systemic and pulmonary
      hypertension, congestive heart failure, and sudden cardiac death (1-3). The cardiovascular
      adverse effects are attributable to the suppression of COX-2-derived PGI2, a potent
      vasodilator and inhibitor of platelet activation (4; 5). The research team has shown that
      global deletion, selective inhibition or mutation of COX-2, or deletion of the receptor for
      PGI2 elevate blood pressure and accelerate thrombogenesis in mouse models (6). The
      investigators have further demonstrated that vascular COX-2 deletion predisposes mice to
      thrombosis and hypertension (7), and that selective deletion of COX-2 in cardiomyocytes leads
      to cardiac dysfunction and enhanced susceptibility to induced arrhythmogenesis (8) that may
      contribute to the heart failure and cardiac arrhythmias reported in patients taking NSAIDs
      specific for inhibition of COX-2.

      This cardiovascular hazard from NSAIDs prompted interest in the microsomal prostaglandin E
      synthase-1 (mPGES-1) as an alternative drug target. mPGES-1 is the inducible PG terminal
      synthase that acts downstream of COX-2 and catalyzes the conversion of the intermediate COX
      endoperoxide product PGH2 to PGE2 (9). The investigators have previously reported that
      similar to the interference with COX-2 expression or function, global or cell-specific
      deletion of mPGES-1 suppresses PGE2 production; but unlike with COX-2, global mPGES-1
      deficiency augments biosynthesis of PGI2 and does not predispose normo- or hyperlipidemic
      mice to thrombogenic or hypertensive events (9-11). Both suppression of PGE2 and augmentation
      of PGI2 in mPGES-1-/- mice result from the rediversion of the accumulated PGH2 substrate to
      PGI2 synthase (10). Furthermore, global deletion of mPGES-1 limits the vascular proliferative
      response to wire injury (12), retards atherogenesis and suppresses angiotensin II-induced
      abdominal aortic aneurysm formation in hyperlipidemic mice (10; 13). The research team has
      also shown that mPGES-1-deficiency does not affect ozone-induced airway inflammation or
      airway hyper-responsiveness suggesting that pharmacological inhibition of mPGES-1 and
      endoperoxide rediversion to PGD2 may not predispose patients at risk to airway dysfunction
      (14). In addition, studies by others indicate that global deletion of mPGES-1 reduces the
      post-ischemic brain infarction and neurological dysfunction in cerebral ischemia/reperfusion
      in mice (15). mPGES-1 deficiency also renders mice less susceptible to excessive inflammation
      and hypersensitivity in rodent models of analgesia (16; 17). Taken together, these findings
      suggest that pharmacological inhibition of mPGES-1 may retain anti-inflammatory effects from
      PGE2 suppression, but due to PGI2 augmentation, targeting of mPGES-1 might avoid the
      cardiovascular risks associated with selective COX-2 inhibitors.

      PGH2 substrate rediversion consequent to mPGES-1 deletion is a ubiquitous event observed at
      the cellular level and systemically (urinary prostaglandin metabolites); the profile of the
      rediversion products, however, varies by cell and tissue type, the disease model, and the
      extent of system perturbation (6; 10-14; 18-21). The investigators have shown that in mice
      deficient in mPGES-1 in endothelial cells (EC) or vascular smooth muscle cells (VSMC), PGI2
      is the predominant substrate rediversion product, whereas deletion of mPGES-1 in myeloid
      cells results in shunting of PGH2 mostly towards TxA2(11). Functionally, mice lacking mPGES-1
      in myeloid cells, exhibited a poor response to vascular injury implicating myeloid mPGES-1 as
      a cardiovascular drug target. Therefore, cell-specific mPGES-1 deletion leads to a
      differential pattern of substrate rediversion and may affect biological function of the
      system, thus complicating drug development. What is unknown is whether genetic deletion or
      pharmacological inhibition of mPGES-1 can directly (through substrate rediversion) or
      indirectly (by effects of prostaglandin rediversion products on enzyme expression or their
      further metabolism to transcellular products (22)) influence the lipidome beyond the
      prostaglandin pathway with functional consequence. For example, disruption of AA-PGE2
      metabolism might influence arachidonate product formation by the cytochrome P450 (23; 24),
      leukotriene, anandamide, 2-arachidonylglycerol (2-AG) and other cascades (25). At the
      cellular level, mPGES-1-/- macrophages, pretreated with LPS and stimulated with arachidonic
      acid (AA), exhibit a 5-fold increase in 12-HHT (12-hydroxyheptadecatrienoic acid), indicating
      substrate rediversion towards thromboxane A synthase (18). Inhibition and deletion of COX-2
      have been reported to augment metabolites of 5-lipoxygenase (5-LO) pathway 5-HETE
      (5-hydroxyeicosatetraenoic acid) and leukotrienes LTB4, LTC4, LTD4 (26-28), and metabolites
      of CYP450 cascade 14,15-DHET/EET (dihydroxyeicosatrienoic/epoxyeicosatrienoic acid) (26).
      Therefore, the substrate AA may be shunted from one pathway to the other when a particular
      branch of the cascade is pharmacologically inhibited or genetically ablated.

      Here, the research team will conduct a broad-spectrum lipidomics screening of
      anti-inflammatory drugs and drug candidates that antagonize receptors (LTC4, LTB4, EP4
      receptors) or inhibit specific components (COX-1, COX-2, mPGES-1, 5-KO, FLAP, LTA4A) of
      arachidonic acid pathway in an in vitro human whole-blood assay (hWBA).

      Preliminary in vitro results from Part A demonstrated that targeting of COX-2 with a
      selective COX-2 inhibitor celecoxib affected not only cyclooxygenase pathway but also
      lipoxygenase cascade. Celecoxib inhibited COX-derived products PGE2, PGF2a and TxB2 and
      significantly reduced levels of 15-HETE, a product of 15-LOX cascade.

      In Part B, the investigators propose to study the effect of celecoxib on plasma lipids ex
      vivo. Healthy, non-smoking, male and female volunteers will be asked to take a single,
      therapeutic dose of 200 mg of celecoxib or a placebo pill and provide blood and urine samples
      before and after the drug administration. Experiments will include (i) the ex vivo whole
      blood assay, in which lipids will be measured in blood collected before and 3 hours (Tmax)
      after administration of celecoxib and stimulated with LPS, (ii) lipid metabolites will be
      measured in pre- and post-celecoxib urine samples, (iii) celecoxib plasma and urine
      concentrations will be measured to evaluate the pharmacokinetic profile of the study drug.

      The investigators expect that lipid profile from ex vivo hWBA done on celecoxib-treated
      subjects will recapitulate findings from the in vitro hWBA received with celecoxib-treated
      human blood.
    

Intervention name(s): Celecoxib; Placebo
Intervention type(s): Drug; Drug
Intervention description(s): Blood and urine collections before and 3 hours after celecoxib administration.; Blood and urine collections before and 3 hours after placebo administration.
Other intervention name(s): Celebrex
Intervention MeSH terms: Celecoxib
Allocation: Randomized
Intervention model: Parallel Assignment
Masking: Quadruple (Participant, Care Provider, Investigator, Outcomes Assessor)
Primary purpose: Screening
Keywords: lipidomics; ex vivo human whole blood assay; celecoxib; selective COX-2 inhibitor