br Methods and materials br Cyclooxygenase COX
Methods and materials
Cyclooxygenase (COX) is the key enzyme required for the conversion of arachidonic Telbivudine synthesis to prostaglandin, prostacyclins and thromboxanes., , , Two distinct isoforms of COX were initially discovered: COX-1 is a constitutive, housekeeping enzyme that is expressed in many tissues and is responsible for the production of prostanoids associated with normal homeostatic functions. The other isoform, COX-2, is inducible in most tissues and is mainly stimulated by inflammatory responses., , , A third COX isoform, COX-3 has also been recently identified as a splice variant of COX-1, but it is not active in human. Traditional non-steroidal anti-inflammatory drugs (NSAIDs) are non-selective COX-2 inhibitors that can cause mild to severe side effects with doses equivalent or lower than clinical dose., , , Large doses of NSAIDs may lead to death via kidney and cardiac malfunctions and fluid retention. The side effects of NSAIDs are most likely due to the inhibition of COX-1, the enzyme required for the normal hemostatic functions of the body, or to the inhibition of COX-2 in the areas where it is constitutively expressed. In order to overcome these side effects, a second generation of selective COX-2 inhibitors (COXIBs) was developed., , Among the COXIBs, celebrex (celecoxib) is the only drug used for arthritis, pain, and other inflammatory processes and has been tested for the treatment of cancer, stroke and Alzheimer’s disease., , , However, the COX-2 selective NSAIDs vioxx (rofecoxib) and bextra (valdecoxib) were pulled from the market due to suspected cardiac toxicity., , , Although COX-2 is the drug target for various diseases, the pathogenesis of the role of COX-2 in many diseases and disorders is not fully understood. Therefore, the ability to quantitatively monitor COX-2 expression in vivo, non-invasively, would lead to a better understanding of the pathophysiology of diseases involving COX-2 and would allow drug development and monitoring of treatment effects targeting COX-2. Among various imaging modalities, PET is one of the most sensitive non-invasive, tomographic imaging techniques used in modern medicine to study various biochemical and biological processes in vivo. A specific PET tracer for COX-2 could serve as a biomarker for the early diagnosis of diseases in which upregulation of COX-2 occurs. In vivo imaging agents for COX-2 could also aid in the development of novel COXIB medications and in monitoring these medications effects in CNS, cardiac and renal functions despite the potential of COX-2 imaging in medicine, no successful in vivo imaging agent is currently available for COX-2 likely due to the lack of qualified ligands with high affinity to COX-2., , , Low COX-2 selectivity over COX-1, complex radiosynthesis procedures resulting in poor molar activity, and de-[F]fluorination may also contribute to the failure of some of the PET radiotracers reported for successful imaging of COX-2 in brain., , , Previous PET imaging experiments indicate that, because of the low baseline expression of COX-2, it is necessary to evaluate high-affinity radiotracers with excellent COX-2/COX-1 selectivity in order to accomplish detection of COX-2 by PET imaging., , Our continued effort in this direction has resulted in the synthesis of [C]3-(4-methylsulfonylphenyl)-4-phenyl-5-trifluoromethyl isoxazole ([C]TMI),, a highly selective COX-2 inhibitor with IC < 1 nM and >500,000 times higher selectivity for COX-2 over COX-1 (COX-1 IC ≥ 500 μM). Herein we describe the in vivo evaluation of [C]TMI in baboon brain using PET imaging. [C]TMI was obtained in 30 ± 5% yield at the end of synthesis (EOS) (based on [C]CHI, n = 10) with a molar activity of 75 ± 9.25 GBq/μmol and radiochemical purity > 99% (). TMI also did not show significant affinity to a variety of competitive brain receptors, transporters, biogenic amines and proteins (Ki > 10 μM) based on the National Institute of Mental Health –Psychoactive Drug Screening Program (NIMH-PDSP) binding assays ().