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Cerebrospinal fluid (CSF) is commonly used for assessing biomarkers of drug efficacy or disease progression in the central nervous system. Studies of CSF from pre-clinical species can characterize biomarkers for use in clinical trials. However, obtaining CSF from pre-clinical species, particularly rodents, can be challenging due to small body sizes, and consequently, low volumes of CSF. Surgical cannulation of rats is commonly used to allow for CSF withdrawal from the cisterna magna. However, cannulae do not remain patent over multiple days, making chronic studies on the same rats difficult. Moreover, CSF biomarkers may be affected by cannulation. Thus cannulation may contribute confounding factors to the understanding of CSF biomarkers. To determine the potential impact on biomarkers, CSF was analyzed from cannulated rats, surgically implanted with catheters as well as from non-cannulated rats. Brain protein biomarkers (αII-spectrin SBDP150 and total tau) and albumin, were measured in the CSF using ELISA assays. Overall, cannulated rat CSF had elevated levels of the biomarkers examined compared to non-cannulated rat CSF. Additionally, the variation in biomarker levels observed among CSF from cannulated rats was greater than that observed for non-cannulated rat CSF. These results demonstrate that in some cases, biomarker assessment using CSF from cannulated rats may differ from that of non-cannulated animals and may contribute confounding factors to biomarker measurements and assay development for clinical use.

The histamine H(4) receptor (H(4)R) is expressed primarily on cells involved in inflammation and immune responses. To determine the potential role of H(4)R in pain transmission, the effects of JNJ7777120, a potent and selective H(4) antagonist, were characterized in preclinical pain models. Administration of JNJ7777120 fully blocked neutrophil influx observed in a mouse zymosan-induced peritonitis model (ED(50)=17 mg/kg s.c., 95% CI=8.5-26) in a mast cell-dependent manner. JNJ7777120 potently reversed thermal hyperalgesia observed following intraplantar carrageenan injection of acute inflammatory pain (ED(50)=22 mg/kg i.p., 95% CI=10-35) in rats and significantly decreased the myeloperoxide activity in the carrageenan-injected paw. In contrast, no effects were produced by either H(1)R antagonist diphenhydramine, H(2)R antagonists ranitidine, or H(3)R antagonist ABT-239. JNJ7777120 also exhibited robust anti-nociceptive activity in persistent inflammatory (CFA) pain with an ED(50) of 29 mg/kg i.p. (95% CI=19-40) and effectively reversed monoiodoacetate (MIA)-induced osteoarthritic joint pain. This compound also produced dose-dependent anti-allodynic effects in the spinal nerve ligation (ED(50)=60 mg/kg) and sciatic nerve constriction injury (ED(50)=88 mg/kg) models of chronic neuropathic pain, as well as in a skin-incision model of acute post-operative pain (ED(50)=68 mg/kg). In addition, the analgesic effects of JNJ7777120 were maintained following repeated administration and were evident at the doses that did not cause neurologic deficits in rotarod test. Our results demonstrate that selective blockade of H(4) receptors in vivo produces significant anti-nociception in animal models of inflammatory and neuropathic pain.

Three novel heterocyclic benzofurans A-688057 ( 1 ), A-687136 ( 2 ), and A-698418 ( 3 ) were profiled for their in vitro and in vivo properties as a new series of histamine H 3 receptor antagonists. The compounds were all found to have nanomolar potency in vitro at histamine H 3 receptors, and when profiled in vivo for CNS activity, all were found active in an animal behavioral model of attention. The compound with the most benign profile versus CNS side effects was selected for greater scrutiny of its in vitro properties and overall drug-likeness. This compound, A-688057, in addition to its potent and robust efficacy in two rodent behavioral models at blood levels ranging 0.2–19 nM, possessed other favorable features, including high selectivity for H 3 receptors (H 3 , K i = 1.5 nM) versus off-target receptors and channels (including the hERG K + channel, K i > 9000 nM), low molecular weight (295), high solubility, moderate lipophilicity (logD pH7.4 = 2.05), and good CNS penetration (blood/brain 3.4×). In vitro toxicological tests indicated low potential for phospholipidosis, genotoxicity, and CYP 450 inhibition. Even though pharmacokinetic testing uncovered only moderate to poor oral bioavailability in rat (26%), dog (30%), and monkey (8%), and only moderate blood half-lives after i.v. administration ( t 1/2 in rat of 2.9 h, 1.7 h in dog, 1.8 h in monkey), suggesting poor human pharmacokinetics, the data overall indicated that A-688057 has an excellent profile for use as a pharmacological tool compound.

Further SAR studies on novel histamine H(3) receptor antagonists are presented. Compound 14bb is a potent antagonist of both the rat cortical and human clone receptors, and is demonstrated to act functionally as an antagonist in an in vivo mouse dipsogenia model.

New and existing methodologies were used to prepare a series of modified CCK analogs in which each amide bond was replaced by a trans-alkene unit. The data indicate that every amide linkage at C-terminal tetrapeptide (CCK-4) region is crucial for biological activity. While the amide bond beyond the Trp residue in the N-terminal direction can be replaced by a trans-alkene and still retain most of the binding potency and functional activity.

The backbone amide bonds in two series of tetrapeptide-based CCK-A receptor agonists were systematically replaced with the methylene amino isostere. Potent and selective pseudopeptides were identified that will facilitate our understanding of how these peptides interact with the CCK receptor and their structural correlations with other CCK ligands. This information will aid in the eventual development of peptidomimetics.

The interaction of the novel CCK analogs JMV-180, JMV-320, and JMV-332 with CCK-B/gastrin receptors on small cell lung cancer (SCLC) cells was investigated. JMV-180, JMV-320, and JMV-332 potently inhibited specific binding of 125I-CCK-8 to CCK-B/gastrin receptors expressed on the SCLC cell line NCI-H345 (H345) with IC50 values of 4.9, 1.8, and 7.0 nM, respectively. JMV-320 and JMV-332 stimulated intracellular calcium ([Ca2+]i) release in a dose-dependent manner in cells preloaded with indo-1. JMV-180 did not stimulate [Ca2+]i but inhibited the [Ca2+]i release elicited by 10 nM CCK-8 in a dose-dependent manner. These data indicate that JMV-320 and JMV-332 function as CCK-B/gastrin receptor agonists while JMV-180 functions as a CCK-B/gastrin receptor antagonist in H345 cells.