Robert W. Wiethe

Fmoc mediated synthesis of Peptide Nucleic Acids

Abstract The syntheses of the Fmoc-protected Peptide Nucleic Acid (PNA) monomer pentafluorophenyl esters of adenine (26), cytosine (23), guanine (29) and thymine (20), and their oligomerization are described. The Fmoc PNA backbone 1 is prepared as a stable hydrochloride salt. The base acetic acids of adenine (4) and cytosine (3) were prepared by Cbz protection of the exocyclic amino groups followed by alkylation with t-butylbromoacetate and subsequent acid hydrolysis of the t-butyl ester. Allylation of 6-chloro-2-aminopurine followed by acid hydrolysis, Cbz protection with N-(benzyloxycarbonyl)imidazole, ozonolytic cleavage, and oxidation afforded the Cbz-protected guanine acetic acid (5). The base acetic acids (2, 3, 4 and 5) were coupled to the backbone (1) with either EDC (2 and 3) or BOP reagent (4 and 5). Acid hydrolysis of the resulting t-butyl esters and transesterification afforded the corresponding pentafluorophenyl esters (20, 23, 26 and 29). Oligomerization is conducted on a 0.05 mmol scale with a mere 2 fold excess of monomer in each coupling cycle. The N-terminal Fmoc group is retained on the final oligomer, following HF cleavage and deprotection, providing a convenient lipophilic handle for HPLC purification.

Identification and Characterization of 4-Chloro-N-(2-{[5-trifluoromethyl)-2-pyridyl]sulfonyl}ethyl)benzamide (GSK3787), a Selective and Irreversible Peroxisome Proliferator-Activated Receptor δ (PPARδ) Antagonist

4-Chloro-N-(2-{[5-trifluoromethyl)-2-pyridyl]sulfonyl}ethyl)benzamide 3 (GSK3787) was identified as a potent and selective ligand for PPARdelta with good pharmacokinetic properties. A detailed binding study using mass spectral analysis confirmed covalent binding to Cys249 within the PPARdelta binding pocket. Gene expression studies showed that pyridylsulfone 3 antagonized the transcriptional activity of PPARdelta and inhibited basal CPT1a gene transcription. Compound 3 is a PPARdelta antagonist with utility as a tool to elucidate PPARdelta cell biology and pharmacology.

N-hydroxyformamide peptidomimetics as TACE/matrix metalloprotease inhibitors: oral activity via P1' isobutyl substitution.

N-Hydroxyformamide-class metalloprotease inhibitors were designed and synthesized, which have potent broad-spectrum activity versus matrix metalloproteases and TNF-alpha converting enzyme (TACE). Compound 13c possesses good oral and intravenous pharmacokinetics in the rat and dog.

2-Amino-4,6-diarylpyridines as novel ligands for the estrogen receptor.

We have prepared a novel series of 2-amino-4,6-diarylpyridines that function as ligands of estrogen receptor alpha (ERalpha) and estrogen receptor beta (ERbeta). These compounds bind to both ERalpha and ERbeta with a modest selectivity for the alpha subtype. The most potent of these analogues, compound 19, has a K(i)=20nM at ERalpha. These molecules represent a novel template for designing potentially useful ligands for the estrogen receptor.

Identification of 1-({[1-(4-fluorophenyl)-5-(2-methoxyphenyl)-1H-pyrazol-3-yl]carbonyl}amino)cyclohexane carboxylic acid as a selective nonpeptide neurotensin receptor type 2 compound.

Compounds active at neurotensin receptors (NTS1 and NTS2) exert analgesic effects on different types of nociceptive modalities, including thermal, mechanical, and chemical stimuli. The NTS2 preferring peptide JMV-431 (2) and the NTS2 selective nonpeptide compound levocabastine (6) have been shown to be effective in relieving the pain associated with peripheral neuropathies. With the aim of identifying novel nonpeptide compounds selective for NTS2, we examined analogues of SR48692 (5a) using a FLIPR calcium assay in CHO cells stably expressing rat NTS2. This led to the discovery of the NTS2 selective nonpeptide compound 1-({[1-(4-fluorophenyl)-5-(2-methoxyphenyl)-1H-pyrazol-3-yl]carbonyl}amino)cyclohexane carboxylic acid (NTRC-739, 7b) starting from the nonselective compound 5a.

Identification of N-[(5-{[(4-methylphenyl)sulfonyl]amino}-3-(trifluoroacetyl)-1H-indol-1-yl)acetyl]-l-leucine (NTRC-824), a neurotensin-like nonpeptide compound selective for the neurotensin receptor type 2.

Compounds acting via the neurotensin receptor type 2 (NTS2) are known to be active in animal models of acute and chronic pain. To identify novel NTS2 selective analgesics, we searched for NTS2 selective nonpeptide compounds using a FLIPR assay and identified the title compound (NTRC-824, 5) that, to our knowledge, is the first nonpeptide that is selective for NTS2 versus NTS1 and behaves like the endogenous ligand neurotensin in the functional assay.

Pyrazole antagonists of the CB1 receptor with reduced brain penetration

Type 1 cannabinoid receptor (CB1) antagonists might be useful for treating obesity, liver disease, metabolic syndrome, and dyslipidemias. Unfortunately, inhibition of CB1 in the central nervous system (CNS) produces adverse effects, including depression, anxiety and suicidal ideation in some patients, which led to withdrawal of the pyrazole inverse agonist rimonabant (SR141716A) from European markets. Efforts are underway to produce peripherally selective CB1 antagonists to circumvent CNS-associated adverse effects. In this study, novel analogs of rimonabant (1) were explored in which the 1-aminopiperidine group was switched to a 4-aminopiperidine, attached at the 4-amino position (5). The piperidine nitrogen was functionalized with carbamates, amides, and sulfonamides, providing compounds that are potent inverse agonists of hCB1 with good selectivity for hCB1 over hCB2. Select compounds were further studied using in vitro models of brain penetration, oral absorption and metabolic stability. Several compounds were identified with predicted minimal brain penetration and good metabolic stability. In vivo pharmacokinetic testing revealed that inverse agonist 8c is orally bioavailable and has vastly reduced brain penetration compared to rimonabant.

Identification of N-{[6-chloro-4-(2,6-dimethoxyphenyl)quinazolin-2-yl]carbonyl}-l-leucine (NTRC-808), a novel nonpeptide chemotype selective for the neurotensin receptor type 2

Compounds acting via the GPCR neurotensin receptor type 2 (NTS2) display analgesic effects in relevant animal models. Using a pharmacophore model based on known NT receptor nonpeptide compounds, we screened commercial databases to identify compounds that might possess activity at NTS2 receptor sites. Modification of our screening hit to include structural features known to be recognized by NTS1 and NTS2, led to the identification of the novel NTS2 selective nonpeptide, N-{[6-chloro-4-(2,6-dimethoxyphenyl)quinazolin-2-yl]carbonyl}-l-leucine (9). This compound is a potent partial agonist in the FLIPR assay with a profile of activity similar to that of the reference NTS2 analgesic nonpeptide levocabastine (5).

The amide linker in nonpeptide neurotensin receptor ligands plays a key role in calcium signaling at the neurotensin receptor type 2.

Compounds acting via the GPCR neurotensin receptor type 2 (NTS2) display analgesia in relevant preclinical models. The amide bond in nonpeptide NTS1 antagonists plays a central role in receptor recognition and molecular conformation. Using NTS2 FLIPR and binding assays, we found that it is also a key molecular structure for binding and calcium mobilization at NTS2. We found that reversed amides display a shift from agonist to antagonist activity and provided examples of the first competitive nonpeptide antagonists observed in the NTS2 FLIPR assay. These compounds will be valuable tools for determining the role of calcium signaling in vitro to NTS2 mediated analgesia.

Identification of 2-({[1-(4-Fluorophenyl)-5-(2-methoxyphenyl)-1H-pyrazol-3-yl]carbonyl}amino)tricyclo[3.3.1.13,7]decane-2-carboxylic Acid (NTRC-844) as a Selective Antagonist for the Rat Neurotensin Receptor Type 2

Neurotensin receptor type 2 (NTS2) compounds display analgesic activity in animal pain models. We have identified the first high-affinity NTS2-selective antagonist (8) that is active in vivo. This study also revealed that the NTS2 FLIPR assay designation for a compound, agonist, partial agonist, and so forth, did not correlate with its in vivo activity as observed in the thermal tail-flick acute model of pain. This suggests that calcium mobilization is not the signaling pathway involved in NTS2-mediated analgesia as assessed by the thermal tail-flick model. Finally, we found a significant bias between rat and human for compound 9 in the NTS2 binding assay.