Target-Mediated Brain Tissue Binding for Small Molecule Inhibitors of Heat Shock Protein 90.

Drug distribution in the brain is generally associated with an affinity for fatty brain tissues and therefore known to be species- and concentration-independent. We report here the effect of target affinity on brain tissue binding for 10 small molecules designed to inhibit brain heat shock protein 90 (HSP90), a widespread protein whose expression is 1-2% of total cytosolic proteins in eucaryotes. Our results show that increasing the test item concentrations from 0.3 to 100 µM increased the unbound fraction 32-fold for the most potent molecules, with no change for the inactive one (1.1 fold change). Saturation of HSP90 led to normal concentration-independent brain tissue binding. In vivo pharmacokinetics performed in rats showed that the overall volume of distribution of compounds is correlated with their affinity for HSP90. The in vitro binding and in vivo pharmacokinetics (PK) performed in rats showed that small molecule HSP90 inhibitors followed the principle of target-mediated drug disposition. We demonstrate that assessing unbound fractions in brain homogenate was subject to HSP90 target interference; this may challenge the process of linking systemic-free drug concentrations to central nervous system unbound concentrations necessary to establish the proper pharmacokinetics/pharmacodynamics (PK/PD) relation needed for human dose prediction.

Synthesis and SAR study of a novel series of dopamine receptor agonists.

The synthesis of a novel series of dopamine receptor agonists are described as well as their in vitro potency and efficacy on dopamine D1 and D2 receptors. This series was designed from pergolide and (4aR,10aR)-1-propyl-1,2,3,4,4a,5,10,10a-octahydro-benzo[g]quinolin-6-ol (PHBQ) and resulted in the synthesis of (2R,4aR,10aR)-2-methylsulfanylmethyl-4-propyl-3,4,4a,5,10,10a-hexahydro-2H-naphtho[2,3-b][1,4]oxazin-9-ol (compound 27), which has a D1 and D2 receptor profile similar to that of the most recently approved drug for Parkinson's disease, rotigotine.

Radiolabelling and PET brain imaging of the α1-adrenoceptor antagonist Lu AE43936.

Cerebral α1-adrenoceptors are a common target for many antipsychotic drugs. Thus, access to positron emission tomography (PET) brain imaging of α1-adrenoceptors could make important contributions to the understanding of psychotic disorders as well as to the pharmacokinetics and occupancy of drugs targeting the α1-adrenoceptors. However, so far no suitable PET radioligand has been developed for brain imaging of α1-adrenoceptors. Here, we report the synthesis of both enantiomers of the desmethyl precursors of the high affinity α1-adrenoceptor ligand (1). The two enantiomers of 1 were subsequently [¹¹C] radiolabelled and evaluated for brain uptake and binding by PET imaging in Danish Landrace pigs. (S)-[¹¹C]-1 and (R)-[¹¹C]-1 showed very limited brain uptake. Pre-treatment with cyclosporine A (CsA) resulted in a large increase in brain uptake, indicating that (R)-[¹¹C]-1 is a substrate for active efflux-transporters. This was confirmed in Madin Darby canine kidney (MDCK) cells overexpressing permeability glycoprotein (Pgp). In conclusion, the limited brain uptake of both (S)-[¹¹C]-1 and (R)-[¹¹C]-1 in the pig brain necessitates the search for alternative radioligands for in vivo PET brain imaging of α1-adrenoceptors.

Identification of the cytochrome P450 and other enzymes involved in the in vitro oxidative metabolism of a novel antidepressant, Lu AA21004.

1-[2-(2,4-Dimethyl-phenylsulfanyl)-phenyl]-piperazine (Lu AA21004) is a novel antidepressant that is currently in late-stage clinical development for major depressive disorder. In the present study, the metabolism of Lu AA21004 was investigated using human liver microsomes (HLM), human liver S9 fraction, and recombinant enzymes. Lu AA21004 was found in vitro to be oxidized to a 4-hydroxy-phenyl metabolite, a sulfoxide, an N-hydroxylated piperazine, and a benzylic alcohol, which was further oxidized to the corresponding benzoic acid [3-methyl-4-(2-piperazin-1-yl-phenysulfanyl)-benzoic acid (Lu AA34443)]. The formation of the 4-hydroxy-phenyl metabolite was catalyzed by CYP2D6 with some contribution from CYP2C9, whereas the formation of the sulfoxide was mediated by CYP3A4/5 and CYP2A6. CYP2C9 and CYP2C19 were the primary enzymes responsible for formation of the N-hydroxylated metabolite. The benzylic alcohol was formed by CYP2D6 only. The oxidation of the benzylic alcohol to the corresponding benzoic acid of Lu AA21004 was catalyzed by alcohol dehydrogenase and aldehyde dehydrogenase, with some contribution from aldehyde oxidase. CYP2D6 was also capable of catalyzing the formation of the benzoic acid of Lu AA21004; however, its overall contribution to this pathway was negligible. Enzyme kinetic parameters revealed that the rate-limiting step in the formation of the benzoic acid from Lu AA21004 is the formation of the corresponding alcohol. Thus, the intrinsic clearance (V(max)/K(m)) in HLM for metabolism of Lu AA21004 to the benzylic alcohol was 1.13 × 10(-6) l · min(-1) · mg(-1), whereas the subsequent metabolism of the benzylic alcohol to the benzoic acid of Lu AA21004 is characterized by an intrinsic clearance (V(max)/K(m)) in S9 fraction of 922 × 10(-6) l · min(-1) · mg(-1).

Exploring the neuroleptic substituent in octoclothepin: potential ligands for positron emission tomography with subnanomolar affinity for α(1)-adrenoceptors.

A series of 1-(10,11-dihydrodibenzo[b,f]thiepin-10-yl)-4-methylpiperazine analogues substituted in the 8-position of the 10,11-dihydrodibenzo[b,f]thiepine scaffold with aryl, heteroaryl, amine, and amide substituents are described. The compounds were designed using the previously reported Liljefors-Bøgesø pharmacophore model for dopamine D(2) and α(1)-adrenoceptor antagonists, with the aim of obtaining selective α(1)-adrenoceptor antagonists suitable for development as radioligands for imaging of central α(1)-adrenoceptors by positron emission tomography. Sixteen aryl and heteroaryl substituted octoclothepin analogues were prepared by a convergent synthesis via coupling of 1-methyl-4-(8-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)-10,11-dihydrodibenzo[b,f]thiepin-10-yl)piperazine with aryl and heteroaryl halides under palladium catalysis. The most selective compound obtained, (S)-N-((11-(4-methylpiperazin-1-yl)-10,11-dihydrodibenzo[b,f]thiepin-2-yl)methyl)isobutyramide (S)-35, showed a similar subnanomolar affinity compared to α(1a), α(1b), and α(1d)-adrenoceptors and a selectivity ratio of 20, 440, and 20 with respect to D(2), 5-HT(2C), and H(1) receptors, respectively.

N,N'-Bis-(2-(cyano)ethoxycarbonyl)-2-methyl-2-thiopseudourea: a guanylating reagent for synthesis of 2'-O-[2-(Guanidinium)ethyl]-modified oligonucleotides.

A guanylating reagent, N,N'-bis-(2-(cyano)ethoxycarbonyl)-2-thiopseudourea, was synthesized and used for synthesis of 2'-O-[2-(guanidinium)ethyl] (2'-O-GE) modified oligonucleotides. A convenient deprotection method for the 2'-O-GE oligonucleotides was developed.

2'-O-[2-(guanidinium)ethyl]-modified oligonucleotides: stabilizing effect on duplex and triplex structures.

[structure: see text] Oligonucleotides with a novel 2'-O-[2-(guanidinium)ethyl] (2'-O-GE) modification have been synthesized using a novel protecting group strategy for the guanidinium group. This modification enhances the binding affinity of oligonucleotides to RNA as well as duplex DNA (DeltaT(m) 3.2 degrees C per modification). The 2'-O-GE modified oligonucleotides exhibited exceptional resistance to nuclease degradation. The crystal structure of a palindromic duplex formed by a DNA oligonucleotide with a single 2'-O-GE modification was solved at 1.16 A resolution.