James J. Sahn

Identification of the gene that codes for the σ2 receptor

Significance Of the many receptors that were pharmacologically described during the 20th century, almost all were cloned by the end of the 1990s. A key exception is the σ2 receptor, a potential therapeutic target for diseases as diverse as schizophrenia, Alzheimer’s disease, and cancer. Despite the development of a rich pharmacopeia, the unknown molecular identity of the receptor has crippled biological investigation. Here, we identify the σ2 receptor as TMEM97, a membrane protein implicated in cancer and a binding partner of Niemann–Pick disease protein NPC1. Our results unite two fields of research, bringing the σ2 receptor into the modern age of biological inquiry and providing the TMEM97 field with a rich pool of ligands and pharmacological tools. The σ2 receptor is an enigmatic protein that has attracted significant attention because of its involvement in diseases as diverse as cancer and neurological disorders. Unlike virtually all other receptors of medical interest, it has eluded molecular cloning since its discovery, and the gene that codes for the receptor remains unknown, precluding the use of modern biological methods to study its function. Using a chemical biology approach, we purified the σ2 receptor from tissue, revealing its identity as TMEM97, an endoplasmic reticulum-resident transmembrane protein that regulates the sterol transporter NPC1. We show that TMEM97 possesses the full suite of molecular properties that define the σ2 receptor, and we identify Asp29 and Asp56 as essential for ligand recognition. Cloning the σ2 receptor resolves a longstanding mystery and will enable therapeutic targeting of this potential drug target.

Facile and unified approach to skeletally diverse, privileged scaffolds.

A novel strategy has been developed to generate a diverse array of privileged scaffolds from readily available tetrahydropyridine precursors that may be prepared by a multicomponent assembly process followed by a ring-closing metathesis. The functionality embedded in these key intermediates enables their facile elaboration into more complex structures of biological relevance by a variety of ring-forming processes and refunctionalizations.

Expedient Synthesis of Norbenzomorphan Library via Multicomponent Assembly Process Coupled with Ring-Closing Reactions

A 124-member norbenzomorphan library has been prepared utilizing a novel multicomponent assembly process (MCAP) followed by a variety of ring-closing reactions to generate norbenzomorphan scaffolds that were readily derivatized via a series of aryl halide cross-coupling and nitrogen functionalization reactions. Biological screening has revealed some novel activities that have not been previously associated with this class of compounds.

Small molecule modulator of sigma 2 receptor is neuroprotective and reduces cognitive deficits and neuroinflammation in experimental models of Alzheimer's disease

Accumulating evidence suggests that modulating the sigma 2 receptor (Sig2R) can provide beneficial effects for neurodegenerative diseases. Herein, we report the identification of a novel class of Sig2R ligands and their cellular and in vivo activity in experimental models of Alzheimers disease (AD). We report that SAS‐0132 and DKR‐1051, selective ligands of Sig2R, modulate intracellular Ca2+ levels in human SK‐N‐SH neuroblastoma cells. The Sig2R ligands SAS‐0132 and JVW‐1009 are neuroprotective in a C. elegans model of amyloid precursor protein‐mediated neurodegeneration. Since this neuroprotective effect is replicated by genetic knockdown and knockout of vem‐1, the ortholog of progesterone receptor membrane component‐1 (PGRMC1), these results suggest that Sig2R ligands modulate a PGRMC1‐related pathway. Last, we demonstrate that SAS‐0132 improves cognitive performance both in the Thy‐1 hAPPLond/Swe+ transgenic mouse model of AD and in healthy wild‐type mice. These results demonstrate that Sig2R is a promising therapeutic target for neurocognitive disorders including AD.

[2 + 2] Photochemical cycloaddition/ring opening of 6-alkenyl-2,3-dihydro-4-pyridones.

During the course of a study aimed at constructing azaspirocycles from 2,3-dihydro-4-pyridones, an unexpected product was obtained in the SET ring-opening reaction of photocycloadduct 1. Differences in reactivity between homologues 1 and 2 were observed in the presence of SmI(2). Tricyclic ketone 2 afforded azaspiro[5.5]undecane 15 when treated with SmI(2); however, when ketone 1 was submitted to similar reaction conditions a double ring-opening/reduction sequence gave cis-piperidinol 10.

Sigma 2 Receptor/Tmem97 Agonists Produce Long Lasting Antineuropathic Pain Effects in Mice

Neuropathic pain is an important medical problem with few effective treatments. The sigma 1 receptor (σ1R) is known to be a potential target for neuropathic pain therapeutics, and antagonists for this receptor are effective in preclinical models and are currently in phase II clinical trials. Conversely, relatively little is known about σ2R, which has recently been identified as transmembrane protein 97 (Tmem97). We generated a series of σ1R and σ2R/Tmem97 agonists and antagonists and tested them for efficacy in the mouse spared nerve injury (SNI) model. In agreement with previous reports, we find that σ1R ligands given intrathecally (IT) produce relief of SNI-induced mechanical hypersensitivity. We also find that the putative σ2R/Tmem97 agonists DKR-1005, DKR-1051, and UKH-1114 (Ki ∼ 46 nM) lead to relief of SNI-induced mechanical hypersensitivity, peaking at 48 h after dosing when given IT. This effect is blocked by the putative σ2R/Tmem97 antagonist SAS-0132. Systemic administration of UKH-1114 (10 mg/kg) relieves SNI-induced mechanical hypersensitivity for 48 h with a peak magnitude of effect equivalent to 100 mg/kg gabapentin and without producing any motor impairment. Finally, we find that the TMEM97 gene is expressed in mouse and human dorsal root ganglion (DRG) including populations of neurons that are involved in pain; however, the gene is also likely expressed in non-neuronal cells that may contribute to the observed behavioral effects. Our results show robust antineuropathic pain effects of σ1R and σ2R/Tmem97 ligands, demonstrate that σ2R/Tmem97 is a novel neuropathic pain target, and identify UKH-1114 as a lead molecule for further development.

Norbenzomorphan Framework as a Novel Scaffold for Generating Sigma 2 Receptor/PGRMC1 Subtype-Selective Ligands

A novel structural class with high affinity and subtype selectivity for the sigma 2 receptor has been discovered. Preliminary structure–affinity relationship data are presented showing that 8‐substituted 1,3,4,5‐tetrahydro‐1,5‐methanobenzazepine (norbenzomorphan) derivatives elicit modest to high selectivity for the sigma 2 over the sigma 1 receptor subtype. Indeed, piperazine analogue 8‐(4‐(3‐ethoxy‐3‐oxopropyl)piperazin‐1‐yl)‐1,3,4,5‐tetrahydro‐1,5‐methanobenzazepine‐2‐carboxylate (SAS‐1121) is 574‐fold selective for the sigma 2 over the sigma 1 receptor, thereby establishing it as one of the more subtype‐selective sigma 2 binding ligands reported to date. Emerging evidence has implicated the sigma 2 receptor in multiple health disorders, so the drug‐like characteristics of many of the selective sigma 2 receptor ligands disclosed herein, coupled with their structural similarity to frameworks found in known drugs, suggest that norbenzomorphan analogues may be promising candidates for further development into drug leads.

APPLICATIONS OF MULTICOMPONENT ASSEMBLY PROCESSES TO THE FACILE SYNTHESES OF DIVERSELY FUNCTIONALIZED NITROGEN HETEROCYCLES.

Several multicomponent assembly processes have been developed for the synthesis of intermediates that may be elaborated by a variety of cyclizations to generate a diverse array of highly functionalized heterocycles from readily-available starting materials. The overall approach enables the efficient preparation of libraries of small molecules derived from fused, privileged scaffolds.

Norbenzomorphan Scaffold: Chemical Tool for Modulating Sigma Receptor-Subtype Selectivity

Some norbenzomorphans exhibit high affinity for sigma 1 and sigma 2 receptors, and varying the position of substituents on the aromatic ring of this scaffold has a significant effect on subtype selectivity. In particular, compounds bearing several different substituents at C7 of the norbenzomorphan ring system exhibit a general preference for the sigma 1 receptor, whereas the corresponding C8-substituted analogues preferentially bind at the sigma 2 receptor. These findings suggest that the norbenzomorphan scaffold may be a unique chemical template that can be easily tuned to prepare small molecules for use as tool compounds to study the specific biological effects arising from preferential binding at either sigma receptor subtype. In the absence of structural characterization data for the sigma 2 receptor, such compounds will be useful toward refining the pharmacophore model of its binding site.

Investigating isoindoline, tetrahydroisoquinoline, and tetrahydrobenzazepine scaffolds for their sigma receptor binding properties

Substituted norbenzomorphans are known to display high affinity and selectivity for the two sigma receptor (σR) subtypes. In order to study the effects of simplifying the structures of these compounds, a scaffold hopping strategy was used to design several novel sets of substituted isoindolines, tetrahydroisoquinolines and tetrahydro-2-benzazepines. The binding affinities of these new compounds for the sigma 1 (σ1R) and sigma 2 (σ2R) receptors were determined, and some analogs were identified that exhibit high affinity (Ki ≤ 25 nM) and significant selectivity (>10-fold) for σ1R or σ2R. The preferred binding modes of selected compounds for the σ1R are predicted by modeling studies, and the nature of substituents on the aromatic ring and the nitrogen atom of the bicyclic skeleton appears to affect the preferred binding orientation of σ1R-preferring ligands.