In a recently published study, Dr PNASThe researchers investigated the selective activity of sigmoid 2 receptor (σ2R) or transmembrane protein 97 (TMEM97) ligands, their effects on nociceptive neurons and the mechanism of action 24 hours later in murine neuropathic pain models.
Neuropathic pain, a chronic condition, presents a significant therapeutic challenge due to the limited efficacy and significant side effects of current medications. A novel drug, TMEM97, with roles in calcium signaling, cholesterol transport and homeostasis, was found to be a potential remedy. TMEM97 is associated with neurodegenerative disorders such as Alzheimer’s and Parkinson’s disease.
About the study
In the current study, researchers investigated whether the anti-neuropathic pain benefits of σ2R/TMEM97 ligands were attributable to binding to σ2R/TMEM97, which exhibits antinociception activity in animal models. They investigated whether targeting TMEM97 could relieve pain by interfering with the integrated stress response (ISR) associated with neuropathic pain produced by traumatic nerve damage, metabolic disease, and autoimmune conditions.
The team used a murine knockout (KO) model of the TMEM97 gene and the TMEM97-binding ligand, FEM-1689, to explore potential causal associations of σ2R/TMEM97 with antinociception in murine neuropathic pain models. They used RNAscope in situ hybridization of human dorsal root ganglia (DRG) from organ donors to investigate σ2R/TMEM97 expression in nociceptors.
Researchers developed FEM-1689 based on favorable results in neuropathic pain models treated with UKH-1114. Whether exploring σ2R/TMEM97 ligands using a global TMEM97-KO mouse acting on σ2R/TMEM97 reduces pain hypersensitivity. They treated male and female wild-type and TMEM97KO mice with intravenous injections of FEM-1689 at 10 nM, 100 nM, and 1,000.0 nM, corresponding to target binding, for 16 h.
The team investigated whether FEM-1689 would block ISR in DRG neurons. They cultured mouse dorsal root ganglia neurons from wild-type and TMEM97KO animals, administered FEM-1689 for 16 h, and assessed changes in phosphorylated eukaryotic initiation factor 2 (p-eIF2α) expression using immunocytochemistry (ICC). They explored the temporal dynamics of the FEM-1689 molecule by reducing p-eIF2α expression. They administered 100 nM FEM-1689 to wild-type murine DRG neurons over 0.5 hour, one hour, three hours, six hours, 12 hours, and 16 hours and measured p-eIF2α immunoreactivity.
The team investigated whether additional drugs binding to TMEM97 reduced the integrative stress response and whether ISR inhibition was specific to TMEM97 regulators that enhance antinociception. Computational docking analysis showed distinct interactions of FEM-1689 and SAS-0132 modulators with the extended binding sites of TMEM97. They investigated whether the effect of compound FEM-1689 to reduce P-eIF2α levels could reduce ISR-dependent and methylglyoxal (MGO)-induced mechanical hypersensitivity. To apply their findings from the murine DRG study to individuals, they treated human dorsal root ganglia neuronal cells from organ donor individuals with a TMEM97-binding ligand for 16 hours.
FEM-1689 requires the presence of the Tmem97 gene to produce antinociception in a rat nerve damage model. FEM-1689 suppresses ISR and promotes neurite outgrowth through a σ2R/TMEM97-specific activity, reduces ISR and p-eIF2α expression in human sensory neurons, and relieves pathogenic ISR engagement by MGO. Human SCN10A-positive nociceptor and satellite glial cells express σ2R/TMEM97.
Murine DRG neurons and satellite glial cells produce Tmem97 messenger ribonucleic acid (mRNA) in a manner comparable to human DRG. The antinociceptive effect of FEM-1689 in the SNI model was absent in TMEM97KO mice. The results support targeting σ2R/TMEM97 for the treatment of neuropathic pain. FEM-1689 was more specific for σ2R/TMEM97 than 40 CNS proteins excluding σ1R and the norepinephrine transporter (NET).
After nerve damage, male and female TMEM97KO and wild-type animals acquire significant and sustained mechanical hypersensitivity. Increasing doses of the TMEM97-binding ligand, FEM-1689, significantly modulated mechanical hypersensitivity in male and female wild-type mice for four days of a single treatment. However, FEM-1689 failed to reduce mechanical hypersensitivity in TMEM97KO mice, indicating that its effects were σ2R/TMEM97-dependent.
FEM-1689, a drug that suppresses the integrative stress response in murine DRG neurons, did not alter p-ACC levels or AMPK-p-ACC pathway activity in cultured mouse DRG neurons after 16 h, indicating a distinct antinociceptive route. TMEM97-KO neurons exhibited lower baseline p-eIF2α levels than wild-type neurons. ISRIB, an ISR inhibitor, reduced p-eIF2α levels in both variants. FEM-1689 effectively modulated MGO-induced and ISR-dependent pain hypersensitivity in human sensory neurons, highlighting the potential of developing TMEM97 modulators for diabetic neuropathic pain.
The results of the study showed that FEM-1689, a σ2R/TMEM97 ligand, can reduce neuropathic pain and block the ISR in mice. This effect requires direct regulation of σ2R/TMEM97, not σ1R or other proteins or receptors. FEM-1689 inhibits eIF2α phosphorylation in human dorsal root ganglia, reducing MGO-induced integrative stress responses. Targeting TMEM97 may reduce mechanical hypersensitivity in pain sufferers by reducing ISR.