Department of Anatomy, Physiology and Pharmacology, College of Veterinary Medicine, Auburn University , Auburn, AL , USA.
Front Endocrinol (Lausanne). 2014 Aug 4;5:131. doi: 10.3389/fendo.2014.00131. eCollection 2014.
Inactivating mutations of the melanocortin-4 receptor (MC4R) cause early-onset severe obesity in humans. Comprehensive functional studies show that most of the inactivating mutants of the MC4R are retained intracellularly. In the present study, we investigated whether a small molecule inverse agonist of the MC4R, Ipsen 5i, could act as a pharmacoperone and correct the cell surface expression and function of intracellularly retained mutant MC4Rs using multiple cell lines, including HEK293 and two neuronal cell lines. We showed that Ipsen 5i rescued the cell surface expression of all 11 intracellularly retained mutant MC4Rs studied herein in at least one cell line. Ipsen 5i functionally rescued seven mutants in all cell lines used. One mutant (Y157S) was functionally rescued in HEK293 cells but not in the two neuronal cell lines. Ipsen 5i increased cell surface expression of three mutants (S58C, G98R, and F261S) but did not affect signaling. Ipsen 5i had no effect on mutant MC4Rs with other defects (Δ88-92, D90N, I102S) or no defect (N274S). It also did not affect trafficking of a misrouted MC3R mutant (I335S). Cell impermeable peptide ligands of the MC4R or cell permeable small molecule ligand of δ opioid receptor could not rescue misrouted mutant MC4R. In summary, we demonstrated that Ipsen 5i was a novel potent pharmacoperone of the MC4R, correcting trafficking and signaling of a significant portion (73%) of intracellularly retained mutants. Additional studies are needed to demonstrate its in vivo efficacy.
黑素皮质素 4 受体 (MC4R) 的失活突变导致人类出现早发性重度肥胖。全面的功能研究表明，MC4R 的大多数失活突变体都被保留在细胞内。在本研究中，我们使用多种细胞系，包括 HEK293 和两个神经元细胞系，研究了 MC4R 的小分子反向激动剂 Ipsen 5i 是否可以作为一种药理学伴侣，纠正细胞内保留的突变型 MC4R 的细胞表面表达和功能。我们表明，Ipsen 5i 至少在一种细胞系中挽救了本文研究的所有 11 种细胞内保留的突变型 MC4R 的细胞表面表达。Ipsen 5i 在所有使用的细胞系中功能性地挽救了七种突变体。一个突变体（Y157S）在 HEK293 细胞中具有功能性，但在两个神经元细胞系中没有。Ipsen 5i 增加了三个突变体（S58C、G98R 和 F261S）的细胞表面表达，但不影响信号转导。Ipsen 5i 对具有其他缺陷（Δ88-92、D90N、I102S）或无缺陷（N274S）的突变型 MC4R 没有影响。它也不影响错误路由的 MC3R 突变体（I335S）的转运。MC4R 的细胞不可渗透肽配体或 δ 阿片受体的细胞可渗透小分子配体不能挽救错误路由的突变型 MC4R。总之，我们证明 Ipsen 5i 是 MC4R 的一种新型有效药理学伴侣，纠正了细胞内保留的突变体中相当一部分（73%）的转运和信号转导。需要进一步的研究来证明其体内疗效。
