Limits...
Ipsen 5i is a Novel Potent Pharmacoperone for Intracellularly Retained Melanocortin-4 Receptor Mutants.

Tao YX, Huang H - Front Endocrinol (Lausanne) (2014)

Bottom Line: 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).

View Article: PubMed Central - PubMed

Affiliation: Department of Anatomy, Physiology and Pharmacology, College of Veterinary Medicine, Auburn University , Auburn, AL , USA.

ABSTRACT
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.

No MeSH data available.


Related in: MedlinePlus

Ipsen 5i rescued the cell surface expression of intracellularly retained mutant MC4Rs in Neuro2a cells. Neuro2a cells transiently expressing WT or mutant MC4Rs were treated with 10−6 M Ipsen 5i. Results are shown as mean ± SEM of at least three independent experiments. See the legend to Figure 3 for details.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC4120685&req=5

Figure 4: Ipsen 5i rescued the cell surface expression of intracellularly retained mutant MC4Rs in Neuro2a cells. Neuro2a cells transiently expressing WT or mutant MC4Rs were treated with 10−6 M Ipsen 5i. Results are shown as mean ± SEM of at least three independent experiments. See the legend to Figure 3 for details.

Mentions: Neuro2a cells transiently expressing WT or mutant MC4Rs were treated with 10−6 M Ipsen 5i for 24 h and then were used for flow cytometry studies. S58C was not further studied in neuronal cells because as described later its function was not rescued by Ipsen 5i. As shown in Figure 4, the cell surface expression of eight mutants (N62S, I69R, P78L, C84R, G98R, W174C, P260Q, and C271Y) was increased with Ipsen 5i treatment compared with the DMSO-treated control group. The cell surface expression of I69R was slightly (although significantly) increased in Neuro2a cells whereas it was not increased in HEK293 cells. The increase of cell surface expression of F261S was not statistically significant in Neuro2a cells. One mutant (Y157S) that was rescued by Ipsen 5i in HEK293 cells was not rescued in Neuro2a cells.


Ipsen 5i is a Novel Potent Pharmacoperone for Intracellularly Retained Melanocortin-4 Receptor Mutants.

Tao YX, Huang H - Front Endocrinol (Lausanne) (2014)

Ipsen 5i rescued the cell surface expression of intracellularly retained mutant MC4Rs in Neuro2a cells. Neuro2a cells transiently expressing WT or mutant MC4Rs were treated with 10−6 M Ipsen 5i. Results are shown as mean ± SEM of at least three independent experiments. See the legend to Figure 3 for details.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4120685&req=5

Figure 4: Ipsen 5i rescued the cell surface expression of intracellularly retained mutant MC4Rs in Neuro2a cells. Neuro2a cells transiently expressing WT or mutant MC4Rs were treated with 10−6 M Ipsen 5i. Results are shown as mean ± SEM of at least three independent experiments. See the legend to Figure 3 for details.
Mentions: Neuro2a cells transiently expressing WT or mutant MC4Rs were treated with 10−6 M Ipsen 5i for 24 h and then were used for flow cytometry studies. S58C was not further studied in neuronal cells because as described later its function was not rescued by Ipsen 5i. As shown in Figure 4, the cell surface expression of eight mutants (N62S, I69R, P78L, C84R, G98R, W174C, P260Q, and C271Y) was increased with Ipsen 5i treatment compared with the DMSO-treated control group. The cell surface expression of I69R was slightly (although significantly) increased in Neuro2a cells whereas it was not increased in HEK293 cells. The increase of cell surface expression of F261S was not statistically significant in Neuro2a cells. One mutant (Y157S) that was rescued by Ipsen 5i in HEK293 cells was not rescued in Neuro2a cells.

Bottom Line: 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).

View Article: PubMed Central - PubMed

Affiliation: Department of Anatomy, Physiology and Pharmacology, College of Veterinary Medicine, Auburn University , Auburn, AL , USA.

ABSTRACT
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.

No MeSH data available.


Related in: MedlinePlus