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Identification of an orally active small-molecule PTHR1 agonist for the treatment of hypoparathyroidism

View Article: PubMed Central - PubMed

ABSTRACT

Parathyroid hormone (PTH) is essential for calcium homeostasis and its action is mediated by the PTH type 1 receptor (PTHR1), a class B G-protein-coupled receptor. Hypoparathyroidism and osteoporosis can be treated with PTH injections; however, no orally effective PTH analogue is available. Here we show that PCO371 is a novel, orally active small molecule that acts as a full agonist of PTHR1. PCO371 does not affect the PTH type 2 receptor (PTHR2), and analysis using PTHR1–PTHR2 chimeric receptors indicated that Proline 415 of PTHR1 is critical for PCO371-mediated PTHR1 activation. Oral administration of PCO371 to osteopenic rats provokes a significant increase in bone turnover with limited increase in bone mass. In hypocalcemic rats, PCO371 restores serum calcium levels without increasing urinary calcium, and with stronger and longer-lasting effects than PTH injections. These results strongly suggest that PCO371 can provide a new treatment option for PTH-related disorders, including hypoparathyroidism.

No MeSH data available.


Related in: MedlinePlus

Effects of PCO371 on PTHR1 signalling in UMR-106 cells.(a) cAMP production by hPTH(1–34) and PCO371 in UMR-106 cells, which natively express rat PTHR1. (b) Duration of cAMP-signalling responses induced by PCO371, hPTH(1–34) and LA-PTH in UMR-106 cells. The time courses of cAMP responses produced by PCO371 (0.1 mmol l−1), hPTH(1–34) (1 μmol l−1 ) or LA-PTH (0.1 μmol l−1) in UMR-106 cells were examined by cAMP washout assay. The cAMP response is expressed as a percentage of the maximal cAMP produced in the cells treated with each ligand (determined by incubating cells concomitantly with the ligand for 10 min, and further incubating with IBMX for 5 min without a washing-out phase). The range of maximum cAMP values was PCO371: 17.4+0.2 pmol per well, hPTH(1–34): 21.3+0.9, LA-PTH: 42.2+0.9, and basal cAMP: 1.8+0.2. (c–g) Effects of PCO371 and hPTH(1–34) on the expression of mRNAs for c-fos (Fos) (c), osteocalcin (Bglap) (d), RANKL (Tnfsf11) (e), osteoprotegerin (Tnfrsf11b) (f) and sclerostin (Sost) (g). mRNA levels are shown as fold-change over control. Gene expression was analysed at 1 h (c) or 6 h (d–g) after treatment of UMR-106 cells with hPTH(1–34) or PCO371. All genes were normalized to 18 S ribosomal RNA. Data are represented as the mean±s.d. of one experiment (n=3 for a–g).
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f2: Effects of PCO371 on PTHR1 signalling in UMR-106 cells.(a) cAMP production by hPTH(1–34) and PCO371 in UMR-106 cells, which natively express rat PTHR1. (b) Duration of cAMP-signalling responses induced by PCO371, hPTH(1–34) and LA-PTH in UMR-106 cells. The time courses of cAMP responses produced by PCO371 (0.1 mmol l−1), hPTH(1–34) (1 μmol l−1 ) or LA-PTH (0.1 μmol l−1) in UMR-106 cells were examined by cAMP washout assay. The cAMP response is expressed as a percentage of the maximal cAMP produced in the cells treated with each ligand (determined by incubating cells concomitantly with the ligand for 10 min, and further incubating with IBMX for 5 min without a washing-out phase). The range of maximum cAMP values was PCO371: 17.4+0.2 pmol per well, hPTH(1–34): 21.3+0.9, LA-PTH: 42.2+0.9, and basal cAMP: 1.8+0.2. (c–g) Effects of PCO371 and hPTH(1–34) on the expression of mRNAs for c-fos (Fos) (c), osteocalcin (Bglap) (d), RANKL (Tnfsf11) (e), osteoprotegerin (Tnfrsf11b) (f) and sclerostin (Sost) (g). mRNA levels are shown as fold-change over control. Gene expression was analysed at 1 h (c) or 6 h (d–g) after treatment of UMR-106 cells with hPTH(1–34) or PCO371. All genes were normalized to 18 S ribosomal RNA. Data are represented as the mean±s.d. of one experiment (n=3 for a–g).

Mentions: We also examined whether PCO371 induces cAMP production via endogenously expressed PTHR1 by using UMR-106 cells (a rat osteosarcoma cell line) that express endogenous rat PTHR1. Although the potency was much weaker than that of hPTH(1–34), PCO371 induced cAMP production as did hPTH(1–34), suggesting that PCO371 acts as a full agonist of PTHR1 (Fig. 2a).


Identification of an orally active small-molecule PTHR1 agonist for the treatment of hypoparathyroidism
Effects of PCO371 on PTHR1 signalling in UMR-106 cells.(a) cAMP production by hPTH(1–34) and PCO371 in UMR-106 cells, which natively express rat PTHR1. (b) Duration of cAMP-signalling responses induced by PCO371, hPTH(1–34) and LA-PTH in UMR-106 cells. The time courses of cAMP responses produced by PCO371 (0.1 mmol l−1), hPTH(1–34) (1 μmol l−1 ) or LA-PTH (0.1 μmol l−1) in UMR-106 cells were examined by cAMP washout assay. The cAMP response is expressed as a percentage of the maximal cAMP produced in the cells treated with each ligand (determined by incubating cells concomitantly with the ligand for 10 min, and further incubating with IBMX for 5 min without a washing-out phase). The range of maximum cAMP values was PCO371: 17.4+0.2 pmol per well, hPTH(1–34): 21.3+0.9, LA-PTH: 42.2+0.9, and basal cAMP: 1.8+0.2. (c–g) Effects of PCO371 and hPTH(1–34) on the expression of mRNAs for c-fos (Fos) (c), osteocalcin (Bglap) (d), RANKL (Tnfsf11) (e), osteoprotegerin (Tnfrsf11b) (f) and sclerostin (Sost) (g). mRNA levels are shown as fold-change over control. Gene expression was analysed at 1 h (c) or 6 h (d–g) after treatment of UMR-106 cells with hPTH(1–34) or PCO371. All genes were normalized to 18 S ribosomal RNA. Data are represented as the mean±s.d. of one experiment (n=3 for a–g).
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Related In: Results  -  Collection

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f2: Effects of PCO371 on PTHR1 signalling in UMR-106 cells.(a) cAMP production by hPTH(1–34) and PCO371 in UMR-106 cells, which natively express rat PTHR1. (b) Duration of cAMP-signalling responses induced by PCO371, hPTH(1–34) and LA-PTH in UMR-106 cells. The time courses of cAMP responses produced by PCO371 (0.1 mmol l−1), hPTH(1–34) (1 μmol l−1 ) or LA-PTH (0.1 μmol l−1) in UMR-106 cells were examined by cAMP washout assay. The cAMP response is expressed as a percentage of the maximal cAMP produced in the cells treated with each ligand (determined by incubating cells concomitantly with the ligand for 10 min, and further incubating with IBMX for 5 min without a washing-out phase). The range of maximum cAMP values was PCO371: 17.4+0.2 pmol per well, hPTH(1–34): 21.3+0.9, LA-PTH: 42.2+0.9, and basal cAMP: 1.8+0.2. (c–g) Effects of PCO371 and hPTH(1–34) on the expression of mRNAs for c-fos (Fos) (c), osteocalcin (Bglap) (d), RANKL (Tnfsf11) (e), osteoprotegerin (Tnfrsf11b) (f) and sclerostin (Sost) (g). mRNA levels are shown as fold-change over control. Gene expression was analysed at 1 h (c) or 6 h (d–g) after treatment of UMR-106 cells with hPTH(1–34) or PCO371. All genes were normalized to 18 S ribosomal RNA. Data are represented as the mean±s.d. of one experiment (n=3 for a–g).
Mentions: We also examined whether PCO371 induces cAMP production via endogenously expressed PTHR1 by using UMR-106 cells (a rat osteosarcoma cell line) that express endogenous rat PTHR1. Although the potency was much weaker than that of hPTH(1–34), PCO371 induced cAMP production as did hPTH(1–34), suggesting that PCO371 acts as a full agonist of PTHR1 (Fig. 2a).

View Article: PubMed Central - PubMed

ABSTRACT

Parathyroid hormone (PTH) is essential for calcium homeostasis and its action is mediated by the PTH type 1 receptor (PTHR1), a class B G-protein-coupled receptor. Hypoparathyroidism and osteoporosis can be treated with PTH injections; however, no orally effective PTH analogue is available. Here we show that PCO371 is a novel, orally active small molecule that acts as a full agonist of PTHR1. PCO371 does not affect the PTH type 2 receptor (PTHR2), and analysis using PTHR1–PTHR2 chimeric receptors indicated that Proline 415 of PTHR1 is critical for PCO371-mediated PTHR1 activation. Oral administration of PCO371 to osteopenic rats provokes a significant increase in bone turnover with limited increase in bone mass. In hypocalcemic rats, PCO371 restores serum calcium levels without increasing urinary calcium, and with stronger and longer-lasting effects than PTH injections. These results strongly suggest that PCO371 can provide a new treatment option for PTH-related disorders, including hypoparathyroidism.

No MeSH data available.


Related in: MedlinePlus