Tedoglutide is a glucagon-like peptide 2 (GLP-2) analogue that reduces gastric emptying and secretion and regulates the growth, proliferation, and repair of small intestinal lining cells. The peptide increases the number of these cells, which increases intestinal absorption and reduces diarrhea. Clinically used for *** short bowel syndrome. The drug became the first drug to be recommended for this rare but extremely debilitating disease.
Tedoglutide chemical formula.
Basic parameters
Name: Teduglutide
Designation: GT-F027
CAS Number: 197922-42-2
Sequence: his-gly-asp-gly-ser-phe-ser-asp-glu-met-asn-thr-ile-leu-asp-asn-leu-ala-ala-arg-asp-phe-ile-asn-trp-leu-ile-gln-thr-lys-ile-thr-asp
MF: C164H252N44O55S
Molecular weight: 375208248
Tedoglutide HPLC graph.
Tedoglutide
At present, there are two main synthesis methods of tildoglutide: sequential coupling and fragment coupling, in which sequential coupling is based on Wang resin as the carrier, and the peptide is synthesized by pure solid-phase sequential coupling, and finally cleaved to obtain teduglutide.
Due to the long peptide chain of teduglutide, the synthesis by sequential coupling will increase the difficulty of synthesis, and produce impurities that are difficult to purify, reducing the ** rate. More importantly, the N-terminal 3rd and 4th amino acids of toxydoglutide are ASP-GLY, and the carboxyl group in the aspartic acid (ASP) side chain can form a five-membered ring with -H in the amide bond, once this five-membered ring structure is formed, it will be degraded and opened under certain conditions, and a variety of impurities will be produced in the process of ring opening, such as: racemic and production of ASP, aspartic acid, etc., resulting in the decline of the purity of teduglutide.
MS plot of texeglutide.
1.Synthesis of fmoc-his(trt)-gly-oh (peptide fragment 1).
Add 1220g (2mol) fmoc-his(trt)-OH and 230g (2mol) HOSU to a 10L three-port glass reaction flask, dissolve with 4000ml DMF for 30 minutes, then add 412g DIC to stir the reaction at room temperature for 2 hours, and finally add 402g (2mol) Gly-obzlHCL reacted with 300ml (2mol) triethylamine for 10 hours, and the reaction was condensed into FMOC-his(TRT)-GlyOBZL protects the dipeptide. React with 10 times the liquid to precipitate with water, filter and collect the precipitate, and wash it with a large amount of water, and dry it under reduced pressure. After passing the inspection, the solid is dissolved with 50 ethanol aqueous solution, 50g of 5 PD C is added, catalytic hydrogenolysis is 16 hours, the ethanol is concentrated, and the filtered product is dried under reduced pressure to obtain the final product 6442 g fmoc-his(trt)-gly-oh in yield 476％。
2.Synthesis of FMOC-ASP (OTBu)-Gly-OH (peptide fragment 2).
Add 422g (2mol) fmoc-asp (otbu)-OH and 230g (2mol) HOSU to a 10L three-port glass reaction flask, dissolve with 4000ml DMF for 30 min, then add 412g DIC and stir the reaction at room temperature for 2 h, and finally add 402g (2mol) Gly-obzlHCl reacted with 300ml (2mol) of triethylamine for 10 hours to complete the condensation of FMOC-ASP (OTBu)-GlyOBZL protects the dipeptide. React with 10 times the liquid to precipitate with water, filter and collect the precipitate, and wash it with a large amount of water, and dry it under reduced pressure. After passing the inspection, the solid is dissolved with 50 ethanol aqueous solution, 50g of 5 PD C is added, catalytic hydrogenolysis is 16 hours, the ethanol is concentrated, and the filtered product is dried under reduced pressure to obtain the final product 4021 g fmoc-asp(otbu)-gly-oh in yield 429％。mw:468。
3.Synthesis of FMOC-ASP (OTBU)-2-Cl TRT-resin
Take the substitution value of 500g as 06 mmolg of 2-Cl TRT-Cl resin with the addition of DMF swelling resin. Take 06mol FMOC-ASP (OTBu), dissolved with an appropriate amount of DMF, added to the above resin, stirred evenly and then added 12mol DIEA, stirring the reaction for 3 hours, extracting the reaction solution, after DMF washing 3 times, DCM washing 3 times, fmoc-asp(otbu)-2-cl TRT-resin was obtained, and the substitution value was 046mmol/g。
Diagram of the finished product of teduglutide.
4.Synthesis of FMOC-ASP (OTBU)-Wang resin
Take the substitution value of 500g as 05 mmol g of Wang resin, with the addition of DMF swelling resin. Take 05mol FMOC-ASP (OTBu), dissolved with an appropriate amount of DMF, added to the above resin, stirred evenly and then added 10mol dic、0.4molhobt、0.04mol 4-N,N-dimethylpyridine, stirred the reaction for 6 hours, the reaction solution was removed, DMF was washed 3 times, DCM was washed 3 times, and FMC-ASP (OTBU)-Wang resin was obtained with a substitution value of 041mmol/g。
5.Synthesis of peptide fragment 3 and ticoliglutide resin
Take 01mol of FMOC-ASP (OTBU)-2-Cl TRT-resin (substitution value of about 0.) in Example 446 mmolg), de-protect with 20 pip DMF solution for 25 min, wash and filter, and obtain fmoc-depleted H-asp(OTBu)-2-Cl TRT-resin.
Take 03mol FMOC-THR (TBU) and 03mol HOBT, dissolved with an appropriate amount of DMF;Take another 03mol DIC, slowly added under stirring, continued to stir the reaction for 30 minutes, added to the above H-ASP (OTBu)-2-Cl TRT-resin, coupling reaction for 120 300 minutes, the end point of the reaction is subject to ninhydrin detection, washing and filtration, and then 20 pip DMF solution to protect for 25 minutes, washing and filtering, to obtain H-thr(TBU)-Lys(BOC)-2-Cl TRT-resin. Peptide fragment 3 is obtained
h-ser(tbu)-phe-ser(tbu)-asp(otbu)-glu(otbu)-met-asn(trt)-thr(tbu)-ile- leu-asp(otbu)-asn(trt)-leu-ala-ala-arg(pbf)-asp(otbu)-phe-ile-asn(trt)-trp (BOC)-Leu-Ile-GLN(TRT)-THR(TBU)-Lys(BOC)-Ile-THR(TBU)-ASP (OTBU)-2-Cl TRT-Resin.
6.Conjugation was performed to obtain teduglutide resin:
h-his(trt)-gly-asp(otbu)-gly-ser(tbu)-phe-ser(tbu)-asp(otbu)-glu(otbu )-met-asn(trt)-thr(tbu)-ile-leu-asp(otbu)-asn(trt)-leu-ala-ala-arg(pbf)-asp (otbu)-phe-ile-asn(trt)-trp(boc)-leu-ile-gln(trt)-thr(tbu)-lys(boc)-ile-thr(tb u)-asp(otbu)-2-Cl TRT-resin.