Date: 7 August 2024 | Category: News
Continuous-Flow Solid-Phase Peptide Synthesis to Enable Rapid, Multigram Deliveries of Peptides
Kyle E. Ruhl*, Michael J. Di Maso*, Harrison B. Rose, Danielle M. Schultz, François Lévesque, Shane T. Grosser, Steven M. Silverman, Shasha Li, Nunzio Sciammetta, and Umar Faruk Mansoor
The Process Research and Development team at Merck in New Jersey have successfully used continuous-flow solid-phase peptide synthesis (CF-SPPS) to prepare 20 g of a single peptide in under 4 hours in high yield and purity over a single run with the Vapourtec SPPS system [1]. For this work, CF-SPPS provided significant advantage when compared with batch mode, including rapid, data-rich optimisation of peptide sequences, and significant reduction of development and process execution timelines and waste generation.
Peptides are becoming increasingly prolific within medicine due to their therapeutical application over a range of diseases and conditions. Examples include semaglutide (Wegovy or Ozempic), a treatment for type 2 diabetes and weight loss, as well as octreotide, a treatment for acromegaly, and insulin. In recent years macrocyclic peptides containing noncanonical amino acids have also been developed, displaying better resistance to protease degradation, reduced entropic barriers to receptor binding and improved oral pharmacokinetic profiles.
The traditional route to preparation of these materials is through solid-phase peptide synthesis (SPPS), but this has several disadvantages including scale limitations and the requirement for significant manual input. In addition, traditional SPPS generates large quantities of waste so is ‘less green’ than other approaches. Until recently, transfer into flow mode was hampered by virtue of the polymeric resins used swelling considerably in organic solvents, causing difficulties when using fixed-volume packed-bed reactors (PBRs).
The development of a variable-bed flow reactor (VBFR) by Vapourtec and Seeberger [2], where the VBFR employs a stepper motor and two pressure sensors to constantly monitor and adjust the bed volume in response to increasing or decreasing pressures, has facilitated development of CF-SPPS as a viable tool. Use of this CF-SPPS system by Merck dramatically reduced synthesis times of complex peptides from 1 to 2 weeks to 1 day, allowing reduced quantities of the custom-made amino acids to be used therefore minimising costs, reducing solvent consumption and giving access to multi-gram quantities of material. In addition, impurities and poorly performing coupling steps, particularly upon scale-up, could be quickly identified through use of a UV-vis detector, also reported by Vapourtec [2]. Finally, on larger scales, as this work was performed before the Vapourtec scale-up VBFR was available, a jacketed heat exchanger could be used to control activation time and temperature, as in this case lower bed temperatures were crucial for preventing thermal cleavage of the growing peptide from the resin.
References
[1] Continuous-Flow Solid-Phase Peptide Synthesis to Enable Rapid, Multigram Deliveries of Peptides (K. E. Ruhl, M. J. Di Maso, H. B. Rose, D. M. Schultz, F. Lévesque, S. T. Grosser, S. M. Silverman, S. Li, N. Sciammetta, and U. F. Mansoor, Org. Proc. Res. Dev., 2024, 28, 7, 2896–2905). https://pubs.acs.org/doi/10.1021/acs.oprd.4c00165
[2] Real-time monitoring of solid-phase peptide synthesis using a variable bed flow reactor (E. T. Sletten, M. Nuño, D. Guthrie, and P. H. Seeberger, Chem. Commun., 2019, 55, 14598–14601). https://doi.org/10.1039/C9CC08421E
