Process Intensification for the Purification of Peptidomimetics: The Case of Icatibant through Multicolumn Countercurrent Solvent Gradient Purification (MCSGP)

Abstract

Biopharmaceuticals are subjected to very strict purity requirements to be marketed. At the same time, peptides and other biomolecules are industrially synthesized through techniques (e.g., solid-phase synthesis) often leading to the formation of many impurities with molecular characteristics very similar to the target product. Therefore, the purification of these mixtures via preparative chromatography can be very challenging. This typically involves ternary or central-cut separations, characterized by chromatograms where the central peak, corresponding to the target product, exhibits significant overlapping on both sides with impurities slightly more or less adsorbable. In single-column (batch) preparative chromatography, this leads to a typical yield-purity tradeoff, meaning that high purity can be obtained at the cost of low yield and vice versa, with obvious consequences on the overall production costs. This study demonstrates how this limitation can be alleviated using the continuous countercurrent operating mode, conducted on a multicolumn system, as a tool for process intensification. In particular, the Multicolumn Countercurrent Solvent Gradient Purification (MCSGP) process has been applied to the purification of an industrial crude mixture of icatibant, which is a peptidomimetic antagonist of bradykinin B2-receptor that has been recently also considered for the treatment of patients affected by COVID-19 disease. It is shown that MCSGP allows conjugating process simplicity (using only two columns) with a significant improvement in process performance, compared to the corresponding batch process. This includes all process performance parameters: yield, productivity, and buffer consumption for a given purity specification of icatibant.

Benefits of a Mixed-Mode Stationary Phase to Address the Challenging Purification of an Industrially Relevant Peptide: A Proof-of-Concept Study

“…For each column, a so-called Pareto curve was also obtained, starting from the offline analysis of the fractions collected along the gradient. This curve reports the variation in purity with recovery [2,16,28]. These two parameters vary inversely: the purest fraction of the peak only contains a small percentage of the peptide mass injected.…”

“…In the last decades, the market of biopharmaceuticals, including protein-, peptideand oligonucleotide-based therapeutics, has experienced an impressive increase due to the biological properties of this new class of molecules being potentially promising for medical applications [1][2][3][4]. Recent advancements in the production strategies, such as in genomics, proteomics recombinant strategies and peptide synthesis, have pushed even further the development of these new drugs [5][6][7][8].…”

UV‐based dynamic control improves the robustness of multicolumn countercurrent solvent gradient purification of oligonucleotides

“…On the other side, Multicolumn Countercurrent Solvent Gradient Purification (MCSGP) is an effective and practical means to overcome the yield‐purity trade‐off for different biomolecules, including peptides [ 12,13 ] monoclonal antibodies [ 14–16 ] and PEGylated proteins [ 17–19 ] Over the past years, the twin‐column embodiment of this continuous chromatography technology has become predominant. The interested reader is referred to the literature [ 20–22 ] and to the Supporting Information for a detailed description of MCSGP, with Figure S1 showing the different steps of the process.…”

“…On the other side, Multicolumn Countercurrent Solvent Gradient Purification (MCSGP) is an effective and practical means to overcome the yield-purity trade-off for different biomolecules, including peptides [12,13] monoclonal antibodies [14][15][16] and PEGylated proteins [17][18][19] Over the past years, the twin-column embodiment of this continuous chromatography technology has become predominant.…”