Sustainable Strategies for Peptide, Oligonucleotide, and Bioconjugate Production in Pharma and Allied Fields

The use of peptides, oligonucleotides, and bioconjugates as therapeutics has experienced rapid and transformative growth in recent years, delivering substantial benefits to patient health and well-being. However, the production of these types of therapeutics has historically utilized processes that involve the extensive use of hazardous reagents, large volumes of solvents, and high process mass intensities. The ACS Green Chemistry Institute Pharmaceutical Roundtable is a forum where global pharmaceutical and allied industries collaborate to advance the sustainability of manufacturing medicines by implementing green chemistry & engineering. An overview of the sustainability initiatives and work advanced by three of the ACS GCIPR Focus teams – Peptides, Oligonucleotides, and Bioconjugates – will be highlighted.

Chemical assembly strategies for the manufacture of long therapeutic peptides typically require side-chain protected segments. In order to combine fully unprotected peptides at arbitrary amino acid junctions, we have developed new variants of the alpha-ketoacid–hydroxyl amine (KAHA) ligation. Through the construction of cyclic hydroxylamines and straightforward methods to install alpha-ketoacid onto synthetic and recombinant peptides, we have established direct routes to therapeutic peptides including tirzepatide. These chemoselective ligations proceed in sustainable solvents including DMSO/water and AcOH and can be translated to GMP manufacturing of therapeutic targets.

Peptide synthesis is facing major global challenges in efficiency and sustainability as demand for peptide therapeutics rapidly increases. To address this, we developed a nature-inspired continuous N-to-C chain elongation platform that assembles peptides directly from unprotected amino acids, delivering high yield couplings. This new platform also features portability, scalability, and no specialized equipment required. Solid phase peptide synthesis (SPPS) has been the gold standard due to its high coupling efficiency and its ability to rapidly assemble long chains, but its C–N elongation strategy requires Fmoc-protected amino acids, which involves multiple nonproductive steps to achieve, leading to high reagent/solvent use (high PMI). An N–C elongation strategy using unprotected amino acids offers much improved atom and step economy by forming amide bonds directly. However, challenges such as epimerization, over addition, and remaining starting materials have limited its application. Here, we report a chemical N–C continuous elongation strategy using free amino acids, achieving seven consecutive couplings with 96% yield per coupling without intermediate isolations. The full chain assembly is more efficient than the Fmoc protection steps required in traditional synthesis. Unlike biosynthesis, this method accepts non-canonical AAs that are critical for drug like properties. Demonstrated on marketed peptide drugs, this approach reduces isolations by ~10 fold and waste generation by ~100 fold. Longer peptides can be readily assembled with continuous fragment couplings. The platform has the potential to offer a greener and more sustainable peptide manufacturing platform and ultimately enable broader patient access.

The construction of amide bonds, ubiquitous in biological molecules and pharmaceutical targets, is typically conducted with peptide coupling reagents or other stoichiometric activators that generate significant waste on process scale and are often associated with safety concerns. Boronic acid catalysts have shown promise as alternatives but are usually limited to elevated temperatures and/or have restricted substrate scopes on account of the energetic cost to eliminating water in the rate-limiting step. We have designed a new bifunctional organocatalyst that incorporates a boronic acid functionality in combination with a P(V) center as the second binding site to facilitate catalysis at room temperature with a scope of substrates that includes acyclic secondary amines and heterocycles of interest in the pharmaceutical industry. In addition to its synthetic utility, we identified novel mechanistic features that showcase how the combination of the boronic acid and phosphine oxide enable catalysis through a different monomeric mechanism to previously reported arylboronic acid catalysts, which form an activated bicyclic intermediate. In this presentation, the optimization and scope of the catalytic conditions will be discussed, applications in peptide synthesis presented, and mechanistic studies that show differences from previously published arylboronic acid catalysts will be presented.

Peptides are reshaping the pharmaceutical landscape, yet chemical manufacturing mainly hinges on the dipolar aprotic solvents N,N-dimethylformamide (DMF) and N-methyl-2-pyrrolidone (NMP) which are toxic to both humans and the environment. We introduce trioxabicyclooctanes (TBOs), tunable dipolar solvents made in two steps from bio glycerol, as drop in replacements.
The lead TBO solvent, MTBO, accelerates couplings and enables reducing amino acid equivalents from 2.0 to 1.1, while maintaining performance. In the synthesis of difficult sequences in batch (Barstar[75–90], 68% vs 65% in DMF) and in flow (Neurokinin A, 68% vs 51% in DMF), crude purities are improved compared to DMF. Practical handling includes moderate boiling points and viscosities, and benign toxicology. Technoeconomic analysis shows that the two-step process can lead to pricing competitive to DMF. Our findings show how waste-glycerol can be turned into performant DMF replacements without relying on N or S atoms, for SPPS, and the broader chemical industry.

This presentation details the development of a dynamic process model designed to align early-stage oligonucleotide synthetic processes with ambitious corporate sustainability targets, specifically focusing on carbon footprint and toxic material consumption. By coupling a Bill of Materials with carbon emission factors and SVHC (Substance of Very High Concern) designations, the model quantifies environmental impacts on both a mass basis and a per-patient-per-year metric.
A case study on oligonucleotide manufacturing is used to demonstrate how this data-driven approach identifies gaps and quantifies the environmental impact of process optimizations. Equipped with these insights, scientists can identify and apply green chemistry opportunities early in oligonucleotide process development to ensure future manufacturing processes meet stringent launch limits.

Downstream separation and cleaning dominate the material intensity of oligonucleotide manufacturing. Conventional solid-phase synthesis exhibits a process mass intensity of ~4,000–5,000 kg per kg product, largely due to purification and wash solvents. Reducing chromatographic steps and developing regenerable separation platforms is therefore essential to lower solvent use and waste generation.
Polymer brush–based affinity materials offer a promising alternative by providing high loading capacities and stimulus-responsive release. Polymer brushes consist of densely grafted polymer chains that create a confined, highly functionalized environment capable of capturing and releasing biomolecules in response to pH.
Here, we report the fabrication and characterization of pH-responsive polymer brush coatings based on poly(2-(diethylamino)ethyl methacrylate) (PDEAMA) and PDEAMA/poly(methacrylic acid) (PMAA) copolymers grafted from gold or glass substrates. A modified surface-initiated polymerization eliminates the need for an inert atmosphere and significantly reduces solvent consumption, enabling synthesis in volumes as low as 3 mL or within a confined droplet between two glass slides.
Interactions with oligonucleotides (10-, 20-, 30-, and 40-mers) were studied as a function of brush composition, thickness, oligonucleotide length, and pH using surface plasmon resonance, quartz crystal microbalance, and fluorescence microscopy. Binding capacity normalized to brush thickness was independent of copolymer composition up to 20% PMAA. In contrast, release behavior depended on oligonucleotide length and brush composition, with an 80:20 PDEAMA:PMAA brush enabling complete release at pH 10 and a pH window of 8–9 identified as optimal for size-based separation.

Bioconjugations play an important role in next-generation therapeutic agents, as exemplified by antibody drug conjugates. New methodologies for site-specific bioconjugations allow for the design and manufacture of improved therapies with enhanced homogeneity, leading to a more consistent therapeutic profile. Biohaven’s Multimodal Antibody Therapy Enhancer (MATE^TM) allows for highly efficient and selective bioconjugations with ‘off-the-shelf’ therapeutic monoclonal antibodies (mAbs) and a range of cytotoxic and non-cytotoxic payloads. The design and application of the MATE^TM technology to ADCs and pathogenic protein degraders, including kilogram scale manufacture and green chemistry implications, will be discussed.