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As a one-stop CRDMO, we conjugate proteins, mainly antibodies with drugs, small molecules, peptides, dyes, or oligonucleotides to create antibody-drug conjugates (ADCs) or other protein conjugates.
The starting point can be in-house protein production, internal antibody discovery programs, or with client-supplied proteins. This way we create ADCs, protein-drug conjugates, antibody-peptide conjugates, and peptide-drug conjugates, to name a few.
We can use various conjugation methods to chemically link a protein with another molecule, utilizing a wide selection of commercially available or client-specific linker-payload combinations. In addition to chemical conjugation, we can fuse cytokines to antibodies, typically through genetic fusion.
By analyzing crucial ADC parameters from the very beginning, we optimize your ADC molecules for solubility, monomeric state, homogeneity, cell-killing potency, and other crucial biophysical and biochemical parameters right from the start.
Using a range of biochemical & biophysical analytical methods, we evaluate conjugates for drug-to-antibody ratio (DAR), typically ranging from 2 to 8, as well as conjugation site occupancy, internalization, and killing potency in cell-based activity assays (CBA). This enables us to produce ADC molecules that meet previously defined specifications for later large-scale clinical and commercial production.
Our dedicated team of cell-based activity assay scientists has a range of CBAs already established or, if needed, can readily design and implement novel ones. Routine assays involve target binding, internalization, and cell-killing potency assays.
To conjugate molecules, we use engineered cysteines (THIOMAB™), random lysines or cysteines, and enzymatic conjugation methods.
Read more from the antibody discovery page.
Antibody-drug conjugates (ADCs) are targeted cancer therapies that combine the specificity of antibodies with the potency of cytotoxic small-molecule drugs. By attaching powerful cell-killing agents to antibodies against tumor-specific cell surface markers, ADCs deliver highly potent drugs precisely to cancer cells, thus minimizing systemic damage to healthy tissues. This targeted approach reduces off-target side effects typically seen in traditional chemotherapy and at the same time confers the otherwise often-lacking effectiveness of an unconjugated antibody. ADCs are increasingly used in treating hematological cancers and solid tumors, offering new therapeutic options for otherwise intractable cancers.
In biotechnology, protein and antibody conjugation techniques are essential for drug development, diagnostics, and research. Understanding and being able to apply different conjugation methods is crucial for achieving the desired specificity, stability, and functionality in biomolecule conjugation, ultimately contributing to therapeutic success. Icosagen currently applies targeted cysteine conjugation, random lysine or cysteine conjugation, and enzymatic conjugation methods.
Targeted cysteine conjugation
Targeted cysteine conjugation is a highly selective method used to modify proteins site-specifically. This technique exploits the thiol (-SH) group in cysteine residues, allowing for precise control over conjugation sites. Since cysteines are less abundant in proteins than lysins, this method enables site-specific labeling, protecting functional or active sites. This approach is widely applied in antibody-drug conjugates (ADCs), where precise targeting is critical for therapeutic efficacy and minimized off-target effects.
Random lysine and cysteine conjugation
Random lysine conjugation involves modifying the amino (-NH2) groups of lysine residues. This method is less specific than targeted cysteine conjugation, as lysine is typically more abundant in proteins. Random cysteine conjugation occurs via thiol (-SH) groups of cysteine residues. It involves the selective, partial reduction of disulfide bridges and coupling the small molecule drug via the initial S=S bond. Despite being less specific, these methods can quickly and efficiently conjugate substances to a wide range of proteins, such as enzymes, antibodies, or peptides, where site specificity is at least initially not critical. It is often used in the development of protein-protein interactions or immunoassays, where random yet stable conjugation is sufficient.
Enzymatic conjugation
Enzymatic conjugation offers a highly specific and biocompatible method for modifying proteins or antibodies. This technique uses enzymes such as sortase, transglutaminase, or glycosyltransferases to attach labels, payloads, or linkers to proteins at defined sites. The enzyme recognizes a specific consensus sequence of amino acid residues for precise modification with minimal chemical impact on the protein’s overall structure or function. Enzymatic conjugation is commonly used in the development of biopharmaceuticals, antibody-drug conjugates, and site-specific labeling for imaging or therapeutic applications.
In antibody-drug conjugates, selecting the right payload (and linker) is critical for optimal therapeutic efficacy and minimal side effects. These payloads are typically cytotoxic agents, often microtubule polymerization or topoisomerase inhibitors, but can also be peptides, or other molecular tools used for specific therapeutic, research, or diagnostic applications. Below are some typical payloads used in ADCs:
Cytotoxic payloads: MMAE, MMAF, EXATECAN
Peptides
Peptides serve as versatile payloads in both therapeutic and diagnostic ADCs. They can be used to block specific signaling pathways, target cancer cells, or deliver therapeutic agents to specific tissues. Peptides are also ideal for targeting cell surface receptors or enzymes in precise targeted therapies.
Fluorophores
Fluorophores, such as those provided in DyLight kits, are commonly used for fluorescent labeling of antibodies or proteins in vivo or in vitro imaging studies. Although not typically used in ADCs for therapeutic purposes, fluorophores are essential in research applications to monitor biomolecule interactions and track cellular processes in real time and can be used as theranostics.
Biotinylation and PEGylation In addition to drug conjugation, we offer:
To provide an integrated ADC service, comprehensive analytical capabilities are a must. At Icosagen this includes:
Cell-based activity assays are integral in understanding the biological potency of ADCs. These assays closely mimic in vivo conditions, allowing for a more predictive assessment of an ADC’s therapeutic performance, such as:
We offer specialized cell-based activity assays to meet your unique ADC development program needs. Our team of CBA scientists have rich experience, and a range of assays already established and can readily adapt existing assays to novel targets, or even design and implement novel cell-based activity assays from scratch. For this, a library of off-the-shelf reporter cell lines are available.
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