Peptide linkers for antibody-drug conjugates (ADCs) are one of the important research directions in the field of cancer treatment. As a bridge connecting antibodies and cytotoxic drugs, peptide linkers are not only crucial for the stability and drug release of ADCs, they can also regulate the release rate, tissue distribution and efficacy of ADCs in vivo. Ideal peptide linkers should have stable chemical properties and the ability to break efficiently under specific conditions (such as acidic environment or enzymatic activity in target cells), thereby ensuring the release of drugs at specific sites. At present, a variety of optimized peptide linkers have been used in clinical and preclinical studies of ADC, some of which have significantly improved the therapeutic effect and safety of anticancer drugs. Research and development of new peptide linkers will continue to promote the development of ADC technology and bring new hope to cancer patients.

Antibody Drug Conjugate

Antibody-drug conjugates represent a cutting-edge class of targeted cancer therapies that combine the specificity of monoclonal antibodies (mAbs) with the potent cytotoxicity of small-molecule drugs. By exploiting the ability of antibodies to recognize and bind to specific antigens expressed on cancer cells, ADCs deliver cytotoxic drugs directly to target cells, minimizing damage to healthy tissue. ADCs are composed of three key components:

Fig. 1. The structure of ADC (Protein Cell. 2018, 9(1): 33-46).

• Monoclonal Antibody (mAb): The antibody serves as the targeting moiety, engineered to specifically recognize and bind to an antigen expressed on the surface of cancer cells. This specificity allows for selective delivery of the cytotoxic payload to cancer cells while sparing normal cells.
• ADC Linker: The linker attaches the cytotoxic drug to the antibody and plays a critical role in ADC stability. A well-designed linker must remain stable in the bloodstream to prevent premature release of the drug but cleave efficiently once the ADC reaches the target site to release the payload. Linkers can be either cleavable or non-cleavable, depending on the intended mechanism of drug release.
• Cytotoxic Drug (Payload): The payload is a highly potent small molecule that can kill cancer cells. These drugs, often too toxic to be administered systemically on their own, include DNA-damaging agents or microtubule inhibitors, which induce apoptosis or disrupt essential cellular functions.

ADCs have emerged as a promising therapeutic strategy for cancer treatment, offering increased efficacy and reduced toxicity compared to traditional chemotherapy. They are particularly useful for treating cancers that express specific surface antigens, such as HER2-positive breast cancer or CD30-positive lymphomas. Beyond oncology, research is expanding into using ADCs for autoimmune diseases and infections, though oncology remains their primary application. The continued refinement of ADC design, particularly in optimizing linkers and improving drug conjugation strategies, is likely to expand their use across a broader range of diseases in the future.

What are Peptides?

Peptides are short chains of amino acids linked by peptide bonds, typically containing 2 to 50 amino acids. They play essential roles in numerous biological processes, such as cell signaling, enzyme activity, and immune function. Due to their versatile nature, peptides have become critical tools in drug development across various therapeutic areas, including oncology, infectious diseases, and autoimmune disorders. In drug development, peptides are valued for their high specificity, low toxicity, and ability to penetrate cells efficiently. One of the key applications of peptides is their use as linkers in ADCs. In ADCs, peptides serve as biodegradable linkers that attach cytotoxic drugs to monoclonal antibodies, ensuring the precise delivery of these potent drugs to target cancer cells. These peptide-based linkers are cleavable by enzymes found in the tumor microenvironment, allowing the controlled release of the cytotoxic payload at the target site. This selective release mechanism reduces systemic toxicity and enhances the therapeutic efficacy of ADCs.

What are Peptide Linker?

Peptide linkers are short sequences of amino acids used in various drug delivery systems to connect two or more functional molecules, such as antibodies and drugs. In drug delivery, peptide linkers help prevent the premature release of a drug, ensuring that the therapeutic payload remains stable as it travels through the bloodstream. Once the drug conjugate reaches the target site, the linker is cleaved by specific enzymes or environmental factors, releasing the drug. Peptide linkers are often designed to be sensitive to proteases in the tumor microenvironment, making them particularly valuable in oncology, especially in ADCs.

• Advantages of Peptide Linkers

Peptide linkers are highly customizable, allowing for the incorporation of specific sequences that respond to environmental stimuli, such as pH changes or enzymatic activity. This tunability makes them ideal for targeted therapies like ADCs, where precise control over drug release is critical. Moreover, their biocompatibility ensures that they degrade into harmless amino acids after performing their function, minimizing any potential toxicity or immune response.

• Applications of Peptide Linkers

Peptide linkers have broad applications in drug development, especially in the creation of advanced therapeutic agents like ADCs, peptide-drug conjugates (PDCs), and nanocarrier systems:

• Antibody-Drug Conjugates (ADCs): In ADCs, peptide linkers connect a monoclonal antibody (mAb) to a cytotoxic drug. The antibody acts as the targeting agent, binding to antigens on the surface of cancer cells, while the peptide linker holds the cytotoxic drug in place until the ADC reaches the tumor site. Enzymes in the tumor microenvironment, such as cathepsins or matrix metalloproteinases, then cleave the peptide linker, releasing the drug and allowing it to exert its therapeutic effects. This targeted release minimizes systemic toxicity and increases the efficacy of the drug.
• Peptide-Drug Conjugates (PDCs): Similar to ADCs, PDCs use peptides as both the targeting moiety and the linker. Peptide linkers in PDCs improve the pharmacokinetics and specificity of these conjugates, ensuring that the drug is released only when it reaches its intended target, such as cancer cells or infected tissues.
• Nanocarriers and Drug Delivery Systems: Peptide linkers are also used in nanocarrier systems, such as liposomes and nanoparticles, to control the release of therapeutic agents. In these systems, peptide linkers provide an additional layer of control, allowing the drug to be released only when the nanocarrier reaches the desired tissue or cell type. This precise release mechanism enhances the therapeutic effect while reducing adverse side effects.

Peptide Linker from BOC Sciences

Peptide Linker for ADCs

In the ADC development process, peptide linkers play a key role. They connect antibodies and toxins and ensure that the drug is released efficiently in a specific environment to avoid toxicity to normal cells. The selection and design of peptide linkers directly affect the effectiveness, specificity and safety of the drug. The following are several commonly used types of peptide linkers and specific examples.

Fig. 2. ADC with peptide linker (MAbs. 2021, 13(1): 1951427).

• Val-Cit Linker

The Val-Cit linker is a widely used dipeptide linker in ADC development. It is composed of the amino acids valine and citrulline and is specifically cleaved by protease B. This linker exhibits excellent plasma stability and efficient cleavage in the presence of protease B, making it one of the most reliable linkers for drug delivery applications. An example of an ADC using the Val-Cit linker is brentuximab vedotin (Adcetris), which is used to treat certain types of lymphoma. The antibody component targets CD30-expressing lymphoma cells, and the cytotoxic drug monomethyl auristatin E (MMAE) is attached via the Val-Cit linker, ensuring the selective delivery and release of MMAE within cancer cells.

• Val-Ala Linker

The Val-Ala linker consists of valine and alanine and can be cleaved by proteases in lysosomes. In contrast to Val-Cit, which has difficulty achieving high DARs due to precipitation and aggregation, the Val-Ala linker can achieve DARs as high as 7.4. Val-Ala has a higher hydrophilicity, which makes this linker more advantageous in the context of lipophilic payloads such as PBD-dimers. The Val-Ala linker has been successfully used in the development of loncastuximab tesirine. In addition to dipeptide linkers, tetrapeptide Gly-Gly-Phe-Gly has also been successfully used in ADC drugs. Compared with dipeptides, tetrapeptide linkers are more stable in blood circulation, and the marketed ADC drug Enhertu uses this type of linker.

Peptide Linker Design

In the development of ADC, the selection and design of peptide linkers are crucial, which directly affects the stability, targeting and effective release of toxins. The design of linkers needs to consider the physiological environment of target cells, such as lysosomal protease activity or the pH of the microenvironment. For example, linkers such as Val-Cit and Phe-Lys utilize protease degradation in lysosomes and are suitable for treating cancers that require targeting the internal environment of cells, while linkers such as GFLG and GGFG are designed for rapid toxin release. In addition, the chemical stability of the linker is also an important factor in the design to ensure that it does not decompose prematurely in the blood circulation to avoid premature release of toxins. Design optimization usually also considers the physicochemical properties of the toxin, as well as the interaction between the antibody and the target, so that the linker is cut in a suitable intracellular environment, thereby precisely controlling the release of the drug. This precision design can improve the efficacy of ADCs and reduce side effects on normal tissues.

Based on this, BOC Sciences provides comprehensive peptide linker development services to meet the various needs of the pharmaceutical and biotechnology industries. Our expertise in designing and synthesizing peptide linkers ensures optimal stability, bioavailability and specificity of the conjugated molecules. We can customize peptide linkers according to customer requirements, and from the initial concept design to the final synthesis, BOC Sciences ensures high-quality products by using advanced analytical techniques and strict quality control measures.

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