The potency, selectivity and high target specificity of peptides and proteins as immunotherapeutic agents had initially been associated with major obstacles towards their drug development and translation. Peptides are intrinsically unstable towards proteolytic degradation, renal clearance and low cell permeability, limiting their in vivo half-life and tissue distribution. Peptides' high target affinity is on one hand a pre-condition for their high target specificity, but on the other hand often also the basis for their high immunogenicity. This constitutes the so called "immunotherapy paradox". Over the years, a range of chemical modifications has been introduced to address these obstacles towards translation. Modifications can be made to the peptide's backbone, amino acids or termini to generate "PEGylation", "lipidation", "cyclization" or other chimeric conjugates that on one hand maintain or even improve their target specificity, but on the other hand also resemble small molecules in terms of their stability and pharmacokinetic properties. These approaches significantly prolong the circulating half-life of peptides in vivo, and improve their tissue penetration and intracellular delivery. The result is a class of peptides with the desired anti-cancer or anti-infectious immune modulation capacity, but without the need for continuous infusion or high dosing frequencies.
Study of molecular mechanisms in experimental immunology requires antigens or other reagents to be stable in serum, in endosomal compartments and, for some purposes, in lysosomal hydrolases long enough to be mechanistically informative. Unmodified peptides often have a very short half-life of a few minutes, which not only makes it difficult to obtain a clear dose–response, but can also be a major source of inter-assay variability. Thus, modified analogues of native peptides often provide an effective surrogate, giving the investigator time to study processes such as T-cell priming, antibody affinity maturation or engagement of the innate sensors. In addition, chemical modification allows orthogonal addition of functional groups such as fluorophores, click-handles for affinity purifications or linkers for controlled release, thereby enabling the peptide to serve as a multifunctional molecular probe.
Fig. 1 Strategies for the design and modification of peptides.1,5
Short half-life is the most important challenge for therapeutic peptides. The extensive distribution of endo- and exopeptidases throughout the body enables the ubiquitous degradation of peptides, as the amide bonds that connect amino acids can be easily hydrolyzed. Modifying the structure of peptides can increase their proteolytic stability. Backbone methylation or N-alkylation, D- or β-amino acids, unnatural amino acids, as well as cyclisation through lactam bonds, disulfide bonds, or ring-closing metathesis can all be used to shield peptides from enzymatic degradation. Cyclisation can also prevent the elimination of these peptides in the kidney. Retro-inverso isomerization, which involves inverting and reversing the sequence of the peptides, can also protect against enzymatic degradation. Reversing and inverting a peptide chain results in an unnatural peptide with the same biological activity. Polymeric modifications, such as PEGylation or hydroxyethyl starch modification, can increase the hydrodynamic volume of the peptides and protect them from enzymatic degradation. This also prolongs the circulation time of the peptide by slowing renal filtration.
One challenge for these hydrophilic entities is the fact that aqueous solubility is often antithetical to membrane crossing and tissue distribution. Efforts to balance these properties led to the development of amphipathic engineering approaches. Lipidation (most often through myristoyl, palmitoyl or cholesteryl groups) enables peptides to partition into cell membranes, promoting paracellular or transcellular absorption, and tether the payload to albumin to promote hitchhiking in the bloodstream. On the other hand, self-assembling peptide amphiphiles can self-assemble into micelles or nanofibrils and allow co-loading of hydrophobic adjuvants or nucleic acids, to achieve co-delivery of immunostimulatory agents. For mucosal or oral administration, cell-penetrating peptide sequences such as penetratin or TAT can be fused to the therapeutic antigen to induce energy-dependent endocytosis while retaining epitope integrity. In addition to enabling new routes of administration, many of these delivery-oriented modifications promote the accumulation of immunomodulators in immunologically relevant anatomical sites (i.e. lymph nodes, mucosal lamina propria, or the hypoxic centers of tumors) where they can exert their activity.
The major drawback of having a short systemic half-life is the need for repeated administrations. There are several strategies to prolong the chemical half-life, mainly by targeting the elimination kinetics. Linking a peptide to albumin, for example, can make use of the FcRn recycling pathway, and gives the peptide an albumin-like half-life in circulation. XTENylation/elastin-like polypeptide fusion can also prolong the half-life, due to the addition of a large, unstructured, hydrophilic moiety that increases the hydrodynamic size and reduces glomerular filtration, without being immunogenic. To provide a pulsatile instead of a sustained delivery of the payload, disulfide-, hydrazone-, or ester-based linker can be used that are sensitive to reductive cleavage, acidic pH, or esterase degradation, respectively. Thus, these peptides become "masked" until they reach the tumor microenvironment (acidic pH) or the cytosol (due to reductive cleavage of the disulfide bonds).
Building on the synergies between synthetic chemistry and translational immunology, we offer a growing library of peptide modifications, which converts potential molecular liabilities into therapeutic opportunities. In place of a static collection of options, we present a continuum of design space in which peptide cyclization, PEGylation and lipidation are tuned as structural motifs. These motifs may be applied independently, or in concert as structural polypharmacy. The unifying concept is one of situational alchemy: every peptide can be seen as a mutable document whose in vivo destiny can be altered by chemical annotations. The end result is a molecule that preserves the information specificity of the amino-acid sequence but that gains the structural stability, vascular-penetrating and membrane trafficking characteristics more commonly associated with larger, or otherwise different drug modalities. In other words, we offer not only modifications, but also passports that allow therapeutic peptides to navigate the complex and often hostile environments of serum, endothelium and cytoplasm, without losing their immunological expressivity.
Cyclization takes these peptides out of an open conformation by forming topologically inner rings of molecules which are not susceptible to enzyme attack. Cyclization can be performed with many types of chemistries such as head-to-tail peptide lactam bonds, disulfide bridges, olefinic crosslinks, and hetero-bifunctional triazole crosslinks. The choice of ring-forming chemistry can be optimized in silico using a flexible molecule to predict the conformation and compare both entropic and enthalpic changes to the molecule. Side chain to backbone cyclization can be performed when contact residues are needed to be kept available for receptor interactions, while backbone to backbone cyclization has been used in cases where very rigid and protein fold mimicking molecules are desired. The final macrocycle is usually re-simulated to make sure that the introduced ring strain does not decrease the activity. However, the closed constrained topology of the cyclized peptide may also relieve entropic loss, expose additional binding sites, or impose helicity in an otherwise unfolded peptide, resulting in increased activity in addition to increased stability.
PEGylation involves more than simply grafting polymers onto a peptide. Linear, branched and comb-shaped polyethylene glycols (PEGs) of different molecular weight are chosen according to the peptide's charge, hydrophobicity and desired half-life. Linkers can be engineered to either permanently shield or allow responsive shedding of the polymer: reductively cleavable disulfides and acid-cleavable hydrazones conditionally restore the peptide in the reducing cytosol and tumor acidic microenvironment respectively. Zwitterionic and polysarcosine alternatives are available to avoid anti-PEG antibodies. The polymer's density can be controlled so that it forms a molecular "corona" which sterically prevents opsonization but does not occlude receptor interaction, thus balancing systemic stability with in vivo potency. Moreover, the polymer shell itself can be decorated with imaging agents or targeting ligands, giving rise to a theranostic platform with features of silent circulation, target recognition and biodistribution tracking.
In our work, lipidation has been a means of membrane poaching: attaching fatty acyl, steroidal or phospholipid groups that enable cell entry without activating the innate immune response. The lipid group is attached through a cleavable ester, carbamate or enzyme-cleavable peptide to enable the removal of the hydrophobic tag once the cargo has arrived at the intracellular site of action. The acyl chain length and saturation is matched to the cholesterol content and fluidity of the target membrane to ensure that the modified peptide will preferentially partition into lipid rafts enriched in uptake receptors. Bile-acid conjugates are used for oral or mucosal delivery to promote trans-epithelial transport, and medium-chain lipids are used to promote lymphatic uptake, making of the gut-associated lymphoid tissue a systemic starting point. The lipid tail can also be used as the anchor for self-assembling nanoparticles that co-encapsulate adjuvants or nucleic acids, enabling multi-modal immunostimulatory depots.
Chemical re-wiring of peptides, initially an engineering strategy only to prolong peptides' half-lives, has in the last ten years emerged as a general design principle that now threads through all strata of modern immunotherapy. Chemical changes that shift peptides away from the 'canonical' amino-acid geometry, such as backbone N-alkylation, side-chain stapling or substitution with mirror-image D-residues, can generate a 'molecular dialect' that is invisible to the cellular dialect (enzyme)-sensitive machinery, potentially prolonging its residence time without increasing off-target toxicity. These can be further decorated with lipid tails, glycan clusters, or nucleic-acid aptamers to form 'smart drugs' which can deliver antigens, danger signals, or metabolic co-factors to specifically target sub-anatomical locations. The result is a form of "programmable immunity" that goes well beyond the classical paradigm of vaccines: one can now encode peptides with lymph-node-targeting peptides to seed adaptive memory, fuse peptides with cytokine muteins to redirect exhausted T cells, or embed them in self-assembling nanofibrils to act as artificial germinal centres. But the grammar is also being applied outside of cancer to autoimmunity (tolerogenic peptide variants to silence autoreactive clones) and chronic infection (viral epitopes cloaked in unnatural stereochemistry to resist neutralizing antibodies while still eliciting sterilizing T-cell responses). In short, the field has moved from the question of whether peptides can be modified to how far the immune system itself can be re-programmed linguistically.
Fig.2 Overview of peptide-based immunomaterials.2,5
Cancer cells develop a dense array of biochemical and physical defenses against endogenous immune recognition as well as exogenous therapeutics. Engineered peptides now act as molecular machetes to cut their way through this jungle. Stapled or hydrocarbon stapled peptides can sequester tumor-suppressor p53 in its transcriptionally active conformation and reactivate apoptotic signaling in cancer cells that have 'forgotten how to die'. Lipidated neoantigen peptides can use albumin as a hitchhiking vehicle to distribute selectively in the tumor interstitium where they can be captured by dendritic cells and cross-presented within hours, instead of days. These peptides can be co-administered with metabolic checkpoint inhibitors (e.g., IDO-1 inhibitory peptides) such that the presentation of antigens happens at the same time as the inhibition of tryptophan catabolizing enzymes that would otherwise induce anergy in responding T cells. A photosensitizer can be added to the same backbone, and then activated with near-infrared light to convert a sub-therapeutic peptide dose into a wave of immunogenic cell death that turns the tumor bed into an in situ vaccine. In neuro-oncology, cell-penetrating peptides, when modified with glioma-homing sequences, can transport siRNA across the blood–brain barrier to silence oncogenic drivers while concurrently releasing GM-CSF and recruiting peripheral dendritic cells into the otherwise immunologically silent brain parenchyma. By converging these functions—targeting, signaling, and remodeling of the tumor microenvironment—into single macrocyclic entities, the conventional distinction between the drug and the delivery vehicle is now beginning to blur.
Challenge, Achilles heel of peptide-based vaccines has always been pharmacokinetic: soluble epitopes drain too rapidly from the injection site, are degraded by ubiquitous peptidases, and reach lymph nodes in quantities too small to ignite robust priming. Chemical grafting of vaccine peptides onto self-assembling β-sheet tapes or α-helical bundles produces nanoribbons that mimic the size and stiffness of viral particles, triggering avid uptake by subcapsular sinus macrophages and rapid transit to B-cell follicles. Further layering of pH-sensitive histidine residues allows these assemblies to swell in the mildly acidic environment of endosomes, bursting the vesicular membrane and releasing epitopes directly into the cytosol for cross-presentation. To ensure targeting specificity, the exterior of the nanostructure can be studded with integrin-binding or chemokine-receptor-targeting ligands that engage trafficking receptors on dendritic cells, guiding the entire payload from skin to draining node within minutes. Importantly, the same construct can embed adjuvant peptides—such as STING agonists—within its hydrophobic core, ensuring that innate activation occurs only after cell entry, thereby avoiding systemic cytokine storms. The net result is a molecular syringe that not only delivers its antigenic cargo intact but also programs the tempo and location of immune activation, transforming subunit vaccines into de facto viral mimics without the biosafety baggage of live vectors.
Acute immune bursts are easily achieved. The future is prolonging high-avidity T-cell clones for years on end. Molecular chronometers are now modified peptides that reset the half-life of immune memory by several complementary mechanisms. (i) Cyclization or head-to-tail lactamization protects the epitope from aminopeptidases so that depot formulations of antigen releasing nanomolar pulses for weeks to months are immunostimulatory. (ii) Fusion to albumin-binding domains prolongs serum half-life and also simultaneously delivers peptides to lymph nodes by trafficking through the subcapsular sinus albumin conduit, where antigenic signal is persistently dripped onto cognate T cells. (iii) Epitopes can be buried in protease-resistant, virus-like peptide cages that are not degraded in lysosomes and are periodically re-presented to the immune system by stromal macrophages, recapitulating an endogenous booster each time the cell divides. (iv) Metabolic signals such as conjugation to short-chain fatty acids bias responding T cells toward central memory phenotypes by engaging fatty-acid oxidation pathways that promote longevity at the expense of terminal differentiation. Taken together, these strategies move vaccination away from a single bolus of antigen and toward a renewable form of communication between antigen and immune system.
Natural peptides often face limitations such as rapid degradation and low stability, reducing their therapeutic potential. Modified peptides overcome these challenges by improving half-life, solubility, and bioavailability. Our modification services include PEGylation, cyclization, and lipidation, tailored to improve therapeutic outcomes. Modified peptides are widely used in cancer immunotherapy, vaccine development, and autoimmune research to ensure drugs reach their targets effectively and remain active longer. With validated modifications, researchers gain reliable, reproducible tools for advancing next-generation therapies.
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Modified peptides unlock new possibilities for drug delivery and immune activation. Collaborate with us to design peptide modifications that ensure maximum therapeutic success in your immunotherapy projects.
1. Why modify peptides?
To enhance stability, bioavailability, and drug efficacy.
2. What modifications are offered?
PEGylation, lipidation, cyclization, phosphorylation.
3. Are modifications safe?
Yes, they enhance performance while maintaining safety.
4. Do you modify custom peptides?
Yes, modifications can be applied to custom sequences.
5. How is quality assured?
All modified peptides are validated with HPLC/MS.
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