Half-life extension

Tackling the challenge of short half-life for biological therapeutics


With more than 180 therapeutic proteins and peptides approved by the US Food and Drug Administration (FDA), the market for biological therapeutics, also called biologics, is booming. But many small biologics, including VHH-based therapies, have one significant challenge that must be overcome: their short half-life.

The short half-life of many promising biologics is attributed to their small size of less than 70 kDa, which makes it easy for the kidney to filter them out. Because of this limitation, many novel small biologics like Blincyto (blinatumomab) – a first-in-class bi-specific antibody T-cell engager for haematological cancers – have a gruelling 28-day dosing schedule that requires in-person visits to the hospital/clinic so patients can continuously receive infusions.

At Isogenica, half-life extension for our VHHs was always top of our list. Improving half-life is essential to realise the potential of smaller biotherapeutics, like VHH-based drugs, by reducing dosing frequency and healthcare costs and, ultimately, improving the quality of life for patients.

In this article, we look at some ways in which the half-life of biological therapeutics can be extended to deliver on this promise, and why we chose an anti-albumin approach – ISOXTEND®.


Using half-life extenders to improve the short half-life of biological therapeutics


Over the years, several strategies have gained momentum in improving the half-life of biologics. While some of them improve the half-life by simply increasing the size of the therapeutic agent, others modify the structure of the drug to reduce the rate at which it is cleared. Many strategies also involve ‘tying’ the biologic to biomolecules that have a better half-life, such as antibodies, transferrin, or human serum albumin (HSA).

Here are some of the common techniques that can be used to increase the half-life of small biologics.




PEGylation was the first successful technology used to extend the half-life of biologics and has been applied to clinical medicine for more than 25 years. Through this process, polyethylene glycol (PEG) molecules are covalently attached to the biologic, increasing its molecular weight and effective size, which prevents the kidneys from quickly filtering the biologic based on size. Additionally, PEGs prevent access to proteases and peptidases that can degrade protein-based drugs, playing a further role in extending the half-life of these therapeutics.



Despite the efficacy of PEGylation, there are several disadvantages to this technology. For instance, research suggests that PEGs are not 100% safe – they are immunogenic and cells treated with PEGylated biologics can develop PEG-containing vacuoles. Additionally, because PEGs change the physicochemical properties of the biological therapeutic, they can also greatly reduce its potency.


Repeating amino acid chains


Repeating amino acid chains, which work in similar ways to PEG, can also be attached to biologics. Like PEG, they increase the effective size of the biological therapeutic to reduce direct filtration by the kidneys, improving the overall half-life of the biologic.

There are two leading amino acid half-life extension technologies – XTEN and ELPylation – although the use of other amino acid chains like PASylation and HAPylation is also being researched.

XTEN is a protein polymer developed by Amunix. It comprises a library of amino acid sequences expressed by E. coli that coil randomly around a small biological therapeutic. Depending on the length of the XTEN sequence used, the half-life of a biologic can be tuned to fit its clinical indication.

ELPylation, a technology developed by PhaseBio, also uses repeating amino acid chains to increase the size of biologics and thereby improve their half-life.

Unlike PEGs, these repeating amino acid technologies are biodegradable and non-toxic. But despite this, they have similar drawbacks to PEGs, including the potential for immunogenicity of the polypeptide repeat units. Additionally, these kinds of half-life extenders can raise a series of potential manufacturing and dosing challenges, including developability, stability, solubility, immunogenicity and loss of activity in comparison with the unmodified drug.


Protein fusion technologies


Small proteins in the blood are normally cleared out of the body by being broken down by cells in the liver or filtered out by the kidneys. But some proteins – such as human IgG antibodies and human serum albumin (HSA) – escape this fate by binding to cell surface receptors (FcRn) that enable them to be taken up by cells and ‘recycled’, prolonging their persistence in the body.

Fusing biologics with HSA or fragments from IgG is a commonly used technique for improving the half-life of many small biologics that would otherwise be lost by filtration or metabolism.


Fusing biologics with IgG to prolong half-life


Fusing a biological therapeutic with IgG enables it to bind to the FcRn receptor and be recycled through cells, bypassing degradation or filtration. Although this is an effective way to improve the half-life of biologics, Fc domains on IgG can cause toxicity in the liver and have unwanted side effects through interactions with immune cells. There are also limitations in applying this across preclinical species.


Utilising Human Serum Albumin (HSA) to extend the half-life of biological therapeutics


HSA is the most abundant plasma protein. It is highly soluble, extremely stable, and has a long circulatory half-life as a result of the FcRn-mediated recycling pathway.

Direct or indirect binding of biologics to Human Serum Albumin (HSA) also recruits the FcRn-mediated recycling pathway, protecting biologics from degradation by recycling them through cells and significantly extending half-life.

Early attempts to exploit the long half-life of HSA involved building fusion proteins combining biologics with serum albumin. However, HSA is a large protein and can exert significant effects on the biophysical properties of the drug, as well as its efficacy being limited to human studies.

However, novel approaches sidestep this issue by linking therapeutics with HSA in a way that preserves the structure and function of the drug.


Introducing ISOXTEND® technology to extend the half-life of biotherapeutics

Isogenica VHH Antibody Libraries

Originally derived from camelids, small-format single-domain VHH antibodies have binding capabilities similar to those of conventional monoclonal antibodies across a wide pH range, but are less immunogenic and can penetrate into smaller spaces than conventional IgG antibodies with Fc domains.

ISOXTEND® is a humanised VHH antibody that binds to albumin with nanomolar affinity maintained across a wide pH range. This enables the antibody to ‘piggyback’ through the endocytic recycling pathway, protecting it from clearance by the kidney while maintaining all other functional characteristics.

ISOXTEND® is in vivo validated and has multi-species cross-reactivity, making it suitable for use in preclinical models including mouse and non-human primates. Our studies have shown that ISOXTEND® extends the half-life of VHH-based therapeutics up to 26 hours in mice – the approximate equivalent of 16-19 days in humans.

The adaptability and flexibility of VHHs mean that ISOXTEND® can easily be chained together with other VHHs to create mono- or multi-specific biotherapeutics, or added to other biologics.

Half-life extenders like ISOXTEND® could make a big difference to the quality of life for patients being treated with cutting-edge biological therapeutics, increasing the time between doses, reducing the time they spend in hospital, and realising the potential of these next-generation treatments.


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