The need for improved half-life extension technologies

In recent years, significant work has been dedicated to studying the half-life of drugs, and while researchers have been successful in lengthening the half-life of proteins and peptides, they have yet to find a way to tailor the pharmacokinetics of certain drugs to specific medical needs.”

Mark Perkins, Novozymes
Today's pharmaceutical market is facing a number ofpressures, in particular relating to moving drugs through clinical trials andto market faster and more cost-effectively. This stems from ever-shrinkingbudgets and growing financial burdens throughout the industry. In recent years,the industry has witnessed a significant increase in the investment into thedevelopment of targeted, biological drugs, with companies looking to take thesetailored, novel therapies to market faster than ever before. The increasedinterest into these therapies is as a result of their decreased side effectsfor patients, which makes them ideal for treating conditions where medicationneeds to be administered frequently, such as chronic diseases.
 
However, biological drugs are often hampered by theircharacteristically short half-lives, which means that once administered, theycan be cleared from the body in a matter of minutes. As a result of this shorthalf-life, patients with chronic conditions such as diabetes, hemophilia andneutropenia are often required to administer higher dosages more regularly,leading to likelihood of reduced compliance, higher costs and greater risks ofside effects. Drugs with a promising therapeutic value are often limited bythis factor. For this reason, the pharmaceutical and biotech sectors are payingincreasing attention to half-life extension strategies, with a number ofresearch institutes and academic papers noting the growing trend in developingtechnologies that extend and improve the circulatory half-life of peptides andproteins.
 
Responding to this issue, many researchers have concentratedtheir efforts in recent years into developing half-life extension technologiesthat modulate the serum half-life of protein-based therapeutics to desiredspecifications. However, while researchers have managed to successfully extendthe actual serum half-life, they have not been able to design flexible proteinhalf-lives to deliver the required pharmacokinetics.
 
Current strategies used for extending half-life are thosethat increase hydrodynamic volume (PEGylation) or those that use FcRn-mediatedrecycling (albumin fusions). Although real progress has been made in thecreation of novel technologies that modulate serum half-life, the market isstill actively searching for a solution that will allow companies to tailortheir therapies in line with specific medical indications.
 
Recent advances inalbumin fusion technology
 
Recent enabling half-life extension technologies are basedon serum albumin, a natural, non-immunogenic plasma carrier protein. Albumin isan optimal material upon which to base half-life extension technologies due toits naturally long half-life of 19 days in humans, in comparison to proteintherapeutics that are often cleared from the body in as little as hours. Apartfrom its size, it is the pH-dependent recycling through the neonatal FcRnreceptor that protects albumin from renal clearance and is responsible for itsextended half-life. Like IgGs, albumin is taken up by cells through nonspecificpinocytosis and is protected from intracellular degradation through pH-dependentbinding to the FcRn receptor in acidic endosomes. This interaction allowsalbumin to then be recycled back to the cell surface where it is released intocirculation due to the physiological pH of the blood.
 
It is the pH-dependent interaction between albumin fusion andthe FcRn receptor that provides the basis for the latest advancements inalbumin fusion technology. The understanding of its impact on albumin fusionhalf-life has enabled the engineering of this interaction with the potential tomodulate albumin's half-life. Previous studies that altered this interactionhave been shown to impact the pharmacokinetics of the IgG.
 
Applying these samekinetic principles, enabling half-life extension technologies are able tomodulate protein half-life through construction of albumin variants withaltered binding affinity to FcRn. With the ability to modulate albuminhalf-life, researchers are provided with the opportunity to tailor therapeuticsto specific disease states and fine-tune their drug design, holding significantbenefits for drug developers and patients alike.
 
Enabling technologies also produce more stable blood levelsin patients, and a reduced risk of side effects at the associated lower doserate means that the toxicity level of the protein may not be reached. Instead,these technologies allow the drug dose to remain within the therapeutic range,increasing the patient's tolerance to the drug.
 
Broad platformapplicability
 
The current approach when using albumin as a half-lifeextension technology is to conjugate, or genetically fuse. Both of thesemethods can be equally as effective, depending on specific drug deliveryrequirements. Lysine, tyrosine and the free thiol residues of the albuminmolecule are used for chemical conjugation to the drug product, with the freethiol at position 34 of albumin the most widely used conjugation route. Thisapproach is particularly useful for peptides containing maleimide groups thatspecifically react with the free thiol, allowing for the formation of a stablethioether bond between albumin and the peptide.
 
Alternatively, proteins can begenetically fused to the N- or C-terminus or even to both ends of the albuminvariant. Using a contiguous cDNA of the target protein or peptide with DNAencoding the albumin variant of choice allows the generation of protein fusionsexhibiting the required binding characteristics. A yeast expression systemprovides a high-quality, consistent and reliable supply of the protein ofinterest when a genetic fusion is applied. 
 
In recent studies, a range of albumin protein fusions hasbeen generated to test that the albumin variants maintain their modified FcRnbinding affinity when fused to a protein or peptide. The variants chosendisplayed a range of binding affinities from low affinity albumins (HSA K500A)to albumins with a 15-fold increase in receptor binding (HSA K573P). Antibodyfragments fused at the C-terminus, N-terminus or bivalent forms, as well asfusions to small or large peptides were compared to unfused albumin variantsfor FcRn affinity by SPR using Biocore technology. All albumin fusions testedpresented different receptor affinities, correlating to their unfused variants.Each showed the same differences in ScRn binding as the control rHSA variant.
 
As a result of the technology, proteins and peptides can bebound at either the C- or N-terminus or both. This creates fusion moleculeswith monovalent, bivalent or bispecific affinity. In addition to protein- orpeptide-based drugs, the technology also serves as a delivery vehicle for smallmolecules, providing a broad scope of usability. It also enables constructionof albumin variants with altered binding affinity to FcRn, making it possibleto modulate half-life extension of a fused target protein, while offering drugdevelopers enhanced flexibility and control.
 
Minutes to hours,hours to days
 
While the market demand for technologies that allow thedevelopment of drugs with novel properties continues to grow, researchers mustdevelop solutions that provide manufacturers with competitive solutions. Due tothe limited number of biological drugs available on the market, researchers arenow looking to adapt and improve upon those that are available to them. Inrecent years, significant work has been dedicated to studying the half-life ofdrugs, and while researchers have been successful in lengthening the half-lifeof proteins and peptides, they have yet to find a way to tailor thepharmacokinetics of certain drugs to specific medical needs. Enabling half-lifeextension technologies provide a solution to these issues by proving a platformin which drug developers are able to fine-tune the half-lives of drugs tocertain therapeutic conditions.
 
Commercially, the latest advancements in half-life extensiontechnologies offer drug manufacturers the opportunity to establish a nicheposition in the market with flexible products that offer improved performancethroughout the drug lifecycle. The ability to modulate and tailor thehalf-lives of drugs offers the potential to significantly improve quality oflife for patients with chronic conditions through lower and less frequentdosage levels. This can lead to increased patient compliance and thepossibility for patients to administer their own drugs. By realizing therelationship between albumin and its receptors for the first time, theseenabling technologies have the potential to revolutionize the wider healthcareindustry by increasing a protein's half-life from minutes to hours, and hoursto days.
 
Mark Perkins is the customer solution manager at NovozymesBiopharma and works with partners who are evaluating Novozymes Biopharma'srecombinant albumin products and associated technologies in the areas ofbiopharmaceutical formulation and half-life extension. He is a formulationchemist with a doctoral degree in pharmaceutical sciences from the University ofNottingham.

Mark Perkins, Novozymes

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