Fusion Proteins for Half-Life Extension of Biologics as a Strategy to Make Biobetters

BioDrugs (2015) 29:215–239 DOI 10.1007/s40259-015-0133-6 REVIEW ARTICLE Fusion Proteins for Half-Life Extension of Biologics as a Strategy to Make Biobetters William R. Strohl1 Published online: 16 July 2015  The Author(s) 2015. This article is published with open access at Springerlink.com Abstract The purpose of making a ‘‘biobetter’’ biologic is to improve on the salient characteristics of a known biologic for which there is, minimally, clinical proof of concept or, maximally, marketed product data. There already are several examples in which second-generation or biobetter biologics have been generated by improving the pharmacokinetic properties of an innovative drug, including Neulasta [a PEGylated, longer-half-life version of Neupogen (filgrastim)] and Aranesp [a longer-halflife version of Epogen (epoetin-a)]. This review describes the use of protein fusion technologies such as Fc fusion proteins, fusion to human serum albumin, fusion to carboxy-terminal peptide, and other polypeptide fusion approaches to make biobetter drugs with more desirable pharmacokinetic profiles. Key Points Biobetters are biologics based on an innovative biologic but with improved properties. Fusion proteins have been used in the biopharmaceutical industry for over 25 years to improve the pharmacokinetic properties of otherwise short-half-life biologics. Biobetter fusion proteins with longer half-lives or with targeting moieties are being developed for several innovative biologic drugs. 1 Introduction to Protein Pharmacokinetics and Elimination & William R. Strohl 1 Janssen BioTherapeutics, Janssen Research and Development, LLC, Pharmaceutical Companies of Johnson & Johnson, SH31-21757, 1400 Welsh and McKean Roads, PO Box 776, Spring House, PA 19477, USA There are now more than 180 therapeutic proteins and peptides approved by the US Food and Drug Administration (FDA) for a wide variety of indications, ranging from alleviation of neuropathic pain to rheumatoid arthritis and replacement enzymes for lysosomal storage diseases. Many of these proteins and peptides have less than optimal pharmacokinetic properties, often because they are smaller than the kidney filtration cutoff of around 70 kDa [1, 2] and/or are subject to metabolic turnover by peptidases, which significantly limits their in vivo half-life [3]. An example of this is the serum half-life of native glucagonlike peptide (GLP)-1, which is about 1–2 min, primarily because of peptidic cleavage by dipeptidyl peptidase (DPP)-4 [4, 5]. Moreover, for virtually all of these proteins and peptides, dosing is parenteral, so each dose is represented by either a subcutaneous or intravenous injection. 216 High dosing frequency, a small area under the curve (AUC), and patient inconvenience are limitations of shortacting peptides. Thus, in many cases, second- or thirdgeneration modifications of those protein or peptide drugs, intended to decrease their sensitivity to proteases [5] and glomerular filtration by the kidney [1, 2], have been developed to improve their pharmacokinetic profiles. Pharmacokinetics is often described as what the body does to the drug, whereas pharmacodynamics is described as what the drug does to the body. The pharmacokinetics of proteins and peptides is governed by the parameters of absorption, biodistribution, metabolism, and elimination. Absorption of peptides and proteins is generally via the lymphatic system [6], biodistribution is generally limited to the extracellular space in the central compartment (e.g., 3–8 L [5]), the volume of distribution is generally \15 L, metabolism occurs through enzymatic cleavage by proteases and peptidases [3–5], and proteins and peptides are eliminated from the serum by several different tissue- and receptor-mediated mechanisms. The most common routes of clearance for proteins and peptides include endocytosis and membrane transport-mediated clearance by liver hepatocytes for larger proteins, and glomerular filtration by the kidney for smaller proteins and peptides [1, 5]. While not all of the parameters involved in glomerular filtration of peptides and proteins are fully understood yet, it is clear that size, shape, hydrodynamic radius, and charge all play significant roles [1, 2]. Generally, proteins and peptides smaller than approximately 70 kDa are more likely to be eliminated by kidney filtration than are larger proteins [1, 2]. Additionally, charge plays a significant role in glomerular filtration. Negatively charged peptides or smaller proteins may be eliminated less readily than neutral polypeptides because of repulsion by the negatively charged basement membrane of the kidney [1, 7]. Cationic polypeptides, on the other hand, tend to be removed even more quickly [7]. Thus, two key strategies have been employed to improve the pharmacokinetics of smaller proteins and peptides, i.e., increasing the size and hydrodynamic radius of the protein or peptide, or increasing the negative charge of the target protein or peptide. A third strategy, similar to that employed with small molecules, is to increase the level of serum protein binding of the peptide or protein through binding to albumin [8, 9] or immunoglobulins [10]. Traditionally, the typical modification made in the past to improve the pharmacokinetics of peptide or biologic drugs was via conjugation to either linear or branchedchain monomethoxy poly-ethylene glycol (PEG), resulting in increases in the molecular mass and hydrodynamic radius, and a decrease in the rate of glomerular filtration by the kidney [1, 2, 11, 12]. PEG is a highly flexible, uncharged, mostly non-immunogenic, hydrophilic, non- W. R. Strohl biodegradable molecule, which generates a larger hydrodynamic radius than an equivalently sized protein [1, 2]. PEGylation has been used widely as a means to lengthen the half-life of proteins, e.g., PegIntron [PEGylated interferon (IFN)-a2b] and Pegasys (PEGylated IFN-a2a) for treatment of hepatitis B, Neulasta (a PEG-conjugated granulocyte colony-stimulating factor [G-CSF] for treatment of chemotherapy-induced neutropenia), and Mycera (a PEGylated form of epoetin-b). While PEG has been approved by the FDA as a GRAS (generally recognized as safe) molecule [13], it has been associated with vacuolization of renal cortical tubular epithelium cells [14], bringing its safety at least somewhat into question. Additionally, PEG is not metabolized by the body. Because of safety concerns—as well as the high cost of PEG itself and the need for chemical conjugation to the protein, followed by repurification of the conjugate [15]—more and more companies are seeking safer and less expensive alternatives to PEGylation. Another approach that has been utilized to improve pharmacokinetic parameters includes modification of glycosylation patterns, resulting in reduced clearance and extension of half-life. The best example of this approach is Aranesp (darbepoetin-a), a second-generation epoetin with modified glyco (...truncated)