It is estimated that the total number of naturally occurring peptides that have been successfully identified is about 7000. These peptides play a crucial role in human physiology including acting as growth factors, neurotransmitters, anti-infective, ion channel ligands, and hormones. Generally, peptides are very selective and highly effective signaling molecules that have the ability to bind to specific cell surface receptors where they may trigger a variety of intracellular effects.

Due to their attractive pharmacological profiles as well as some of their very intrinsic properties, peptides are seen as a perfect starting point for designing a variety of novel therapeutics. Since they have proven to possess exceptional specificity, they also come with excellent safety, tolerability, and very high efficacy in humans. These features and attributes are the main factors that differentiate peptides from the bulk of most of the traditional small molecules.

Also, peptide therapeutics are mostly associated with low production complexities, unlike other protein-based biopharmaceuticals that naturally have high production complexities. As such, the production costs of peptides are relatively low, just like that of most small peptides, making them very attractive in the biopharmaceutical arena.

Peptides that occur naturally are not normally preferred for use in various therapeutics, owing to some of their certain intrinsic weaknesses such as poor physical and chemical stability, and short plasma half-life. Before they can be considered suitable candidates for the design and development of a variety of medicines, some of these weaknesses have to be addressed and proper mitigation measures put in place to circumvent them.

Though some of the intrinsic weaknesses have been successfully resolved, others are still a work in progress. Apart from the traditional peptide designs, a wide range of peptide technologies has been on the rise in recent years, and these technologies have provided a lifeline for the future development of peptide-based therapies. Some of these promising emerging technologies include the discovery and applications of cell-penetrating peptides, multifunctional peptides, and the development of peptide drug conjugates and technologies that focus on using alternative routes of administration.

In this piece, we focus on some of these emerging peptide areas and technologies and their relevance as far as the formulation of peptide therapies is concerned.

A Brief on Traditional Peptide Technologies

Over the years, peptide technologies have evolved, leading to the development of highly potent signal transduction molecules, with very powerful physiological effects. They are primarily characterized by their short circulating plasma half-life, and their suboptimal chemical and physical properties when being considered for application in the development of various types of medicines. As such, most of the traditional rational design of peptides has always evolved around techniques designed to mitigate some of the known weaknesses of peptides.

Rational Design of Peptide Therapeutics

In ration design of peptide therapeutics, the starting point for the therapeutic can be a known crystal structure that is then further developed into secondary and tertiary structures. Once this is accomplished, what then follows is that inputs from various analyses such as those obtained from alanine substitution and small focused libraries are designed in sequential steps which ultimately leads to the identification of essential amino acids as well as potential sites for possible substitution. During this process, especially during the process of liquid drug formation of the desired final product, it is vital to identify the amino acids that are chemically labile and prone to events such as oxidation, glycosylation, and isomerization – processes which should always be avoided.

Another vital aspect of the rational design of peptide drugs is the need to improve the physicochemical properties of the natural peptides. This is usually considered as a way of aggregating the peptides which are sometimes water-soluble. Some of the chemical design strategies that can be used to avoid aggregation may include corrupting hydrophobic patches. It is possible to achieve this through substitution or N-methylation of certain specific amino acids. If some of the peptide candidates are faced with solubility issues, then the point of focus would be on the charge distribution and the isoelectric point of the peptide, with regards to the pH of the desired formulation of the final product.

It is also possible to improve certain physiochemical properties of peptides by introducing stabilizing a-helixes or a salt bridge formation or other kinds of chemical modifications. Any modification introduced during the rational design of a peptide with the aim of improving the physicochemical properties of that peptide must be in tandem with the overall design of the desired pharmacological properties of the peptide therapeutic in question. With the current application of predictive IT software tools in the facilitation of such rational peptides, design has shown immense potential in leveling the playing field, and it is believed that it will lead to an increase of design prospects in the future, hence creating more room for the design of additional therapeutics.

Owing to the challenges represented by the intrinsic physicochemical properties of peptides, it has been possible to somehow get suboptimal solutions that have been incredibly useful to both physicians and patients. One good example worth considering is the use of glucagon used in treating severe hypoglycemia for patients suffering from diabetes. Presently, the glucagon rescue kit in the market for treating unconscious hypoglycemic patients features a lyophilized peptide powder in a sterile vial that can be quickly constituted and then administered to unconscious patients.

There is a general feeling that second-generation peptide drugs made through rational design may lead to more user-friendly peptide therapeutics. One prime example of this approach is the invention and the subsequent design of a glucagon analog that has displayed immense stability in liquid dosage form and is considered as an application that is ready for use in the form of a rescue pen.

Also, products of this nature may be useful when used as part of a closed-loop dual-chamber artificial pancreatic device for the delivery of both glucagon and insulin in therapeutics designed to be released centrally using an automatic pump in tandem with continuous blood glucose sensors. With the fast development, as well as the rapid miniaturization of devices and pumps, it may be possible for a pulsatile delivery system to be developed in the future, and this could be used for the smart delivery of peptides.

How to extend plasma half-life for peptides

Natural peptides generally come with a relatively short circulating plasma half-life. As such, there are various techniques that are currently under exploration in an attempt to increase the plasma half-life of the peptides. One of the approaches under serious consideration is to try and limit the enzymatic degradation of the peptide. This is achieved by identifying certain molecular cleavage sites where the relevant amino acid is substituted. It is possible to achieve the protection of certain enzymatic cleavages by enhancing the secondary structure of the peptide. With this approach, there will be a need for the insertion of a structure-inducing probe, a lactam bridge as well as clipping or stapling of the peptide sequence or through cyclization.

Most of the strategies which are currently under exploration use binding to the circulating protein albumin as the main driving factor for extending the plasma half-life which makes it possible for the peptide therapeutics to be administered less frequently. Some of these strategies include the insertion of albumin-binding peptide elements in the peptide backbone, peptide acylation, and peptide conjugation to albumin-binding antibody fragments.

Polyethylene glycol (PEG)-ylation may sometimes be used as a means of slowing down or limiting globular filtration, which in turn helps to increase the plasma half-life by simply limiting the elimination of peptides. Due to increasing concerns over safety and tolerability surrounding the use of PEG as one of the constituents for injectable therapies, however, PEGylation is no longer one of the strongly preferred methods.

Emerging Peptide Areas and Technologies

Currently, there is an immensely large pool of natural peptides, some of which have shown great potential as the perfect starting point for the development of various peptide therapeutics. The metabolic arena, such as the gut, for example, has attracted a lot of interest due to its diverse bacteria that could be helpful in identifying new peptides from sources such as degradation products, protein sources, and signaling molecules. With the ongoing microbiome research, there is hope that there will be new and significant opportunities for peptide therapeutics as solutions for a variety of metabolic diseases.

However, when it comes to innovative peptides for drug development, it is important for chemists and researchers to widen their scope and not focus on the traditional peptide technologies alone. One of the emerging technologies and approaches involves the use of multifunctional peptides which represent more than one pharmacological activity such as triple or dual agonism. This approach comes with a lot of sense based on the information from genomics. As such, it can be seen that knock-out animals often present with no distinct phenotype.

Additionally, although there are tremendous industrial efforts in the GPCR field that have made it possible to successfully identify several selective agonists and antagonists, only a few ligands have made it all the way to approval for use as medicines. All these tend to indicate some kind of redundancy in the biological systems and they also tend to favor multi-target approaches when it comes to the development of peptide medicines. Another method that involves the use of polypharmacology is the ability to use more personalized and individual treatments of differentiated patient groups.

Some of the multifunctional peptides that are currently in development include various antimicrobial peptide drug candidates that feature additional biological functions such as wound healing and the stimulation of the immune system. Additionally, there is a general trend gravitating towards multifunctional peptides, and this is evident in the fields such as GLP-1 agonists that have been used in the establishment of a drug class which has seen the introduction of various drugs in the market today, including Bydureon, Lyxumia, and Victoza. Since the introduction of these drugs into the market, they have grown in popularity, making them be commercially successful for developers as well as manufacturers.

One of the chemical strategies used in the development of multifunctional peptides may feature a hybrid of two peptides which are then bound together like modules. The binding can be directly or indirectly where a linker or a chimera may be used. Recently, GLP-1-GIP-GCG triple incretin agonist was exhaustively described as an anti-obesity agent following the strong beneficial effects it showed on model rats.

However, the development of multifunctional peptides could still present a challenge since the prediction of the in vivo outcomes of the drug candidates tends to be more complex when dealing with dual-target pharmacological compared to dealing with just a single target pharmacological. One of the challenges likely to be encountered with this is the translation of in vitro to in vivo where the potentially biased signaling may be encountered from novel ligands targeting two or more receptors.

Additionally, the process of translating results from animal models to human beings comes with many risks for multifunctional peptides when the same is compared to a single receptor peptide. The main reason for this challenge is the uncertainty associated with two or more targets. Within the antibody field, such challenges have also been encountered in the development of certain specific antibodies targeted for treating cancer. Due to these and other reasons, it is realistic to expect that established paradigms may be the ultimate source for the origins of multifunctional peptides.

References

1. C. Delgado, et al.
   The uses and properties of PEG-linked proteins
   Crit. Rev. Ther. Drug Carr. Syst., 9 (1992), pp. 249-304
2. R.R. Henry, et al.
   Continuous subcutaneous delivery of exenatide via ITCA 650 leads to sustained glycemic control and weight loss for 48 weeks in metformin-treated subjects with type 2 diabetes
   J. Diabetes Complicat., 28 (2014), pp. 393-398
3. M. Hattori, T.D. Taylor
   The human intestinal microbiome: a new frontier of human biology
   DNA Res., 16 (2009), pp. 1-12
4. Pocai, et al.
   Glucagon-like peptide 1/glucagon receptor dual agonism reverses obesity in mice
   Diabetes, 58 (2009), pp. 2258-2266
5. K. Fosgerau, et al.
   The new glucagon-GLP-1 dual agonist ZP2929 in combination with long-acting insulin improves glycemic control without causing weight loss in db/db mice
   Diabetes, 60 (2011), pp. A408-A442
6. B. Finan, et al.
   Unimolecular dual incretins maximize metabolic benefits in rodents, monkeys, and humans
   Sci. Transl. Med., 5 (2013), p. 209ra151
7. K. Fosgerau, et al.
   The novel GLP-1-gastrin dual agonist, ZP3022, increases beta-cell mass and prevents diabetes in db/db mice
   Diabetes Obes. Metab., 15 (2013), pp. 62-71
8. R. Just, et al.
   The novel GLP-1-GLP-2 dual agonist ZP-GG-72 increases intestinal growth and improves insulin sensitivity in DIO mice
   Diabetes, 63 (2014), pp. A1-A102
9. Pocai
   Unraveling oxyntomodulin, GLP1’s enigmatic brother
   J. Endocrinol., 215 (2012), pp. 335-346
10. V.A. Gault, et al.
    A novel glucagon-like peptide-1 (GLP-1)/glucagon hybrid peptide with triple-acting agonist activity at glucose-dependent insulinotropic polypeptide, GLP-1, and glucagon receptors and therapeutic potential in high fat-fed mice