The Benefits of Using Peptide Drug Delivery

Enhancing peptide stability and bioavailability through structural modifications

Introduction to structural modification

Peptide therapeutics, owing to their natural amino acid-based composition, exhibit inherent advantages. However, despite these advantages, peptides face substantial challenges, primarily related to their structural properties.184 Generally, peptides have two major limitations as therapeutics: poor membrane permeability and instability in vivo.2 Their size and amino acid composition hinder crossing cell membranes to reach intracellular targets.185 Moreover, the lack of secondary and tertiary structures makes peptides susceptible to enzymatic degradation. Additionally, their amide bonds are also prone to hydrolysis. These two factors result in short half-lives and rapid elimination in vivo for peptides.184 These limitations, coupled with the high manufacturing costs associated with peptide drug discovery, create hurdles in their widespread application. To overcome these limitations, structural modifications have emerged as a key strategy. A variety of modification methods (Fig. 7) have been widely applied to enhance stability against enzymatic cleavage, improve bioavailability by overcoming biological barriers, and fine-tune specificity for selective interactions with target molecules.184

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Structural modifications for stability and improved bioavailability

Overcoming the instability against proteolysis and limited bioavailability associated with peptide therapeutics are among the most pressing issues in the field of structural modifications. Various structural modification methods have been applied, and here we provide a brief overview of these approaches and their applications.

A) Backbone modifications

Backbone modifications are the earliest structural modification method in the field that targets the masking or removal of amide bonds in the peptide backbone. When it comes to this approach, the most commonly employed tactics include substituting D-amino acids for L-amino acids,186,187 inserting N-alkylated amino acids,188 and incorporating β-amino acids or α/β-substituted α‑amino acids.189,190 Each strategy is implemented through different principles to address proteolytic stability issues that often plague native peptides.191 Substituting metabolically labile L-amino acids with their D-amino acid counterparts enhances resistance to enzymatic degradation, significantly extending the peptide’s half-life.186 The insertion of N-alkylated amino acids and incorporation of β-amino acids or α/β-substituted α‑amino acids strategically fortify the peptide structure, leading to improved stability.188 In the process of synthesizing lanthipeptides, the backbone modifications are elegantly presented, for instance, d-amino acids are successful introduced into lanthipeptides via two enzymatic reactions catalyzed by the dehydratase domain of lanthipeptide synthase,192 showcasing the potency of backbone modifications in overcoming challenges associated with proteolytic instability in peptide therapeutics.

Replacing one or more peptide bonds with an isosteric or isoelectronic substitute are also strategies that modify the peptide’s backbones. The isosters that are most frequently used include azapeptides, retro-inverso peptides, and peptoids,193,194,195 bringing forth diverse modifications and offering unique functionalities to peptide drugs. Azapeptides have a similar structure to natural peptides, but with the key difference of having a nitrogen atom rather than a carbon atom bonded to the amino group, which makes azapeptides useful synthetic mimics of natural peptides. For instance, a powerful new covalent inhibitor of the SARS-CoV-2 main protease (Mpro) using an azapeptide scaffold capped with a cysteine residue was reported recently, enabling targeted, irreversible labeling of the Mpro active site and makes it one of the most potent Mpro inhibitors reported so far.196 Retro-inverso peptides, with reversed N- and C-termini sequences and L- to D-amino acid substitutions, find applications as versatile immunomodulators, anti-inflammatory agents, and more.197,198,199 Peptoids, featuring N-alkylated glycines, provide flexibility and have applications in cancer, neurological and autoimmune disorders.200

B) Side chain modifications

Side chain modifications are another strategic maneuver in the enhancement of stability and bioavailability of peptide drugs. The principles underlying this approach involve the replacement of natural amino acids with their analogues during the synthesis of peptides. This substitution seeks to bring about advantages such as the augmentation of binding affinity and target selectivity.201 Notable applications of side chain modifications include the incorporation of analogues like β-phenylalanine,202 benzyloxytyrosine,184 and homoarginine.203 These modifications have found practical implementation in near-infrared (NIR) dye-peptide conjugates, for instance, it has been reported that the use of acetamidomethylcysteine to replace the cysteine residue in a near-infrared fluorescent dye conjugated with the type I collagen targeting peptide RRANAALKAGELYKCILY successfully disrupted the self-assembly of the peptide and thereby changed the performance of the molecular probe in aqueous solution, ultimately improving the contrast of arthritic joints against the background.204

C) Peptide cyclization

Among the six peptide drugs approved in , three are cyclic peptide drugs (Rezafungin, Motixafortide and Zilucoplan), emphasizing that cyclic peptides have become an important modality in the development of peptide drugs.205,206,207 Delving into the peptide cyclization unfolds diverse approaches, most commonly used including head-to-tail, backbone-to-side chain, and side chain-to-side chain.208 The advantages stemming from peptide cyclization encompass heightened proteolytic stability,209,210 and the facilitation of secondary structure formation.211,212 These principles have been applied tangibly in the development of S-tert-butylation of the free thiol group of cysteine. Which was done in a rationally designed peptide using the evolutionarily conserved γ-core region (GXC-X3-9-C) of the antifungal protein from Aspergillus.213 S-tert-butylation not only improved stability, but also extended the peptide’s antifungal activity to the mold Aspergillus fumigatus.

A significant advantage of cyclic peptides lies in their pharmacokinetic properties. Specifically, their unique characteristics in stability, hydrophilic/lipophilic balance, cell permeability, etc. make cyclic peptides a key focus for the development of orally bioavailable peptide drugs.214 Among the currently marketed oral peptides, three are cyclic peptides - Cyclosporine A, Voclosporin, and Desmopressin. The favorable pharmacokinetics conferred by their cyclic structure contributes to their success as orally bioavailable peptide therapeutics.215,216,217 In addition to these marketed drugs, some recently reported cyclic peptide drugs also show potential. For instance, MK- is an orally bioavailable, renally excreted cyclic peptide inhibitor of PCSK9. In clinical trials, MK- has demonstrated dose-dependent reductions in LDL cholesterol, non-HDL cholesterol, and apolipoprotein B levels. Besides, it can lower Lp(a). MK- is currently in Phase 3 clinical trials and shows promise for the treatment of cardiovascular diseases given its oral bioavailability, potent lipid-lowering effects, and renal clearance.218 Meanwhile, stapled peptides, which are also cyclic peptides in nature, skillfully “stapled” into α-helix and β-sheets shapes, exhibit proteolytic resistance and extended plasma half-life,219 exemplified by stapled peptides that resemble Helix 1 of the human ACE2 receptor, have shown varying degrees of efficacy in preventing SARS-CoV-2 infection.220 In addition, cyclized analogues adopted type I β-turn structures achieved by substituting select glycine residues with N-(2-thioethyl) glycine and stapling the peptides using bifunctional reagents was reported. Which yielded cyclic analogues with improved analgesic activity compared to the parent enkephalins after introducing benzyl substituents on the trithiocyanurate stapling reagents.221 Besides, stapling natural peptides by cross-linking two amino groups via different imidazolium linkers with various α-ketoaldehyde reagents has been reported recently,222 providing more possibilities for the design of stapled peptides.

Another frequent use of peptide cyclization is to stabilize various secondary structures like α-helixes and β-sheets, known as “mimicking secondary structures.”2,223 Mimicking secondary structures employing strategies like cross-linking or hydrogen bond surrogates to replicate α-helices and β-sheets.2 Key approaches for mimicking and stabilizing α-helices in peptides include lactam-based crosslinks, disulfide bonds, and biselectrophilic linkers. Lactam-based crosslinks form a lactam bridge by substituting lysine for the side chain of glutamic acid or aspartic acid, enhancing hydrogen bond formation and restricts conformational freedom of the peptide chain to mimic and stabilize alpha helical conformations.224 Disulfide bonds can form covalent connections between peptide chains when N-terminal serine residues are replaced with homocysteine or cysteine.225 Biselectrophilic linkers can react with two amino acid side chains on the peptide chains simultaneously to generate connections.226,227 To stabilize β-sheets, peptides are modified by introducing D-amino acids, a successful example is the recently reported use of mirror-image phage display technology to select D-amino acid peptide ligands for aggregation-prone proteins that play a role in neurodegenerative diseases, thereby avoiding the accumulation of endogenous proteins such as amyloid beta peptide (Aβ), achieving therapeutic purposes.228 As for β-sheet mimicking, there are also reports that have created β-sheet mimics via macrocyclization or amyloid.229,230 In conclusion, mimicking secondary structures enhances stability and specificity, enabling a dynamic shift in peptide therapeutic design, allows creating structures that closely mimic protein-protein interfaces, demonstrating the potential of secondary structure mimicry.

D) Conjugation strategies

Conjugating peptides to larger molecules are also a method that has become quite popular in recent years, which not only increase lipophilicity, but the enhanced steric hindrance also prevents filtration of the conjugate through the kidneys and prolongs circulation time. Units commonly used to conjugate include lipids and polymers. Lipid conjugation strategy entails tying peptides to lipids such as glycerides, steroids, and fatty acids.231,232 The principles behind this approach include forming stable ester or amide bonds, with fatty acids such as squalenoic acid and docosahexaenoic acid playing prominent roles.233,234 The advantages offered by lipid conjugation include reduced toxicity and improved bioavailability.235 An illustrative example involves the conjugation of the Poly (L-glutamic acid) (PGA), as the hydrophilic backbone, with the peptide antigen Ag and the molecular adjuvant imidazoquinoline (IMDQ) TLR7/8 agonist, respectively. The polyanionic properties of PGA were used to bind the electrostatic interaction of ionizable lipids condenses PGA-Ag and PGA-IMDQ into lipid nanoparticles.236

Conjugation with polymers is another useful tactic to increase the stability, prolong the in vivo half-life, and lessen the immunogenicity of peptides, with PEGylation being a widely used technique.237 This approach operates on the principles of extending half-life and mitigating immunogenicity, providing a shield against enzymatic degradation.237 For instance, FDA-approved PEGylated proteins Krystexxa and PEGASYS have elevated the standard for stability and in vivo efficacy of peptide therapeutics.238 Despite these gains, it has also been suggested that whereas conjugation shields targeting peptides from proteolysis to some extent, altering the peptide sequence to increase protease resistance can significantly increase homing and transport efficiency.239

E) Terminal modifications

As exopeptidases typically break down peptide sequences at either the N- or C-terminus, it is possible to increase a peptide’s resistance to protease hydrolysis by modifying its terminus. N-acetylation, C-amidation, and fusion with albumin are the main concepts when it comes to terminal alterations and fusion tactics.188,240,241,242 These modifications orchestrate an extended plasma half-life, amplifying the potential of peptide-based therapeutics. Exemplifying this, with site-specific albumin conjugation, a “clickable” non-natural amino acid named azide-l-phenylalanine (AzF) was added to three specific sites (V16, Y19, and F28) of the GLP-1 variant and then linked to HSA through a strain-promoted azide-alkyne cyclization reaction. The resulting three HSA-conjugated GLP-1 variants (GLP1_16HSA, GLP1_19HSA, and GLP1_28HSA) have serum half-lives comparable to HSA in vivo.243

Conclusion and future directions

The future of structural modification of peptide drugs is poised to witness several exciting advancements. First, precision targeting is gaining prominence, with researchers designing peptides that exhibit high specificity for their intended targets, enabling precision targeting of disease-related proteins or receptors.98 Additionally, we’re witnessing the emergence of multi-functional peptides that combine therapeutic effects with diagnostic capabilities or even serve as drug delivery vehicles.244 Another promising trend involves stapled peptides, which stabilize peptide structures through covalent bonds, are expected to see further advancements to enhance peptide stability and bioavailability.184,219 Besides, the exploration of novel peptidomimetics, synthetic molecules that mimic the properties of peptides, may help overcome limitations associated with natural peptides.245,246

However, the field faces several research challenges that require innovative strategies to address. Improving the oral bioavailability of peptides, which often struggle with enzymatic degradation and poor absorption, remains a critical priority.247 Prolonging the typically short half-lives of peptides without compromising their efficacy is another key challenge.248 Overcoming immunogenicity, where some peptides can trigger unwanted immune responses, is also crucial.195 As researchers explore more intricate peptide structures, the complexities of synthesis and characterization present additional hurdles.184,249

To address these challenges, various technologies and strategies are being explored. Backbone modifications alter the peptide backbone to enhance stability and resistance to enzymatic degradation. Side chain modifications, achieved by substituting specific amino acids with modified analogs, can improve properties like solubility and binding affinity, might find applications in tailored therapeutics for diverse diseases. Conjugation with polymers extends half-life and improves pharmacokinetics, may see refinements for specific drug delivery needs. Cyclization, creating cyclic peptides, enhances stability and reduces susceptibility to proteases. Terminal modifications and fusion strategies are likely to be refined, offering extended half-lives for peptide therapeutics. Lastly, novel delivery systems, such as nanoparticles or liposomes, enhance tissue penetration and overall bioavailability. By leveraging these innovative strategies, the future of peptide drug development holds great promise in precision, multifunctionality, and overcoming existing limitations.250

Advance of peptide-based therapeutics in diabetes

Obesity is a chronic progressive disease that affects nearly 760 million adults worldwide. Obesity not only affects human health and quality of life, but also increases the risk of T2DM, and cardiovascular disease. Diabetes is a chronic metabolic disease that is prevalent globally, with 537 million people already living with diabetes as of , and T2DM accounts for approximately 90% of all people with diabetes, a number that is expected to continue to increase in the coming decades.251 Diabetes imposes a huge burden on the health of individuals and society. In addition to obesity, complications such as diabetic cardiomyopathy, diabetic nephropathy, and diabetic foot can occur, increasing the rate of disability and death.252

T2DM is primarily caused by insulin resistance and insufficient insulin secretion, and usually develops in adults, especially middle-aged and older adults. Weight management can be used to control the onset and progression of the disease in both T2DM and obese patients. Weight management is a lifestyle-based treatment that combines a personalized low-calorie diet, physical activity, and behavioral counseling. This intervention is able to reduce patient weight moderately (5%-10%) and control the onset of T2DM, but further weight loss is needed for effective control and most patients treated with weight management have difficulty adhering to it.253 Although drugs such as exenatide and simethicone are now marketed for the treatment of diabetes, they are not yet sufficient, so the development of novel drugs remains important.254,255

History of peptide-based therapeutics in diabetes

Insulin, the prototypical pharmaceutical agent for the treatment of diabetes, was initially derived and purified from the pancreatic tissue of a canine in and subsequently validated in a 12-year-old adolescent in . In , Eli Lilly and Company’s insulin (Insulin®) was launched on the market; in the same year, the Nobel Prize in Medicine and Physiology was awarded to the discoverers of insulin, such as Banting and MacLeod. However, the initial production method was not only complicated, costly and low-yielding, but also susceptible to immune reactions and viral infections.256 With the advent of genetic engineering, researchers recombinantly obtained insulin in E. coli. Unlike extracted animal insulin, this type of insulin is attributed to human proteins, and the immune response is greatly controlled.257 In , Insulin human® was approved by the FDA as the first recombinant therapeutic protein.258 With the development of genetic and chemical engineering, the number of engineered insulins gradually increased and insulin modifications followed. saw the approval of Lispro® (Humalog) as the first industrially engineered insulin. Detemir (Levemir®), which is a long-acting insulin, adds a fatty acid chain to the carboxyl terminus of the insulin β-chain.259 Monnier et al. successfully prolonged the duration of insulin’s action by acylating insulin at LysB29, which allowed this peptide hormone to bind with endogenous serum proteins. In recent years, researchers have partially electrostatically mediated under the skin by introducing mutations at the C-terminal end of the β-chain leading to the aggregation of mutant insulins, allowing them to be gradually secreted into the bloodstream.260 Many new insulin analogs have also been developed, such as the long-acting insulin analog Insulin glargine, which has a duration of action of up to 30 h, and the rapid-acting insulin analog insulin aspart, which has an immediate immediate onset of action after 10–15 min and peaks after 1–2 h (Table 6).261

In , Charles Kimball and John Murlin found and identified glucagon. In the s, Joel Habener262 described a new glucagon-related peptide encoded in the pre-glucagon cDNA of the pipefish. In the s, Joel Habener described a new glucagon related peptide encoded in the pipefish pre-glucagon cDNA. Two glucagon related peptides were subsequently identified in rat, bovine, hamster and human glucagonogen. These two peptides are now referred to as GLP-1 and glucagon-like peptide-2 (GLP-2). GLP-2 is an enterotrophic hormone released by enteroendocrine cell (EEC) L cells,263 and its receptor is mainly distributed in the digestive tract, exerting enteroprotective effects through different pathways GLP-2 acutely prevents endotoxin-related increased intestinal paracellular permeability in rats. Currently, the field of diabetes is dominated by the study of GLP-1. GLP-1 has a wide range of pharmacological and therapeutic uses. In addition to its ability to reduce gastric emptying, inhibit food intake, or control metabolism, the activation of the GLP-1 receptor induces a protective effect, the regulation of the hypothalamic-pituitary-adrenal axis, heart and lungs. It also reduces the production of inflammatory cytokines, chemokines, and the infiltration of immune cells in tissues, resulting in a broad range of neuroprotective and anti-inflammatory effects. GLP-1 interacts with its receptors to stimulate insulin secretion from pancreatic β-cells and inhibit glucagon release from pancreatic α-cells, increasing satiety and delaying gastric emptying in a glucose-dependent manner. Endogenous GLP-1 is degraded and rapidly inactivated by dipeptidyl peptidase-4 (DPP-4). To prolong the stimulation of GLP-1 receptors, GLP-1 RAs need to be synthesized to prevent their degradation.

GLP-1 RAs are emerging drugs for glycemic control and have been widely used in the treatment of T2DM in recent years. Currently, GLP-1 RAs are mainly categorized into peptide and non-peptide. Based on the similarity of their amino acid sequences, peptide agonists are mainly categorized into GLP-1 and derivatives and exendin-4 and derivatives.252 In recent years, with the growing market size of the GLP-1 RA class of drugs, the U.S. glucose-lowering drug market has undergone a trend shift, evolving from insulin-based drugs as the star drugs before to GLP-1 drugs leading the way.

Since the FDA approved the first GLP-1 RA, Exenatide (Eli Lilly, Exenatide®), in , six GLP-1 RAs have entered the clinic, including liraglutide (//, Novo Nordisk A/S, Victoza®/Saxenda®/Xultophy®), lixisenatide (, Sanofi-Aventis, Lyxumia®/Adlyxin®/Soliqua®), albiglutide (, GSK Plc, Eperzan®/Tanzeum®), dulaglutide (, Eli Lilly, Trulicity®), semaglutide (, Novo Nordisk A/S, Ozempic®/Rybelsus®) and tirzepatide (/, Eli Lilly, Mounjaro®/Zepbound®), in addition to efpeglenatide and taspoglutide which are in clinical studies. When injected, these GLP-1 RAs help the pancreas release the right amount of insulin when blood glucose is high, effectively lowering glycated hemoglobin and average blood glucose levels and improving fasting glucose.

As the first GLP-1 RA that can be used orally, semaglutide is able to effectively control blood glucose levels and achieve appetite reduction and weight loss by slowing down gastric emptying through brain regions that regulate appetite and food intake.264 For patients with T2DM, the use of semaglutide has multiple implications. First, as a hypoglycemic drug, it can help patients effectively control their blood sugar levels and reduce the risk of stroke, heart attack, or death in patients with T2DM, as well as in patients with cardiovascular disease, and it is suitable for use in overweight/obese patients suffering from hyperglycemia, as well as, with a proper diet and exercise program. Secondly, semaglutide also has cardiovascular protective effects,56 which is certainly an important therapeutic option for diabetic patients with comorbid cardiovascular disease or at high risk of cardiovascular disease. In addition, Novo Nordisk has made three modifications to semaglutide: 1. alpha-aminoisobutyric acid has been used to replace alanine at position 8. 2. a C18 lipoic acid side chain has been linked to lysine at position 26, with glutamic acid as the linker. 3. arginine has been used to replace lysine at position 34. The above three modifications enable the ultra-long-lasting characteristics of semaglutide, which needs to be taken orally only once a week, which greatly improves the convenience of dosing and patient compliance.

In addition to the hypoglycemic effect, the weight loss effect of semaglutide has also attracted much attention. By slowing down gastric emptying, increasing satiety and acting on the hypothalamus to suppress appetite, semaglutide is able to effectively reduce dietary intake, thus achieving weight loss. Studies have shown that semaglutide can reduce body weight by 15 to 20%, which is far more effective than previously approved weight loss medications. For obese type 2 diabetic patients, semaglutide can not only help them control blood glucose, but also effectively reduce weight, which is of great significance to improve the metabolic status of diabetic patients.

Efpeglenatide, currently in development, is a long-acting GLP-1RA used to control blood glucose levels in patients with T2DM. Efpeglenatide consists of a modified exendin molecule and is coupled to a fragment of human immunoglobulin 4 by a special technology called long-acting peptide. This special coupling technology allows for a more flexible dosing frequency of efpeglenatide.265 Epernatide is also less desensitizing than other therapeutic agents, which means that epernatide has a longer-lasting therapeutic effect. However, eperinatide is still in the developmental stage, and its other related properties and characterization need to be developed in subsequent studies.

Similar to GLP-1, GIP is an intestinally secreted peptide that promotes insulin biosynthesis and islet β-cell differentiation and reduces apoptosis. Clinical trials have shown that the GIP/GLP-1 dual-targeting combination has shown effective weight loss and glucose-lowering effects. Based on these two important targets, Lilly developed tirzepatide (Zepbound®), the first marketed dual-target agonist (GIP/GLP-1) obesity treatment injection, which provides better weight control than semaglutide and dulaglutide (Fig. 8b).266 The drug has previously been approved as an adjunctive therapy to diet and exercise to improve glycemic control in T2DM.267,268 Tirzepatide consists of 39 amino acids, 37 of which are naturally occurring (or coded) and two are non-naturally occurring. Non-coding amino isobutyric acid residues at positions 2 and 13. Besides, it is amidated at the C-terminus, which binds the C20 lipoic acid portion through a spacer region attached to Lys20.269 This side-chain structure has been cleverly designed to not only improve the stability and bioavailability of the molecule, but also to enhance its affinity for the target. For dual activity, it incorporates amino acid residues primarily from GLP-1 and GIP and uses some unique amino-acid residues. Studies have shown that tirzepatide improves pancreatic β-cell function and insulin sensitivity, and compared to traditional GLP-1 RA, these drugs have a longer duration of action, allowing for a reduction in the frequency of weekly dosing.270

Meanwhile, the research and development of triple-target agonists is also a current hot issue. It has been found that retatrutide, a 39 amino acid peptide, can resist cleavage by dipeptidyl peptidase IV, which is responsible for breaking down GLP-1 and GIP,271 and it also stimulates GLP-1, GIP and glucagon receptors, which is potentially useful for treating obesity and T2DM. However, studies have shown that retatrutide can increase heart rate by 6.7 beats per minute,272 which may be harmful and counteract some of the benefits of weight loss. Retatrutide is currently in the developmental testing phase and more detailed data is pending subsequent studies, but the availability of retatrutide still offers a new direction in the treatment of T2DM.

In addition, there are some potential drugs that may also have an effective effect on diabetes. C-peptide, as a bioactive peptide, can reflect the indicators of pancreatic β-cell function as well as bind to signaling molecules on the surface of the cell membrane and activate its own signaling pathway, exerting antioxidant, anti-apoptotic, and anti-inflammatory effects, or regulating cellular transcription through internalization. It has been found that the bioactivity of C-peptide can be used to prevent and treat the complications of diabetes and thus influence the comprehensive treatment of T2DM. Currently, the relationship between C-peptide and chronic complications of diabetes is complex and clinical trials have been unsatisfactory, requiring control of baseline C-peptide levels. Efpeglenatide, currently in development, is a long-acting GLP-1 RA for controlling blood glucose levels in patients with T2DM.273 Efpeglenatide consists of a modified exendin molecule and is coupled to a fragment of human immunoglobulin 4 by a special technology called long-acting peptide This special coupling technology allows for a more flexible dosing frequency of efpeglenatide. At the same time, eptifibatide is less desensitizing than other therapeutic agents, which means that eptifibatide has a longer-lasting therapeutic effect.

Progress of GLP-1 receptor agonist repositioning studies

In recent years, GLP-1 has made many important advances in a number of research areas, including the cardiovascular system, central nervous system, obesity and metabolic syndrome, insulin secretion, muscle and liver (Fig. 9). Studies have found that GLP-1 RAs have multiple beneficial effects in the cardiovascular system, including improving cardiovascular function and inhibiting the development and rupture of atherosclerotic plaques, and thus GLP-1 RAs may have cardiovascular protective potential, slightly reducing the risk of death due to cardiovascular disease and any cause, and slightly reducing the risk of stroke compared to placebo. Recent studies have shown274 that GLP-1 may also regulate appetite and energy balance by affecting neuronal activity and synaptic transmission, which is critical for appetite and weight regulation. In addition, GLP-1 plays an important role in the pathogenesis of obesity and metabolic syndrome. It has been found that obese patients often have GLP-1 resistance, resulting in reduced GLP-1 bioactivity. Therefore, researchers are exploring ways to increase GLP-1 activity to improve the treatment of obesity and metabolic syndrome. Some scholars’ research shows that Liraglutide can reduce the occurrence of neuroinflammation to a certain extent while reducing Aβ plaques, and the above two are important causes of Alzheimer’s disease (AD), so GLP-1 RAs have great potential in the treatment of AD.275 Because Liraglutide has certain anti-inflammatory effect, can reduce the occurrence of neuroinflammation, some scholars believe that it will also play an irreplaceable role in the treatment of Parkinson’s disease. Some experimental studies have shown that Liraglutide can also activate SIRT1 to a certain extent, which is an important factor in regulating muscle cell metabolism, and can prevent muscle atrophy, so GLP-1 RAs provides new ideas for the treatment of muscular dystrophy and other diseases. In addition, Liraglutide has been verified to reduce the possibility of hepatic steatosis by participating in autophagy-lysosomes. The use of semaglutide prior to total hip arthroplasty has been shown to reduce postoperative prosthetic joint infections and readmissions. All of the above suggests that there is great potential for GLP-1 RAs in the treatment of a wide range of conditions.

Advance of peptide-based therapeutics in cancer

Cancer is one of the primary factors affecting human health and leading to mortality. According to World Health Organization statistics, nearly 10 million deaths (almost one in six) are caused by cancer in .276 Despite the breakthroughs in immunotherapy in recent years, the current clinical treatment of tumors is still dominated by chemotherapy, surgery and radiotherap. In this regard, surgical procedures are prone to trauma and bleeding, as well as posing risks of infection and weakened immunity.277 Radiotherapy is prone to a variety of complications, as well as high costs and long treatment times.278 Chemotherapy has significant side effects and fails to differentiate between normal and tumor cell. In addition, chemotherapy often leads to the development of drug resistance and is prone to relapse279; Although immunotherapy has significantly fewer side effects than chemotherapy,280,281 the effectiveness of treatment varies with each individual, and it also has the potential to trigger autoimmune myocarditis282 or induce cytokine storms.283 Therefore, new treatments or anti-tumor drugs are urgently needed to be researched and developed to meet the various needs. Anti-cancer peptides (ACPs) have gradually received attention from researchers in the field of tumor therapy because of their high specificity and safety advantages.

ACPs are a class of anti-tumor active peptides within antimicrobial peptides (AMPs), typically possessing a positive charge, high hydrophobicity, and strong penetrability.284 Then, it can be used as a hormone and inhibitor or as a peptide vaccine to activate anti-tumor immune responses.285,286 Besides, these peptides can be used for imaging, cancer diagnosis, and targeted drug delivery, even some specific peptides can be engineered to attack cancer cells and prevent tumors from worsening.287 Based on the types, quantities, and structures of amino acids, ACPs can be classified into four categories: α-helical, β-pleated sheets, random coil and loop structures.288 From the perspective of source, it mainly includes natural peptides, peptides obtained from combinatorial libraries (more details in section “Display library technology”), as well as synthetic or modified peptides.284 It has complex mechanisms of action, including inhibiting tumor angiogenesis, disrupting cell membranes, interfering with metabolism, targeting cytoplasmic components and mediating immune cell regulation, etc. An overview of the research on peptides in the direction of anti-tumor is presented in Fig. 10.

Peptides for anti-cancer

A) Natural anti-cancer peptides

Natural ACPs are widely found in animals, plants and microorganisms. For example, oncolytic peptides (e.g., anoplin) can selectively cleave tumor cell membranes and remain effective against drug-resistant tumors.289 Some studies have also proved that oncolytic peptides have potential role in activating anti-tumor immunity. The first drug named “oncolytic peptide”, LTX-315 (Ruxotemitide, Oncopore), is currently in phase II clinical trials and can induce complete ablation of a variety of tumors.290,291 Wang KR et al. isolated a cationic ACPs, Polybia-MPI, which was originally isolated from the venom of the social wasp Polybia paulista. Its primary sequence is IDWKKLLDAAKQIL-NH2.292,293 It targets non-polar lipid cell membranes, forming ion-permeable channels and leading to depolarization, irreversible cytolysis, and eventual cell death. The results showed that polybia-MPI exhibit antitumor activity by disrupting the cell membrane of cells, while it has lower cytotoxicity to erythrocyte and normal fibroblast.292 It is also unaffected by conventional multi-drug resistant mechanisms and has the potential to be used as a chemotherapeutic agent against multi-drug resistant tumors.294 In addition, anti-tumor peptides, such as Apicidin(cyclo(N-O-methyl-L-tryptophanyl-isoleucinyl-D-pipecolinyl-L-2-amino-8-oxodecanoyl)), can be obtained from fungal metabolites. It is a selective inhibitor of histone deacetylase (HDAC) and has been shown to have potent anti-angiogenic activity and reduce the level of hypoxia inducible factor-1α (HIF-1α) in human and mouse tumor cell lines.295,296 In the HCT-116 xenograft tumor model of human colon cancer, Apicidin inhibited tumor growth. Moreover, apicidin also showed anti-tumor activity in Ishikawa cell xenograft tumor model, which could inhibit the proliferation of eight tumor cell lines, including HeLa, MCF-7 and HBL-100.297

B) Synthesized or modified anti-cancer peptides

Natural peptides as potential drug candidates have the advantages of high selectivity, biocompatibility, diversity of targets of action, and low toxicity, however, natural peptides also have some disadvantages. The instability of some natural peptides can lead to their susceptibility to degradation by gastric acids and enzymes, which reduces their effectiveness in oral drug delivery. In addition, peptides derived from natural sources usually have high production costs, limiting their large-scale application. In order to overcome the limitations of natural peptides, the development of peptide synthesis and modification techniques has become crucial. Synthetic technologies can produce peptides with specific sequences and structures, improving their stability and biological activity. Additionally, modification techniques can change the physical and chemical properties of peptides and enhance their drug compatibility, bioavailability and targetability.298 The development of peptide synthesis and modification technologies can further expand the application of peptides as drugs, improve their therapeutic efficacy and reduce their side effects. Here we described typical and some new synthetic peptides. For more details on peptide modification techniques are described in section “Enhancing peptide stability and bioavailability through structural modifications”, where we have summarized them in more detail.

In the s, since du Vigneaud et al.11 synthesized the first peptide, Pitocin® (oxytocin), the technology of peptide synthesis has developed rapidly. Peptide synthesis techniques include two categories: biosynthesis and chemical synthesis. Furthermore, Biosynthesis methods can be divided into natural extraction, enzymatic, fermentation and genetic recombination methods, etc. The chemical synthesis of peptides is further divided into SPPS299 and Liquid Phase Peptide Synthesis (LPPS).300 Both of them play important roles in the field of peptide synthesis.

As far as biosynthesis is concerned, the desired peptides are mainly obtained by microbial fermentation, enzyme catalysis and genetic recombination, etc. For example, the synthesis of formadicin, ramoplanin, vancomycin, and teicoplanin.301 Besides, the synthesis of the bis-intercalators family is highly dependent on biosynthetic methods. Bis-intercalators family are a class of C2-symmetric cyclic non-ribosomal peptides produced by actinomycetes, which can be inserted into DNA molecules through two unique chromophores in their structures, hence possessing good biological activities such as antimicrobial and antitumor activities. The complex molecular structure of the double-embedded family of non-ribosomal peptides makes chemical synthesis very challenging, while microbial fermentation is the main method for the production of this family of compounds. Thiocoraline,302 Triostin A,303 and Echinomycin304 are members of the bis-intercalators family, their synthesis is often via biosynthetic methods. Their key core skeletons, such as 3-hydroxyquinaldic acid, (3HQA), quinoxaline-2-carboxylic acid, (QXC) and 6-methoxy-3-hydroxyquinaldic acid, are synthesized in the presence of a range of enzymes (e.g. thioesterase) are synthesized. Finally, a series of fragments are assembled stepwise to obtain the corresponding antitumor peptides.305

On the other hand, with respect to chemical synthesis methods. LPPS is mainly composed of two strategies: stepwise synthesis and fragment combination. The advantages of LPPS include low cost, a wide choice of protecting groups, easy to scale up the synthesis, suitable for the synthesis of short peptides, and convenient purification.300 SPPS was proposed by Merrifield299 in and has been greatly developed to date. According to the different α-amino protecting groups, it can be divided into tert-butoxy-carbony (Boc) method and 9-fluorenyl-methoxycarbonyl (Fmoc) method.38,306 Compared with liquid-phase synthesis, solid-phase synthesis is convenient to operate, easy to realize automated processing, higher product yield and purity, which greatly promotes the development of peptide drugs.307 Nowadays, the synergies between solid-phase and liquid-phase synthesis methods play an important role in the field of peptide synthesis. In addition, as auxiliary technologies, microwave-assisted synthesis as well as advances in microchannel flow technology have made it possible to precisely control the reaction time (<1 s) and temperature, which greatly enhances the efficiency.38,308,309 For example, the synthesis of feglymycin using a linear method of microfluidic amide bond formation has enabled the preparation of biologically active oligopeptides with highly racemized amino acids, which are attractive drug candidates.310

Reniochalistatin E, the only tryptophan-containing cyclo-octapeptide in the reniochalistatins family. To date, it is the only peptide with antitumor activity (including RPMI-, MGC-803, HL-60, HepG2, and HeLa) among all the members of the reniochalistatins family, and it is particularly cytotoxic to myeloma RPMI- and gastric MGC-803.311,312 It uses 2-chlorotriphenyl chloride (CTC) resin as the carrier and HCTU/DIPEA as the condensation system to synthesize the straight-chain peptide by Fmoc solid-phase synthesis and liquid-phase cyclization to obtain the cyclic peptide crude product. Then it was purified through preparative liquid chromatograph. Next, the product was characterized by HPLC, MS and NMR. Finally, the purity of the target product as well as the overall yield was significantly higher than that of the peptide obtained from this reniochalina stalagmitis.313 This method is simple and easy to operate, applicable to the synthesis of the series of cyclic peptides.

HYD-PEP06 is a peptide compound obtained via Solid-Phase synthesis by Yang et al.314 It is an RGD-modified endostatin-derived synthetic peptide consisting of 30 amino acids. Its N-terminal RGDRGD fragment can specifically bind to different integrins in endothelial cells, which has been shown to have anti-tumor effects in colorectal cancer, oral squamous cell carcinoma and hepatocellular carcinoma (HCC).315,316 Based on the “one drug and two targets” design, Yang et al. selected the anti-tumor active fragment of endostatin and carried out structural modification and optimization on this basis, so that it could not only inhibit vascular activity, but also further have de-integrin effect. The mechanism of HYD-PEP06 is to block PI3K/AKT to inhibit the epithelial-mesenchymal transformation of liver cancer cells. Moreover, it also can inhibit the glomus formation and migration of liver cancer tumor stem cells by blocking the Wnt/β-catenin signaling pathway, so as to delay the development of liver cancer. Research has shown that HYD-PEP06 can inhibit the growth of subcutaneous graft tumor remnants of hepatocellular carcinoma in nude mice and has significant therapeutic effects on lung metastasis of hepatocellular carcinoma.314 It is a promising drug for the treatment of hepatocellular carcinoma and colorectal cancer by regulating the action of tiny nucleotides, ion channels and tumor stem cells, thus comprehensively inhibiting tumor recurrence and metastasis. Currently, the drug has completed undergoing clinical phase I trials (CTR), and is actively undergoing clinical phase II trials (CTR).

Furthermore, there are a number of globally approved drugs that utilize chemical synthesis techniques, including Sandostatin® (octreotide), Velcade® (bortezomib), Aphexda® (motixafortide).

Octreotide (Sandostatin®) is a typical octapeptide derivative synthesized using the Fmoc-SPPS method. The synthesis of octreotide acetate by solid-phase peptide synthesis and the adoption of the trans-salt process has the characteristics of simple operation, easy to operate, higher yield, etc., which are suitable for industrialized production.317 It is pharmacologically mimicking natural growth hormone. However, it is more potent than the natural hormone.318 Octreotide acts on the somatostatin receptors and causes vascular smooth muscle contraction by inhibiting the coupling of G proteins to phospholipase C. Like somatostatin, it induces an increase in calcium entry through L-type calcium channels, which leads to an increase in calcium-induced calcium release from the sarcoplasmic reticulum in smooth muscle cells through calcium-induced calcium release channels of the ryanodine receptor, then, it initiates the contractile cycle through activation of myosin light-chain kinase via interaction with calcium-calmodulin.319 Besides, octreotide can directly inhibit tumor angiogenesis by down-regulating growth hormone (GH) release.318,320 And its two formats, injection and slow-release microspheres, were approved by the FDA for the treatment of carcinoid tumors, vasoactive intestinal peptide tumors, in and , respectively.321 Additionally, octreotide is a classic therapeutic peptide that is also used in chemotherapy-associated refractory or intractable diarrhea, graft-versus-host disease, and HIV-associated diarrhea due to cryptosporidiosis, but is not approved by the FDA.322 Similarly, there are drugs such as lanreotide and pasireotide that exert the same effect. Furthermore, octreotide has been successfully used in imaging (neuroendocrine, endocrine, breast, small cell lung and prostate cancers) and more recently in targeted radiation therapy.323 It is one of the more prominent examples.

Bortezomib (Velcade®), a reversible inhibitor of the 26S proteasome, was obtained by chemical synthesis. It can be synthesized by convergent approach, and the use of TBTU (a condensation reagent, O-Benzotriazol-1-yl-N,N,N’,N’-tetramethyluronium tetrafluoroborate) inhibits the racemization during fragment condensation. It was approved by the FDA as a borate peptide for the treatment of multiple myeloma in . Then Approved for mantle cell lymphoma in . This drug exerts its antitumor effects mainly by inhibiting key substances in nuclear factor κB (NF-κB) pathway involved in cell proliferation, apoptosis, and angiogenesis.324 However, it is associated with hematotoxicity and peripheral neuropathy, as well as exhibiting poor permeability and pharmacokinetic parameters in solid tumors. Additionally, its chemical stability and bioavailability are low. Recent studies have demonstrated that nanoparticle delivery can overcome these limitations, offering a solution to circumvent the challenges encountered by conventional cancer chemotherapy drug administration.325

Motixafortide (Aphexda®) is a synthetic cyclic peptide approved by the FDA in September .326 It is utilized in combination with filgrastim (granulocyte colony-stimulating factor G-CSF) to facilitate the mobilization of hematopoietic stem cells into the peripheral blood for subsequent collection and autologous transplantation in patients diagnosed with multiple myeloma. Notably, motixafortide is the first innovative drug in nearly a decade to be approved by the FDA in the field of stem cell mobilization in multiple myeloma. It was shown that motixafortide blocks the interaction between CXCL12 and CXCR4 by binding to CXCR4 on hematopoietic stem cells (HSCs) and having a long receptor occupancy (>48 h).206,327 Peripheral blood and stored HSCs collected by apheresis were injected back into the patient (autologous transplantation) or into the recipient patient (allogeneic transplantation) to repopulate the bone marrow. Furthermore, motixafortide has been demonstrated to effect “cold” tumors in multiple modes of action, such as immune cell migration, tumor infiltration by immune effector T cells, reduction of immunosuppressive cells (e.g., Myeloid-derived suppressor cells, MDSCs) in the tumor microenvironment, etc. which can turn “cold” tumors (e.g., pancreatic cancer) “hot” (i.e., sensitizing them to immune checkpoint inhibitors and chemotherapy).206,328,329 And then, in a Phase II clinical trial, motixafortide in combination with cemiplimab and gemcitabine showed very significant results in the treatment of patients with pancreatic cancer.206,326

In the case of chemically modified peptides, for example, Polyethylene glycol (PEG) modification, Amino acid substitution modification and Cyclization modification and, etc. More details on the content of modified peptides are in section “Enhancing peptide stability and bioavailability through structural modifications”. Here we won’t describe it too much.

The occurrence and development of human tumors are related to the inhibition and intracellular degradation of p53 by negative regulatory proteins murine double minute 2 (MDM2) and murine double minute 4 (MDM4 or MDMX) of tumor suppressor p53. Therefore, antagonizing MDM2 and MDMX to activate and stabilize p53 is an important strategy for anti-tumor drug design.330 The peptide PMI (TSFAEYWNLLSP), which inhibits p53-MDM2 interaction with high solubility and specificity, was synthesized by phage display technology.331,332 However, the side chains of multiple residues of PMI are prone to interact with each other. Based on this, Lu et.al. screened and designed a peptide PMI-M3 (LTFLEYWAQLMQ) with low affinity for MDM2 and MDMX in pmol/L level through systematic mutation analysis and free energy addition principle.175 In addition, the researchers also obtained modified peptides of PMI-2K (KTSFAEYWNLLSPK) and M3-2K (KLTFLEYWAQLMQK) by adding lysine residues at both ends of PMI and PMI-M3 to improve their cellular uptake. Moreover, M3-2K can significantly improve the anti-tumor activity of p53-dependent in vitro and in vivo, and this p53-MDM2/MDMX interaction is expected to be further developed as a peptide inhibitor.

Peptides used in tumor diagnosis

Peptides can not only be used as drugs, but also used as molecular probe tools for molecular diagnosis and imaging of tumors. Peptides scintigraphy and peptide receptor radiotherapy have been developed based on the fact that tumor cells express one of the five growth inhibitory receptor subtypes. Currently, radioactive elements such as indium 111 (In), yttrium 90 (Y), gallium 68 (Ga), and technetium 99 m (Tc) can selectively bind to different somatostatin analogs via chelating groups.333 Here we focus on peptides developed in recent years for use in imaging and diagnostics.

In , [68Ga]Ga-DOTA-TOC was approved by the EMA for the specific imaging of tumor cells expressing somatostatin receptors (SSTRs), a radiopharmaceutical that combines the radionuclide 68Ga with the somatostatin analog DOTA-TOC to function. Then, the FDA approved [68Ga]Ga-DOTA-TOC in as the first 68Ga radiopharmaceutical in the U.S. to use positron emission tomography (PET) to image the somatostatin receptor (SSTR)-positive gastroenteropancreatic neuroendocrine tumors. This radioactive probe will help locate tumors in adults and children with a rare disease, somatostatin receptor-positive neu-endocrine tumors (NETs).334

Similarly, Illuccix®, also called gallium (68Ga) gozetotide or gallium (68Ga) PSMA-11 was approved by the FDA in for PET imaging of prostate-specific membrane antigen (PSMA)-positive prostate cancer, in which this radiopharmaceutical combines the radionuclide 68Ga with a mimetic peptide, Glu-NH-CO-NH-Lys(Ahx)-HBED-CC, which allows for the PSMA-expressing tumor cells for specific imaging. This targeting approach can also be used to develop treatment plans and potentially to assess treatment response. Notably, it is the first drug to use radioactive 68Ga for PET imaging of PSMA-positive prostate cancer.335

In addition, Pluvicto® (lutetium Lu 177 vipivotide tetraxetan, 177Lu-PSMA-617) is also available for the treatment of adult patients with prostate-specific membrane antigen (PSMA)-positive metastatic castration-resistant prostate cancer (mCRPC), who have received prior androgen receptor (AR) inhibitor and paclitaxel-based chemotherapy, which was approved by the FDA in March .92 Similarly, Posluma® (flotufolastat F-18), a high-affinity PSMA-targeted radio-diagnostic reagent based on a novel radio-hybridization technology, was approved by the FDA in May for testing in males with suspected metastatic prostate cancer for PET (Positron Emission Tomography) of PSMA prostate-specific membrane antigen (PSMA)-positive lesions.336

Generally, possessing the advantage of peptides in nature, anti-tumor peptides can inhibit the proliferation, migration and invasion of tumor cells and promote the apoptosis of tumor cells by preventing protein interaction, regulating the conformation of biomolecules, competing for receptor binding and destroying the cell membrane. In addition, anti-tumor peptides are able to be used as molecular probe tools for tumor molecular diagnosis and imaging in the field of tumor drug therapy. At the same time, peptides from different sources can be modified to make them have longer half-life, stronger resistance to enzymatic hydrolysis, relatively complete pharmacokinetic properties. With the in-depth understanding of tumor pathology and the development of new drug discovery technology, it is believed that more anti-tumor peptide drugs will be developed in the future.

Advance of peptide-based therapeutics in cardiovascular disease

Cardiovascular disease (CVD) is the leading cause of death in the world, accounting for about 50% of all deaths.337 The World Heart Report, launched at the World Heart Summit, shows that CVD deaths jumped globally from 12.1 million in to 20.5 million in , with four in five of those deaths occurring in low and middle-income countries.338 Diabetes mellitus, obesity, hypertension, hyperlipidemia, and other factors are high risk factors for cardiovascular disease, which manifests itself clinically as impaired cardiac function due to myocardial hypertrophy and fibrosis, and ultimately, death resulting from heart failure. A variety of ways have been used in preclinical studies for the treatment of cardiovascular disease, including small molecule drugs, protein drugs, and gene therapy to address the pathologic process.339,340,341,342,343 Peptides, due to their advantages, have become important therapeutic agents for cardiovascular diseases focusing on the symptoms of hypertension, vascular function and coronary artery disease, and acute coronary syndromes (ACS). Here, we briefly presented the overview of peptides in cardiovascular diseases.

Peptide drugs acting on G protein-coupled receptors

GLP-1 is an incretin secreting hormone secreted by intestinal L cells. GLP-1 receptor is a class B G protein-coupled receptor, which plays an important role in glucose homeostasis and the treatment of T2DM.344 GLP-1 exerts its effects through GLP-1 receptor binding. Studies have found that GLP-1 receptor is not only expressed in islet β cells, but also widely distributed in brain, lung, gastrointestinal tract, kidney, liver, heart and other organs in the body.252,345 Therefore, GLP-1 RAs may have effects on multiple organs, and the development of such drugs opens new ideas for the treatment of various clinical diseases. The first GLP-1 RAs is exenatide, which was approved by the FDA for the treatment of T2DM in . Studies have shown that GLP-1 RAs is not only effective in glycemic control, but also beneficial in the prevention of cardiovascular disease and weight loss similar to sodium-glucose transporter 2 (SGLT2) inhibitors.346 For example, liraglutide, dulaglutide and semaglutide can significantly reduce the incidence of major cardiovascular events (MACE) in patients with T2DM. In addition, the study showed that the risk of MACE in T2DM patients with a history of cardiovascular disease was significantly reduced by 14%, while the risk of MACE in patients without a history of cardiovascular disease was significantly reduced by 6%.345,347 In addition, the use of GLP-1 RAs in nondiabetic patients has focused on a modest improvement in left ventricular function after 7 days of acute treatment with the GLP-1 RA in patients with ST-segment elevation myocardial infarction (STEMI).348,349 Similar results were seen in patients without STEMI, independent of diabetes status. This suggests that GLP-1 RAs has some preventive effect on cardiovascular events in nondiabetic populations, but more study data are needed to support this. Although the peptide GLP-1 RAs is not FDA-approved for the treatment of cardiovascular disease, the potential therapeutic pleiotropic effects of the peptide GLP-1 RAs in patients with cardiovascular disease may extend beyond the treatment of diabetes in the future.350

Human urotensin-II (UII) is the strongest vasoconstrictor found in mammals.351 and it acts through the activation of the UII receptor (UT), the orphan G protein-coupled receptor (GPR14), collectively referred to as the UII/UT system.352 Urantide is a peptide UT antagonist derived from Urotensin II(UII).353 Urantide contains a core peptide consisting of six amino acid residues (Cys-Phe-Trp-Lys-Tyr-Cys), which has certain biological activity. Therefore, urantide has a high affinity with the UTs of human, mouse, monkey and other animals. Studies have shown that urantide can reduce the content of oxygen free radicals and anti-lipid peroxidation in myocardial tissue by activating PI3K /Akt and PKC signal transduction pathways, further regulate the expression of Bcl-2-associated X (Bax) and B-cell lymphoma 2 (Bcl-2) proteins, and inhibit the apoptosis of myocardial cells in rats with myocardial ischemia/reperfusion.198 Attenuating cardiotoxicity induced by doxorubicin and protecting primary cardiomyocytes depended on the down-regulation of p38 in the MAPK pathway.354 The protective effect of urantide on DOX-induced myocardial injury was more obvious. In studies where allantoin was administered to rats with atherosclerosis, it was found that allantoin reduced myocardial injury and lowered, serum creatine kinase (CK) and lactate dehydrogenase (LDH) levels by blocking the UII/UT system and regulating the mitogen-activated protein kinase (MAPK) signaling pathway. Urantide also reduced the levels of UII and its receptor, p38, p-extracellular signal-regulated kinase (ERK) and p-c-Jun N-terminal kinase (JNK) in myocardial tissue to protect cardiovascular function.354,355 At present, the research of urantide in cardiovascular diseases is still in the stage of animal experiments.

Peptide drugs acting on natriuretic peptide receptors

Natriuretic peptides (NPs) family consists of atrial natriuretic peptide (ANP), brain natriuretic peptide (BNP), and C-type natriuretic peptide (CNP).356 play an important role in the prevention of plasma volume expansion and hypertension.357 Natriuretic peptide regulates cardiovascular homeostasis mainly through three membrane receptors: natriuretic peptide receptor A (NPR-A) and natriuretic peptide receptor B (NPR-B) are guanylyl cyclase-coupled receptors, and natriuretic peptide receptor C (NPR-C) is a non-guanylyl cyclase-receptor on the cell surface.358 NPs mainly act through NPR-A and/or NPR-B receptors, while NPR-C is mainly used to remove NPs.359 The cardiovascular effects of natriuretic peptides include a reduction in peripheral vascular resistance and cardiac preload.360,361 Nesiritide, a recombinant human brain natriuretic peptide (rh-BNP) that mimics brain natriuretic peptide (BNP) action, plays a role in patients with decompensated heart failure. Clinical studies with intravenous injections have shown that, Nesiritide has a potent, dose-related vasodilator effect that is rapid and long-lasting. It was approved by the FDA in for the treatment of acute decompensated heart failure in patients at rest or with mild dyspnea.362,363 However, nesiritide is not widely used due to, which has side effects of headache and decreased blood pressure, low specificity and safety, now withdrawn from the market.362,364

Carperitide, a cyclic, recombinant α-human atrial natriuretic peptide (hANP), was approved by the PMDA in for the treatment of acute congestive heart failure (ADHF).365,366 ANP often induces a biological response by binding to guanylate cyclase-coupled receptor NPR-A. The use of carperitide in patients with ADHF can reduce central filling pressure and plasma aldosterone concentration, increase cardiac output, diuresis, and improve hemodynamics in the acute phase of ADHF.367 Hypotension was considered to be the most common adverse effect of caperitide, followed by renal dysfunction, and the increased in-hospital mortality in ADHF patients was significantly associated with the use of caperitide. However, a low dose (0.025-0.050 μg/kg/min, 0. μg/kg/min in some cases) of continuous intravenous infusion of carperitide can reduce side effects and in-hospital mortality, but this report still needs to be confirmed by a large number of samples and multi-center trials.368 In addition, carperitide can also increase coronary blood flow (CBF), reduce myocardial contraction and metabolic dysfunction, and limit infarct size. Among them, nitric oxide (NO) plays an important role in carperitide-induced ischemic heart vasodilatation and cardioprotection.369

Peptide drugs acting on angiotensin-converting enzyme (ACE)

Hypertension is a common chronic cardiovascular disease characterized by persistently elevated arterial blood pressure. It is a major risk factor for cardiovascular diseases such as atherosclerosis, coronary heart disease, stroke and myocardial infarction, besides, it is also a major cause of premature death worldwide. Angiotensin-converting enzyme inhibitors, angiotensin II receptor blockers, calcium channel blockers, β-adrenoceptor blockers and diuretics are commonly used antihypertensive drugs in the world.370 These antihypertensive drugs have good antihypertensive effects, but there are also many side effects. For example, some drugs that normally lower blood pressure may abnormally raise blood pressure, or increase blood pressure after stopping the drug, and cause symptoms such as dry cough, rash, edema, and acute renal impairment.371,372 Among the active peptides that have antihypertensive effects, most of them achieve their antihypertensive effects by inhibiting the activity of ACE. Active peptides with antihypertensive effects are similar to other peptide drugs in that they exert their therapeutic effects (lowering blood pressure) without any side effects on the body. Ace-inhibiting peptides are derived from a wide range of animal, plant and Marine organisms through the hydrolysis of hydrolases (e.g., pepsin, chymotrypsin and trypsin) and microbial enzymes (e.g., alkaline protease, thermolysin, conditase and proteinase K).373

LKPNM, a peptide derived from Katsuobushi, is an ACE inhibitor with antihypertensive activity, it has been used as a pharmaceutical ingredient in antihypertensive capsules.374,375 ACE-inhibiting peptides have also been identified from other species. For example, ACE-inhibiting peptides have been identified from other species, such as four peptide sequences (Gly-Gly-Pro-Ala-Gly-Pro-Ala-Val, Gly-Pro-Val-Ala, Pro-Pro, and Gly-Phe) isolated and extracted from salmon gelatinase hydrolysate (SC-1), which possess ACE and dipeptidyl peptidase IV (DPP-IV) inhibitory activity as well as oxygen radical uptake capacity. The peptides showed strong antihypertensive effects in a rat model of spontaneous hypertension, suggesting that the peptides could be used as pharmaceutical ingredients for the treatment of hypertension and related diseases.376 The results indicate that this polypeptide may be used as a drug preparation for the treatment of hypertension and related diseases.

Novel ACE-inhibiting peptides, VVLASLK, LTLK, LEPWR, ELPPK and LPTEK, were screened, identified and synthesized from cutlass, among which peptide LEPWR exhibited the best ACE-inhibiting ability. The antihypertensive effect of the ultrafiltration fraction was confirmed by using a spontaneously hypertensive rat (SHR) model. LEPWR antagonize ACE in a mixed competitive mode and forms six hydrogen bonds with ACE. The present study demonstrates that the hypotensive effects produced by the cutlery are attributed to these peptides.377 Identification of novel ACE inhibitory peptides from Pacific saury: In vivo antihypertensive effect and transport route. Dual inhibitory properties of mechanically boneless chicken residue (MDCR) hydrolysate on angiotensin-I converting enzyme (ACE-1) and dipeptidyl peptidase 4 (DPP4). Using food-grade protease to hydrolyzate MDCR, a potent peptide with dual inhibitory effects on ACE-1 and DPP4 was identified and then isolated to obtain IY (ACE-1-inhibitor) and VL (DPP4-inhibitor) peptides with dual effects on blood pressure and blood glucose regulation. Low Molecular Weight Peptide Fraction from Poultry Byproduct Hydrolysate Features Dual ACE-1 and DPP4 Inhibition.378

Peptide drugs acting on GPIIb /IIIa receptors

Eptifibatide, a ring heptapeptide derived from a protein in the venom of crotchtails, can be specifically recognized by GPIIb /IIIa receptors on the surface of platelets, blocking the binding of fibrinogen to GPIIb /IIIa receptors and inhibiting the final pathway of platelet aggregation. Its ring structure increases the bioavailability of the drug and its resistance to plasma proteases.379,380 It has been approved by the FDA for clinical use in Acute Coronary syndrome (ACS), including heart attacks and other emergencies such as sudden cessation of blood supply to the heart.381,382

Acute coronary syndromes are recognized by damage to the walls of the coronary arteries, resulting in the formation of intraluminal thrombi that block one or more coronary arteries, leading to unstable angina, non-ST-segment elevation myocardial infarction, and ST-segment elevation myocardial infarction.9 Antiplatelet therapy may be a therapeutic basis for the prevention and treatment of recurrent cardiovascular events in patients with acute coronary syndrome and undergoing percutaneous coronary intervention (PCI). In addition, it has been shown that intracoronary eptifibatide combined with PCI in the treatment of non-ST-segment elevation-acute coronary syndrome (NSTE-ACS) can effectively improve blood flow, increase myocardial perfusion to a certain extent, reduce perioperative platelet aggregation, which can improve cardiac function, with better results than PCI alone, besides, it has a better safety profile than that of abciximab.383,384 Furthermore, compared with the non-peptide small molecule GPIIb /IIIa receptor antagonists orbofiban, xemilofiban, sibrafiban and roxifiban (high side effects, such as thrombogenicity and high mortality),385 eptifibatide has little antigenicity.386 Eptifibatide has stronger binding ability to GPIIb /IIIa receptors, better safety, and faster action. The results of comparative experiments showed that intravenous administration of eptifibatide avoided the pre-systemic metabolism of liver and gastrointestinal enzymes, resulting in complete systemic availability. At a given dose, the pharmacokinetics of eptifibatide is linearly dose-dependent, and the antiplatelet activity of regular doses of eptifibatide is superior to that of tirofiban (GPIIb/IIIa inhibitor).379

Peptide drugs acting on tyrosine kinase receptors

Neuroglial protein (NRG)-1, also known as neural differentiation factor (NDF) or glial growth factor (GGF), is expressed in the cardiovascular system, nervous system, gut, kidney, and mammary gland.387,388 It is a ligand for tyrosine kinase receptors of ErbB3 and ErbB4 and structurally belongs to the epidermal growth factor (EGF) family. It directly binds to the ErbB4 receptor on cardiomyocytes, which activates the receptor and produces the corresponding bioactivities. The NRG-1/Erb B signaling system is not only involved in the regulation of cardiac embryonic development, but also closely related to the formation of cardiac structure, the maintenance of cardiac function, and the development of heart failure. Recombinant human neuromodulin (rhNRG)-1 peptide can attenuate myocardial injury in various animal models of cardiomyopathy. Additionally, it has therapeutic effects on heart failure. Phase II (CTR) and phase III (NCT) clinical trials of rhNRG-1 in heart failure are in progress.

Advance of peptide-based therapeutics in infection

Anti-infection is an important area of therapeutic peptides. The infection is caused by pathogens such as bacteria, viruses, fungi, parasites, etc. Infectious diseases are one of the major challenges in current clinical medicine. Penicillin is the first anti-infective drug applied in the clinic in the world. After years of development, more and more antibiotics have sprung up and made significant contributions to the cause of resisting bacterial infections for all mankind, but the problem of drug resistance caused by the misuse of antibiotics has also gradually come to the fore. In , it is estimated that nearly 5 million deaths were related to antimicrobial resistance, of which 1.27 million were directly caused by antimicrobial resistance. At the same time, “superbugs” are constantly expanding outwardly. As a result of drug resistance, antibiotics and other antimicrobial medicines become ineffective and infections become increasingly difficult or impossible to treat.389,390 The development of new antibiotics is insufficiently motivated globally, and there is an urgent need for new anti-infective drugs, which makes the research on antimicrobial peptides one of the current hotspots. More than antimicrobial peptides (AMPs) have been identified to date. Among them, gramicidin, daptomycin, colistin, vancomycin, oritavancin, dalbavancin and telavancin has been approved by the FDA for the treatment of bacterial infections.391

Antimicrobial peptides (AMPs), as a class of possible alternatives to antibiotics, are good candidates for overcoming antibiotic resistance due to their high antimicrobial activity, broad antimicrobial spectrum, variety, specificity, and the fact that the target strains are not prone to resistant mutations.392 It is a small molecule peptide that plays an important role in the host innate immunity. Most AMPs are short (10-50 amino acids), possess a positive charge (ranging from 2 to 11), and contain a large percentage of hydrophobic residues (usually 50%).391,393,394 Structurally, AMPs have α-helical, β-sheet, or extended/random-coil structures.395,396,397 Traditional antibiotics target an enzyme or a protein along a metabolic pathway, however, bacteria can produce new proteins through genetic mutations and thus become resistant to the drug, most antimicrobial peptides act by disrupting the membrane integrity of the target organisms and/or by transmigrating across microbial membranes to reach intracellular targets.398 Most antimicrobial peptides can kill microbial pathogens directly and the antimicrobial action of antimicrobial peptides tends to be very rapid, e.g., tick-defensin-derived Os(3-12)NH2 and computer-designed PaDBS1R1 peptides both kill microorganisms within 5-10 minutes of the exposure.399,400 Moreover, many of these antimicrobial peptides have a wide spectrum of antimicrobial effects, including activity against gram-positive and gram-negative microorganisms, fungi, unicellular protozoa and viruses. Also, some AMPs exhibit immunomodulatory activity, resulting in the indirect facilitation of pathogen clearance from the host.400,401,402,403

In terms of AMPs structure, the net positive charge of AMPs is capable of electrostatic interaction with negatively charged microbial membranes, and it has low selectivity to neutrally charged mammalian cell membranes. The hydrophobic residues enable them to penetrate cells and induce membrane cleavage. Meanwhile, the increased hydrophobicity of amino acid sequences can also reduce the selectivity and toxicity to mammalian cells.404 In fact, AMPs still contain some anions. However, its mechanism of action is difficult to determine, and it still has a significant role in anti-infection. In addition to their antimicrobial properties, AMPs play a pivotal role in intracellular processes such as angiogenesis, arteriogenesis, inflammatory responses, cell signaling, and wound healing, which makes them interesting candidates for research and development of the innovative drug.405,406,407

Antibacterial mechanisms

As mentioned above, most AMPs contain a net positive charge, a considerable proportion of amino acid hydrophobic residues, and contain α-helix, β-sheet and other secondary structures, which play an important role in antibacterial and anti-infection.404 It has great prospects in the field of anti-infection. At present, the mechanism of action of AMPs mainly includes two modes of action: Membrane action model and non-membrane action model.408,409 In membrane interaction models, disruption of microbial membranes by AMPs can occur through different mechanisms, including disruption of the lipid bilayer (barrel-stave model and toroidal-pore model), thinning of the membrane lipid bilayer, and subsequent membrane solubilization (carpet model, and aggregate model).410 On the other hand, some AMPs do not rely on a direct membrane breakdown mechanism but pass through the bacterial cytoplasmic membrane without necessarily disrupting it, then it interferes with fundamental processes such as DNA and protein synthesis, or inhibit other intracellular targets.411,412 Of these, binding of AMPs to bacterial cell membranes occurs through electrostatic interactions between cationic AMPs and anionic lipopolysaccharides (lipopolysaccharides from gram-negative bacteria) or (lipoblasts and peptidoglycans from gram-positive bacteria), and the subsequent invasion of AMPs into the cell exerts their respective interactions to achieve anti-infective properties.413 Nowadays, a variety of peptide antibiotics and non-antibiotic anti-infective drugs are under pre-clinical feasibility study or marketed for the treatment of infectious diseases.

Classical AMPs

Daptomycin, a cyclic lipopeptide antibiotic developed and marketed by Cubist Pharmaceutical Company in , is a classic antimicrobial peptide drug. It has become the world’s first cyclic lipopeptide antibiotic and it is the first-line drug for the treatment of infections caused by drug-resistant Gram-positive bacteria. In , daptomycin was approved for use in the United States for the treatment of complex and structural skin infections caused by gram-positive bacteria, as well as bacteremia and right-sided endocarditis caused by Staphylococcus aureus.414 Daptomycin mainly acts on the cell membrane of gram-positive bacteria. Within neutral pH, daptomycin has a negative charge and its antibacterial activity is dependent on calcium ions. When daptomycin is close to the bacterial cell membrane, it oligomerizes under the action of calcium ions and forms an “ion channel” like structure on the cell membrane, which causes intracellular ions to flow out, rapidly depolarizes the cell membrane, and blocks the synthesis of RNA, DNA and macromolecular proteins, and finally, bacterial death due to these factors.415,416,417 In addition, surotomycin, optimized by the fatty acid side chain of daptomycin, has been shown to have a rapid bactericidal effect on Clostridium difficile infection (CDI). It is currently in phase III clinical trials for the treatment of CDI.418,419,420

Dalbavancin (Xydalba®), a second-generation, semi-synthetic lipoglycopeptide antibiotic, has strong antibacterial activity against a variety of gram-positive bacteria (including methicillin-resistant Staphylococcus aureus MRSA and Streptococcus pyogenes) and some streptococci, with a long half-life and good tolerance.421,422 Xydalba® was approved by the EMA in for the treatment of acute bacterial skin and skin structure infections (ABSSSI) caused by gram-positive bacteria, including methicillin-resistant Staphylococcus aureus (MRSA). Interestingly, dalbavancin, one of the star drugs in the field of antibiotics, was first discovered and developed by Vicuron company in the United States, and then passed through Pfizer and Durata Company before it was approved by the FDA in May . Based on the positive results of two randomized, double-blind and multi-center clinical trials (SOLO I and SOLO II trials), FDA approved a new antibiotic, which is named Oritavancin in the same year.423,424,425 Oritavancin injection for the treatment of ABSSSIs caused by gram-positive bacteria (including methicillin-resistant Staphylococcus aureus, MRSA) in adult patients. Oritavancin and dalbavancin (approved by the FDA in ), as well as the vancomycin (approved by the FDA in ), are belong to the new second-generation, semi-synthetic glycopeptide antibiotics. Notably, it is the first and only antibiotic for the treatment of ABSSSIs with a single-dose regimen.423

Rezafungin (Rezzayo®) was approved by the FDA in March for the treatment of candidemia and invasive candidiasis in adults, the first therapeutic treatment approved for invasive candida infections in nearly a decade.426 Rezafungin is a novel echinocandin that acts by inhibiting β-1,3-glucose synthetase thereby disrupting the integrity of fungal cell wall.427,428 The approval of this drug further expands the application of AMPs in the field of anti-infectives. Besides, there are some promising AMPs under study.

The antimicrobial peptide peceleganan (PL-5) spray is a non-antibiotic anti-infective drug. As a new type of peptide broad-spectrum anti-infective drug, it has a unique bactericidal mechanism.429 It acts as a bacterial penicillin binding proteins family inhibitor (PBPs inhibitor), showing strong bactericidal advantages against gram-positive, Gram-negative and various antibiotic-resistant bacteria. It can be used for various wound infections such as burns, traumatic ulcers, venous ulcer infections and Wagner class II diabetic foot ulcers, etc. Currently, antimicrobial peptide PL-5 spray is in the U.S. Phase II clinical study (NCT), which is expected to achieve better results.

Teixobactin, a novel peptide antibiotic obtained by screening bacteria from soil samples has a wide range of activity against multi-drug resistant gram-positive pathogens such as methicillin-resistant staphylococcus aureus, streptococcus pneumoniae, and vancomycin-resistant enterococci.430 Teixobactin targets the peptidoglycan precursor lipid II, which has the dual effect of inhibiting peptidoglycan synthesis and disrupting cell membranes, enabling an effective attack on bacterial cell membranes.431,432,433 Antimicrobial experimental studies in vitro have shown that teixobactin has a good inhibitory effect on gram-positive bacteria and is non-toxic to mammalian cells, as well as non-hemolytic and non-genotoxic, which make it a promising drug lead-compound.434

Recently, Li et al. reported that CT-K3K7, a scorpion antimicrobial peptide derivative, was able to inhibit the growth of Candida both in vitro and in vivo. CT-K3K7 can kill Candida by destroying cell membrane or nucleus and interacting with nucleic acid. It can also induce candida cell necrosis and inhibit its mycelium and biofilm formation through reactive oxygen species (ROS)-related pathways. In the mouse model of subcutaneous infection, CT-K3K7 significantly prevented skin abscess formation and reduced the number of recovered Candida cells in the infected area. Therefore, CT-K3K7 is expected to be a promising drug for the treatment of Candida skin infections. In addition, there are a number of AMPs derived from natural sources, as well as AMPs from engineered sources, that play an important role in the field of anti-infection. Such as the rhesus theta-defensin-1 (RTD-1),435,436 a natural macrocyclic AMP expressed in primate leukocytes. Its macrocyclic structure confers drug stability and resistance to cleavage by the large number of proteases found in the saliva of cystic fibrosis (CF) patients, furthermore, it is more effective than the usual natural AMPs. In addition, RTD-1 showed rapid in vitro bactericidal activity against mucoid, non-mucoid and multi-drug resistant clinical isolates of Pseudomonas aeruginosa, without cross-resistance to colistin. Nowadays, it is considered as a potential therapeutic agent for CF airway infections. WLBU2 is a cationic amphipathic peptide with membranolytic activity composed of Arginine, Valine and Tryptophan in different arrangements.437 Due to its broad-spectrum antimicrobial activity against ESKAPE (Enterococcus faecium, Staphylococcus aureus, Klebsiella pneumoniae, Acinetobacter baumannii, Pseudomonas aeruginosa and Enterobacter species) pathogens.438,439,440 the molecule is currently in Phase I clinical trials (NCT) for the treatment of periprosthetic joint infections.

Overall, AMPs have been initially used in the field of biomedicine research with their unique advantages. AMPs not only have certain inhibitory effects on bacteria, fungi and anti-parasites, but also a certain inhibitory and killing effects on enveloped viruses and tumor cells. The broad-spectrum antibacterial activity of AMPs has been studied as an alternative to antibiotics. It is believed that AMPs will be more rapidly developed in the field of biomedicine in the near future.

Advance of peptide-based therapeutics in anti-coronavirus

Currently, the coronaviruses(COVs) that infect humans (HCoVs) contain seven COVs.441 Within these, Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2), Middle East Respiratory Syndrome Coronavirus (MERS-CoV), and SARS Coronavirus (SARS-CoV) are highly pathogenic COVs that can cause serious illness and even death. In contrast, HCoV-229E, HCoV-OC43, HCoV-NL63 and HCoV-HKU1 are considered low pathogenic HCoVs as they usually cause mild disease in humans.442 Novel Coronavirus (COVID-19), which is caused by SARS-CoV-2 infection, has led to about 770 million diagnosed cases and more than 7 million deaths worldwide as of January 24, (From https:// covid19.who.int). For the prevention and treatment of COVID-19, there are three dominant antiviral options: vaccines, neutralizing antibodies, and small molecule drugs.443 Among them, vaccines can realize early prevention to a certain extent, neutralizing antibodies are expected to be able to directly block the invasion of the virus into the host cell, while small molecule drugs can block the replication of the virus in the host cell by targeting the core shared proteins, which play a therapeutic role in post-infection treatment and disease control.444 It is hoped that long-term effective and broad-spectrum small molecule drugs can be developed. With regard to vaccines and neutralizing antibodies, SARS-CoV-2 continues to spread globally with the mutation of the new COVs, and new variants are still emerging rapidly, posing a major challenge to current vaccine and therapeutic approaches. Thus, this calls for a sustained supply of highly effective, low-toxicity antiviral drugs to combat SARS-CoV-2 and its variants. Peptide drugs are gaining attention in the development of anti-SARS drugs due to its low immunogenicity, low cost, specificity and modularity enabling it to be tailored to the virus.

Development of anti- COVs candidate targets

In the process of viral invasion, viruses need to enter the target cells through the steps of receptor recognition and membrane fusion or endocytosis, in which the receptor-binding subunit of the viral surface protein (SP) plays a role in mediating the recognition between the virus and the cell receptor, and the transmembrane subunit of the viral SP plays a role in mediating the fusion of the membrane, which are the common antiviral targets in this process. In addition, there are receptors on the cell, host cell proteases needed to cleave the SP, and so on. After the virus enters the target cell, it releases its own DNA/RNA into the cell, which serves as a template to direct the synthesis of viral proteins. At the same time the viral genome undergoes simultaneous self-replication. During this process, certain viral proteases act to cleave viral precursor proteins, while RNA viruses require RNA-dependent RNA or DNA polymerases (RdRp and RdDp) for replication. The RdRp and RdDp are not present in the human body, so viral proteases as well as RdRp and RdDp are also important antiviral targets. Then, the newly synthesized viral genome and viral proteins assemble into new viral particles, which are released to the outside of the cell for dissemination through budding or cell trafficking pathways. Within the same viral genus, the sequences and structures of proteins that perform the same function are often highly similar, such as the spike (S) protein structures of SARS-CoV and SARS-CoV-2. These proteins often serve as common targets for the development of antiviral drugs.445

The protein structure of COVs includes structural proteins and Non-structural proteins (NSPs). The structural proteins are composed of the spike (S) protein, envelope (E) protein, membrane (M) protein and nucleocapsid (N) protein.446 Theoretically, all the COVs enzymes and proteins involved in viral replication and control of the host cell machinery are potential drug targets, including S protein, papain-like protease (PLPro) and master protease (Mpro/3 CLpro), and viral RNA-dependent RNA polymerase (RdRP). This chapter focuses on the use of peptides in COVID-19, with a brief overview of hot peptide targets in COVID-19.

A) The S protein, a key protein in COVs, and its role

The coronavirus spike S protein plays an important role in the invasion of COVs. Similar to other COVs, SARS-CoV-2 infection requires the fusion of the viral envelope and cell membrane, which is mediated by the viral spike (S) glycoprotein.447 It regulates viral entry into host cells and is also the major antigenic determinant, often the target of the host antibody response. The COVID-19 is caused by severe acute respiratory SARS-CoV-2 infection. S protein is an important protein that mediates virus entry into cells. The receptor binding domain (RBD) on the S1 subunit is responsible for the binding of the virus to the cell surface receptor.448,449,450,451 Many neutralizing antibodies target this region to inhibit RBD binding to the viral receptor, thereby blocking COVs infection. However, this region is not conserved and there is great variability among COVs, which makes most of the neutralizing antibodies targeting this region lack broad-spectrum and can only inhibit one or a few COVs.446 The heptad repeat 1/2 (HR1/HR2) region of the S2 subunit can interact with each other to form a six-helix structure, which mediates the fusion of the virus with cells and the entry into cells for replication. The formation of the homologous hexahelix can be blocked by the addition of exogenous peptides that can interact with the HR1 or HR2 region of the S2 subunit, thereby inhibiting the invasion of the virus into cells. In addition, the mechanism of membrane fusion between virus and target cells is very conserved in different COVs, and the six-helix bundle formation between the HR1 and HR2 domains plays a key role in driving membrane fusion, which also makes this region an important target for the development of broad-spectrum viral fusion/invasion inhibitors.

In addition, researchers found that the affinity of the S protein of SARS-CoV-2 to the human Angiotensin-converting enzyme 2 (ACE2) receptor was much higher than that of SARS.448 Based on the researchers’ elucidation of the electron microscopic structures of the full-length proteins of the ACE2-B0AT1 complex and the ACE2-RBD complex, which revealed how the S protein of -nCoV interacts with the receptor ACE2 at the atomic level, the structural basis and functional characterization of the entry of SARS-CoV-2 into the target cells were further clarified. The above results show that -nCoV enters human cells by targeting the receptor domain (RBD) of the viral transmembrane S protein to the receptor protein, ACE2. In other words, the interaction between S protein and ACE2 is the key way for the virus to enter cells. ACE2 is like the “door handle”, and SARS-CoV-2 opens the door to enter human cells by grasping the “door handle”. Therefore, blocking the interaction between ACE2 and -nCoV S protein can be used as one of the potential effective ways to prevent and control COVID-19.

B) 3CL protease (3C-like protease, 3CLPRO, or Mpro)-a key enzyme in COVs replication

COVs are enveloped viruses with a positive-stranded, single-stranded RNA genome of 26 to 32 kb in length. During SARS-CoV-2 infection of the host, the S protein on the outer surface of the viral particles is responsible for binding to the host receptor for attachment to the cell membrane, followed by fusion of the viral and host cell membranes and release of the viral genomic RNA into the cell. Next, there is fusion of the virus and host cell membranes and release of viral genomic RNA into the cell. Subsequently, two viral replicase polyproteins are translated into two polyproteins, pp1a and pp1ab, by controlling the production of two viral replicase polyproteins by host ribosomes.452 These can be further processed by viral-encoded proteases in 16 mature nonstructural proteins (NSPs). The main protease (Mpro), or 3C-like protease (3CLpro), is an extremely important protease in the propagation of other coronaviruses, such as Middle East Respiratory Syndrome Coronavirus (MERS-CoV), which is responsible for the cleavage of protein precursors of the translated viral genome, resulting in a number of nonstructural proteins (NSPs), that assemble to form the viral replication-transcription enzyme complex NSPs. Finally, these non-structural proteins are assembled to form the viral replicase-transcriptase complex (RTC).453 Therefore, 3CLpro inhibitors can be developed to inhibit the activity of 3CLpro and interfere with the viral replication process. Moreover, since of 3CLpro is highly conserved among different genera of COVs and has no homologous proteins in the human body, it implies that the design of inhibitors by using it as a drug target can achieve a better expectation of selectivity and safety, and it has a great potential to be explored, which has become the direction of research in the field of human in the anti-coronaviral field.

Peptides for coronaviruses (COVID-19 as an example)

Paxlovid® is currently the only peptidomimetic drug approved by the FDA for severe symptoms caused by COVID-19. In December , Emergency Use Authorization (EUA) of Paxlovid by the FDA for the treatment of SARS-CoV-2 in mild-to-moderate adults and pediatric and adult patients ≥12 years of age with a body mass of ≥40 kg, and in patient populations at higher risk of severe disease.454,455 Preliminary results of a phase III clinical trial showed that patients who were treated within 3 days of symptom onset had an 89% lower risk of hospitalization and death from any cause than those who received placebo.456 Until May , the drug was formally approved by the FDA from Emergency Use Authorization (EUA).

In fact, Paxlovid® is a COVID-19 combination product, a combination of the 3CL protease inhibitor PF- (also known as Nirmatrelvir) and low-dose Ritonavir, both peptide analogues.455,457 Development of PF- began with the SARS-CoV outbreak in . During the SARS-CoV epidemic period, Pfizer’s researchers tried to design SARS-CoV 3CL protease inhibitors based on Rupintrivir (an irreversible inhibitor of human rhinovirus 3CL protease) as the starting point. After a series of optimization, PF- was obtained. However, the inhibitor has not yet been tested in animals, and it has been shelved due to the demise of the SARS-CoV epidemic. It was restarted in due to the outbreak of COVID-19. Ritonavir was developed to solve the problem that multiple sites of PF- are oxidized and metabolized by CYP3A4 (cytochrome P450 3A4 enzyme) in liver microsomes.457,458 Ritonavir, as a pharmacokinetic enhancer of Nirmatrelvir, slows down its breakdown in vivo and enhances its half-life and bioavailability by inhibiting the degradation of Nirmatrelvir by CYP3A4. so that it remains active in the body for a longer period of time and at higher concentrations to help fight the virus.455,459 This strategy is also borrowed from previous antiviral drug delivery strategies. Efficacy data from the phase II/III study of Paxlovid showed an 86% reduction in the risk of COVID-19-related hospitalization or death from any cause among patients who received Paxlovid® within five days of symptom onset. Oral Paxlovid is superior to intravenous treatments such as Remdesivir (approved by the FDA in October ),460 as well as lower hospitalization and mortality rates than Molnupiravir (approved by the FDA in ), since then, Paxlovid® (PF- and Ritonavir) was born and remains relevant for the treatment of COVID-19 and some of its variants. This antiviral therapy has also been derived from the oral anti-COVID-19 innovative drug Simnotrelvir/Ritonavir, which is safe and effective, and has shown remarkable efficacy and safety in clinical II/III trials.

EK1 polypeptide, a peptide inhibitor developed by Jiang et al., broadly inhibits multiple human COVs capable of infecting humans, which binds to the HR1 region of a variety of coronaviruses to form a hetero-hexamer. Thus, it competitively inhibits the formation of hexamers from the HR1/HR2 interaction of the virus itself, thereby inhibiting the fusion of the virus with the host cell and preventing the entry of the virus’ genes into the cell for replication. After the outbreak of COVID-19, they demonstrated that EK1 peptide can also bind to the HR1 of SARS-CoV-2 and efficiently inhibit the invasion and infection of SARS-CoV-2 into host cells.461 Currently, the EK1 nebulizer is about to enter a Phase IIa clinical trial. Besides, by modifying with palmitic acid or cholesterol, Jiang/Lu et al. have further developed a series of more efficient and broad-spectrum lipopeptide fusion/invasion inhibitors, such as EK1C4, EKL1C, and EK1-C16, which are equally effective in inhibiting the infection of various SARS-CoV-2 variants without being affected by variants.462,463

In addition to the HR1 region, the HR2 region is also an important target for the development of viral fusion/invasion inhibitors. However, the HR1-derived peptides contain too many hydrophobic amino acids to maintain a stable α-helix structure in aqueous solution. Therefore, they designed a protein inhibitor targeting the HR2 region of SARS-CoV-2, 5-helix (composed of two HR2 and three HR1-derived protein fragments), which can bind to the HR2 region of SARS-CoV-2 to form a heterologous six-helix bundle and inhibit the invasion of the virus into host cells. Like the EK1 peptide, nCoV-5-helix can also effectively inhibit the infection of various SARS-CoV-2 variants, including Omicron and Delta.464 Compared with common neutralizing antibody drugs, peptide-based inhibitors have the following obvious advantages: (1) Peptide-based inhibitors act on the conserved HR1/HR2 locus, which allows peptide-based inhibitors to demonstrate broader-spectrum COVs inhibitory activity than targeting RBD/NTD-neutralizing antibodies; (2) Peptide-based inhibitors have shorter amino acid sequences and smaller molecular weights, which can be directly synthesized by chemical synthesis, greatly reducing the time and production cost; Moreover, it can directly inhibit the replication of COVs in the patient’s lung by intranasal administration. (3) high safety and efficacy, basically equal to antibody drugs. The only drawback is that the half-life of peptide drugs is shorter than that of antibody drugs. (4) Peptide-based inhibitors are more stable and can be stored for a long time at room temperature, with lower storage and transportation costs. Therefore, the development of Pan-CoV fusion/invasion inhibitors can effectively prevent and treat SARS-CoV and its variants, SARS-CoV and MERS-CoV infections.

HY was developed in the early stage of the COVID-19 outbreak. Based on the analysis of the membrane fusion mechanism of SARS-CoV-2, SARS-CoV,MERS-CoV.465,466,467 Gao et al. designed a polypeptide inhibitor P3 targeting SARS-CoV-2 HR1, which can inhibit theinfection of SARS-CoV-2,467 but peptide P3 has insufficient activity. In order to further improve the activity of P3 peptides and develop anti-SARS-CoV-2 polypeptide membrane fusion inhibitors, researchers designed many P3 derivatives based on their structure, finally, they found that the antiviral activity of P315 peptide was the best, which was about 8-fold higher than that of P3. Furthermore, through PEG and cholesterol modification, the P315V3 lipopeptide with about -fold higher activity was obtained (HY).468 It can bind to the HR1 region of the S protein of SARS-CoV-2 and inhibit membrane fusion to prevent the virus from entering the cell interior. At the same time, the hydrophobic tail of HY can be fixed to the surface of the cell membrane, forming a protective barrier at the cell surface. Besides, it can effectively inhibit novel COVs variants, including XBB.1.5. In addition, it can inhibit SARS-CoV, MERS-CoV and seasonal COVs HCoV-NL63, HCoV-229E and HCoV-OC43, HY shows a broad spectrum of COVs. Based on the effective antiviral effect, HY nasal spray was approved by NMPA for phase I clinical trial in August . Now, the phase I clinical trial has been completed and the drug shows enough safety. Based on it, it is currently under clinical phase II trial in China. Furthermore, HY was accepted by the US FDA clinical trial in January .

PeptiOrigin Product Page

As mentioned above, targeting the most conserved HR1 region of SARS-CoV-2 can be an important target for the development of broad-spectrum viral fusion/invasion inhibitors. By using the membrane fusion mechanism, researchers developed a peptide YKYY017 (from SARSCoV-2 HR2-deriving lipopeptides, IPB20)468,469 that targeted the conserved HR1 region and competitively bound to the viral HR1 region, then YKYY017 prevents the formation of the viral homologous six-helix bundle (6-HB) structure, and blocks the fusion process between the virus and host cells. It can inhibit two membrane fusion pathways, cell surface and endosomes, to exert antiviral effects. In vitro pharmacodynamic studies showed that YKYY017 had significant and broad-spectrum inhibitory effects on the original SARS-CoV-2 strain and its epidemic variants (Delta, Omicron BA.1, BA.2, BA.4 and BF.7). YKYY017 has a broad spectrum against cold-causing coronaviruses (HCoV-NL63, HCoV-229E, etc.), severe acute respiratory syndrome coronavirus (SARS-CoV), Middle East respiratory syndrome coronavirus (MERS-CoV), and bat-derived coronavirus RaTG13), pangolin-derived coronaviruses (PCoV-GD, PCoV-Gx), it has also shown significant inhibitory effects, reflecting the broad-spectrum anti-coronavirus activity. Toxicological studies showed that YKYY017 had no obvious toxicity and related adverse reactions, as well as a good therapeutic and preventive effects. It is currently in Phase II/III clinical trial in China (ChiCTR) and was approved for clinical investigation on May 12th, . As a new generation of COVs membrane fusion inhibitor with broad spectrum, YKYY017 aerosol inhalation is expected to be well used in the fight against SARS-CoV-2 and its variants.

Recently, Dang et al. developed a new broad-spectrum SARS-CoV-2 fusion inhibitor, A1L35HR2m-Chol, which can effectively inhibit different SARS-CoV-2 mutant strains, as well as SARS-CoV and MERS-CoV.470 The researchers interconnected the ACE2-derived peptide A1 with the N-terminus of the HR2m peptide via a flexible junction (GGGGS)7 of the appropriate length to generate A1-(GGGGS)7-HR2m (A1L35HR2m), (HR2mL35A1) was then obtained by exchanging the positions of the A1 peptide and the HR2m peptide. In addition, two other fusion peptides were constructed by shortening the length of the junction to derive A1L5HR2m and A1HR2m. The researchers then evaluated the ability of A1L35HR2m, HR2mL35A1, A1L5HR2m, and A1LHR2m to inhibit SARS-CoV-2 by the SARS-CoV-2 pseudotyped virus infection assay. It was found that the addition of ACE2-derived peptide A1 to the N-terminus of HR2m peptide with a long flexible junction significantly increased the anti-SARS-CoV-2 activity.

This research revealed that the antiviral mechanism of A1L35HR2m may be similar to that of HR2-derived peptides, which can enhance the competitive interaction with the affinity of HR1 structural domains to block the fusion of viruses with target cell membranes. Meanwhile, adopting the S protein-mediated cell-cell fusion assay, it was found that A1L35HR2m-Chol effectively inhibited SARS-CoV-2 D614G S protein-mediated cell fusion between 293 T/EGFP/S and Caco-2 cells, which contributes to the enhancement of the viral activity; moreover, the researchers also performed site-specific modification of the C-terminus of A1L35HR2m to generate A1L35HR2m, which is an anti-viral peptide that can be used to prevent the virus from fusing with the target cell membrane. The results showed that A1L35HR2m-Chol exhibited broad and effective inhibitory activity against different SARS-CoV-2 VOC, SARS-CoV and MERS-CoV, and did not show in vitro or in vivo toxicity.As well, the level of A1L35HR2m-Chol dose had no effect on the hepatic and renal functions of the mice.

In general, peptides against acute respiratory distress syndrome (ARDS) caused by coronaviruses and related respiratory problems caused by SARS-COV-2 infection, most of them are target-based synthetic peptides. Researchers have been working on the development of peptide therapeutics for a variety of diseases, including Covid-19, due to their ease of synthesis, high target specificity, selectivity, and low toxicity, etc. In addition, a peptide-based Covid-19 vaccine is in clinical trials. Finally, a number of novel peptide drug delivery systems with high potential for other disease development. Peptide drugs are expected to be further developed in the future.

Advance of peptide-based therapeutics in digestive diseases

The use of peptide drugs in gastrointestinal diseases is mainly focused on the control of acute upper gastrointestinal bleeding (such as somatostatin analogues and vasopressin analogues) and other intestinal diseases (Linzess®, Trulance®).Vapreotide (Sanvar®), a synthetic somatostatin analogue, was approved in Mexico in .471,472 In addition, a phase III clinical trial found that the combination of Vapreotide and endoscopic therapy was more effective than endoscopic therapy alone in controlling acute bleeding in patients with cirrhosis and variceal bleeding.473 Terlipressin (Terlivaz®) is a synthetic vasopressin analogue developed by Ferring Pharmaceuticals, which can be used to treat gastrointestinal variceal bleeding. In terms of anti-gastrointestinal bleeding, the main focus is on bleeding caused by cirrhosis.474 It was approved by the FDA in to improve the rapid decline in renal function in adult patients with hepatorenal syndrome (HRS).475

In addition, peptides have been shown to be effective in irritable bowel syndrome (IBS). Irritable bowel syndrome (IBS) is a chronic functional gastrointestinal disease characterized by smooth muscle dysfunction, which is prone to relapse and has a high incidence worldwide. “IBS is characterized by persistent or intermittent episodes of abdominal pain, bloating, and abnormalities in bowel habits and stool form, without morphological or biochemical abnormalities.” Linaclotide (Linzess®) is a guanylate cyclase-C (GC-C) agonist, which is a 14-amino acid polypeptide.476,477 Linzess® was approved by the FDA in December for the treatment of adult patients with constipation-predominant irritable bowel syndrome (IBS-C) and chronic idiopathic constipation (CIC). It is also the first new prescription drug approved for the treatment of adult patients with moderate to severe IBS-C in Europe. Linzess® and its active metabolites have been shown to bind to guanylate cyclase-C (GC-C) receptors on the luminal surface of the small intestinal epithelium, leading to the activation of GC-C and the increase of intracellular and extracellular concentrations of cyclic guanosine monophosphide (cGMP). Intracellular cGMP increases the secretion of chloride and bicarbonate in the small intestine by activating the cystic fibrosis transmembrane conductance regulator (CFTR), which eventually leads to increased secretion of intestinal fluid and accelerated intestinal transport. Through GC-C activation, linaclotide reduced intestinal pain and increased gastrointestinal transit speed in animal models. Extracellular cGMP can reduce the activity of painful nerve fibers and alleviate visceral pain in animal models.478,479

In January , the FDA approved Trulanc® (plecanatide), a GC-C (guanylate cyclase-C) agonist with linaclotide, for CIC (Chronic Idiopathic Constipation).479,480 Plecanatide is as effective as linaclotide in the treatment of CIC but not as effective as linaclotide in the treatment of IBS-C.481 In terms of structure, Plecanatide and Linaclotide are both small molecule monopeptide, but with one less disulfide bond, which is easier to synthesize. However, Plecanatide has a higher dosage than linaclotide in the indication of IBS, which is expected to be further optimized in future studies.

Elsiglutide is an orally active and selective GLP-2 receptor agonist, which is an analogue of GLP-2. It can increase cell proliferation, reduce intestinal cell apoptosis, and regulate intestinal balance.482,483 In a rat model, Elsiglutide ameliorated Lapatinib (HY-) -induced diarrhea in rats. In addition, clinical phase II studies testing elsiglutide in colorectal cancer (CRC) patients receiving chemotherapy (CT) induced diarrhea with 5-fluorouracil (5-FU) have not yet yielded results.484 Therefore, peptide drugs also occupy a certain proportion in the field of digestive tract diseases.

Advance of peptide-based therapeutics in Alzheimer’s disease

Alzheimer’s disease (AD) is a multifactorial neurodegenerative disease with insidious onset, clinically characterized by memory impairment, aphasia, loss of recognition, visuospatial skills impairment, behavioral changes and other comprehensive dementia manifestations and more serious neurological impairments, which seriously affects the patient’s quality of life, and in severe cases even death.485 It is estimated that by , this number will reach 130 million. This will create a huge social burden.486 Worse still, the main challenge in the treatment of Alzheimer’s disease is that its pathogenesis has not been fully clarified, and the more recognized hypotheses of the pathogenesis include deposition of β-amyloid (Aβ)487 and intracellular Tau protein aggregates,488 neuroinflammation489 and lack of acetylcholine (Ach), oxidative stress,490 and bio-metal dyshomeostasis, etc.491 The Aβ deposition hypothesis suggests that the normal mechanism of Aβ removal from the brain tissue of AD patients is disrupted, leading to the accumulation of toxic Aβ in the brain tissue to form Aβ plaques,492 and when the accumulation reaches a certain level, it triggers a series of neurodegenerative processes, such as inflammation, oxidative stress, and the deposition of Tau proteins,493 which ultimately leads to a series of clinical conditions of AD. The establishment of the Tau protein deposition hypothesis is inextricably related to the Aβ hypothesis, which suggests that Tau protein as an important protein involved in stabilizing neuronal structures in the brain and regulating internal neuronal transport systems. Tau protein plays an important role in normal brain cell activity.494 However, due to the Aβ deposition mentioned in the above hypothesis, Tau protein accumulates and forms abnormal protein clumps (NFTs), which severely disrupts the normal function of neuronal activity and ultimately leads to a series of AD disorders in patients. In addition, Aβ accumulations in neurons can also activate glycogen synthase kinase 3-β (GSK-3β), which phosphorylates Tau proteins and ultimately forms NFT,495 making the clinical symptoms of AD appear gradually. At the same time, neuroinflammation exacerbates the pathological processes of Aβ accumulation, Tau accumulation and Tau phosphorylation,496 and the pathological processes of the three hypotheses produce a synergistic effect, which leads to a persistent vicious cycle, thus exacerbating the disease process of neuronal damage. In addition, the hypothesis of reduced acetylcholine levels is also recognized as an important cause of AD. In patients with AD, cholinergic neurons in the brain degenerate, resulting in reduced Ach levels,497 which in turn leads to a range of clinical symptoms of AD, including reduced learning and memory abilities. Currently approved therapies include cholinesterase inhibitors (rivastigmine, donepezil, tacrine and galantamine), and NMDA receptor antagonists, but even their combination provides only temporary relief of symptoms, not a cure.

The key clinicopathologic indicators of AD are amyloid plaques formed by amyloid accumulation in extracellular regions or hyperphosphorylated Tau proteins aggregated in intracellular regions, which form neural protofibrillary tangles (NFTs) in affected neurons. Amyloid plaques mainly contain amyloid β40/42 peptides, which further accumulate leading to neuroinflammation and mishandling of Tau protein.498,499 Based on this, a large number of domestic and international researchers have tried to develop drugs that can remove these two toxic proteins from the brain in the hope of treating AD or slowing down the process.500 Among them, Aβ is produced by hydrolyzing and processing amyloid precursor protein (APP) by β-secretase (BACE1) and γ-secretase, which means that inhibition of β- and γ-secretase cleavage can inhibit the production of Aβ protein. In this regard, many drugs targeting secretase have entered clinical trials,501 including CTS- (CoMentis), PF- (Pfizer), LY (Lilly), and AZD (AstraZeneca). The results of the clinical phase 1 trial showed that CTS- reduced plasma levels of Aβ protein502 665, and AZD was also shown in the clinical phase 2/3 trial. AZD also achieved the target efficacy in clinical phase 2/3 trial.503 However, the results of Phase II/III clinical trials for many other related new drug developments have been unsatisfactory.

Peptides can intervene in the process of Aβ aggregation in a variety of ways, and thus researchers continue to plow through the field. Driven by advances in drug design and synthesis techniques, a common strategy for peptides used in the treatment of AD is to design peptides with high affinity to bind to Aβ and prevent its aggregation. These peptides can alter the conformation of Aβ or inhibit its aggregation process by interacting with it. Phages can search for potential therapeutic peptides for various diseases based on the affinity between the produced peptide and the target molecule, so researchers have used it to search for peptides with the potential to inhibit Aβ aggregation. Kiessling et al.504 then identified several Aβ-affinity peptide ligands by phage display.505 Another strategy is to block amyloid or peptide production by designing peptides with the core sequence of the Aβ fibrillization to suppress protein or peptide aggregation. In , Prof. Kapurniotu of the Technical University of Munich, Germany506 designed a series of peptide-based small molecule inhibitors based on the protein sequences of islet amyloid polypeptide (IAPP), a type of small molecule inhibitor that can inhibit the root causes of amyloid proteins (amyloid- beta peptide and IAPP) aggregation, thus reducing the cytotoxicity of amyloid fibrils, and providing a new idea for the treatment of Alzheimer’s disease in the clinic. Prof. Kapurniotu further deepened his research on this basis,507 and developed a new type of cyclic peptide-based small-molecule inhibitor based on the straight-chain peptide-based small-molecule inhibitors. The optimized cyclic peptide-based small molecule inhibitor not only has high efficiency and specificity in binding and recognizing amyloid proteins, but also shows a high degree of protein hydrolysis stability in human plasma, and can cross the human blood-brain barrier in cellular models, making it a potentially effective anti-amyloid drug for the treatment of AD. In addition, chiral biomolecules have the advantage of inherent stereoselectivity, whereby the aggregation of Aβ can be hindered by modifications of chiral amino acids or peptides. Amino acid chirality can determine the folding of the peptide backbone, hydrogen bonding interactions, and even the biological functions of proteins in vivo.508 Xue et al. borrowed graphene to structurally modify a peptide to make it pharmaceutically active and regulate Aβ40 aggregation in vitro.509

Intracellular neuroprogenitor fiber tangles containing phosphorylated Tau proteins are a hallmark protein of AD, and hyperphosphorylation of Tau proteins causes the deposition of this characteristic protein. Meanwhile, Tau function is regulated by a variety of post-translational modifications at more than 50 sites, and Tau in healthy neurons carries multiple phosphorylation motifs that are located predominantly in its microtubule assembly domain. It has also been shown that Aβ can initiate a deleterious cascade of responses involving Tau pathology and neurodegeneration, i.e., Tau proteins are mediators of Aβ cytotoxicity.510 Possible synergistic effects of Aβ and Tau proteins on microglia and astrocytes, whose interactions mediate cognitive dysfunction in AD patients. Researchers have proposed multiple mechanisms of action against Tau, including specific removal of pathological Tau species, reduction of Tau production, promotion of Tau physiological function through microtubule stabilization or inhibition of post-translational modifications.511 However, most of the Tau protein-targeting therapies tested to date have been immunotherapies that can target Tau proteins intracellularly and/or extracellularly, but targeting extracellularly alone is unlikely to be effective,512 and no Tau-targeting therapies have yet to show definitive clinical efficacy in preclinical or early-stage AD. From the perspective of the aberrant aggregation of toxic Tau proteins, Zheng jie et al. designed and synthesized a peptide chimera drug, DEPTAC, which can specifically reduce the phosphorylation level of Tau proteins and thus promote their degradation to avoid aggregation.513 This drug can selectively recruit protein phosphatase 2 A (PP2A) to the periphery of Tau proteins and thus induce their dephosphorylation.

Several studies have shown that neuroinflammation plays a prominent role in the pathogenesis of Alzheimer’s disease. There is a correlation between neuroinflammation and amyloid and Tau pathology. When innate immune cells are activated, programmed cell death can be induced through a variety of pathways, and cell death usually leads to the release of pro-inflammatory cytokines that propagate the innate immune response and can eliminate beta plaques and aggregated Tau proteins.514 However, chronic neuroinflammation caused by cell death is associated with neurodegenerative diseases and may worsen Alzheimer’s disease. The continued elucidation of cell death pathways and central innate immune sensor signaling pathways involved in regulating neuroinflammation and Aβ/Tau clearance will have a major impact on the field of AD research. Recent evidence suggests an association between AD and T2DM.515 Numerous reports have found that GLP-1 RAs improve cognitive behaviors and pathological features in AD patients and animals, which may be related to the improvement of glucose metabolism in the brain. GLP-1 RAs not only reduce Aβ deposition by inhibiting Aβ production and facilitating its clearance, but also reduces inflammatory mediator. GLP-1 RAs also exert neuroprotective effects by inhibiting oxidative stress and reducing neuronal apoptosis.515 In addition, GLP-RAs can improve the cognitive function of AD patients by enhancing learning memory and spatial orientation.516

Though strides have been made, peptides still face some challenges in the treatment of AD at present. First, AD is a complex disease whose pathogenesis and pathological processes have not been fully clarified, so further in-depth studies are needed to identify the best therapeutic targets for peptide drugs. Second, the stability, pharmacokinetics and bioavailability of peptide drugs also need to be addressed to ensure that the drugs can effectively exert therapeutic effects in vivo.

Looking ahead, with a deeper understanding of the pathogenesis of AD and the continuous advancement of drug development technology, peptide drugs will have a broader prospect in the treatment of AD. More peptide drugs targeting AD may enter clinical trials in the future and are expected to provide new and more effective treatment options for AD patients. Meanwhile, research on the combined application of peptide drugs with other therapeutic methods also needs to be strengthened, with a view to achieving better therapeutic effects.

Advance of peptide-based therapeutics in rare diseases

Currently, there are approximately identified rare diseases. Although it is individually rare, it generally affects a significant portion of the general population (10% of the population). However, less than 6% of all rare diseases have approved treatment options, highlighting their huge unmet need for drug development.517,518 Rare diseases (RDs) were often chronic, leading to lifelong disability or early death; many RDs had pediatric onset, and about 30% of children with RDs died before age 5 years.519 70% of rare diseases are hereditary, caused by germline and somatic gene mutations.520 A small number of rare diseases are also caused by environmental, infectious or immunologic factors (e.g., African trypanosomiasis),521 but these will not be discussed here.

Peptides also play an important role in the treatment of rare diseases. Due to the diversity and complexity of the etiology of rare diseases, conventional drugs have limited therapeutic effects on their treatment. However, some specific peptide drugs, such as synthetic analogs or biosynthetic peptides, may have specific therapeutic effects on certain rare diseases. For example, certain rare genetic diseases may result in abnormal hormone levels, while other peptide drugs are able to regulate these hormone levels, thereby alleviating the symptoms or slowing down the progression of the disease. Therefore, the research and application of peptide drugs in the treatment of rare diseases has also attracted much attention and is expected to provide more effective treatment options for these patients. Here we focus on more recent developments.

Rett syndrome

Rett syndrome, originally proposed by Andreas Rett in , is a rare progressive neurodevelopmental disorder that results in severe intellectual disability, loss of mobility, and autism-like symptoms, other features include slowed head growth, seizures, autistic features, and respiratory abnormalities.522,523 Trofinetide (Daybue®) is a novel synthetic analog of the amino-terminal tripeptide of insulin-like growth factor I (IGF-1) that can be administered orally and is neuroprotective at minimal doses.524,525 It was approved by the FDA in March for the treatment of Rett syndrome for children 2 years of age and older, Notably, it is the first and only FDA-approved treatment for Rett syndrome.526 Studies have shown that trofinetide not only inhibits inflammatory cytokine production and overactivation of microglia and astrocytes, but also increases the amount of available IGF-1 that can bind to the IGF-1 receptor, which is beneficial for the treatment of core symptoms of Rett syndrome.524

Generalized myasthenia gravis (gMG)

Generalized myasthenia gravis (gMG) is a disorder of neuromuscular junction transmission caused by the destruction of the postsynaptic membrane of skeletal muscle by autoantibodies at the neuromuscular junction.527,528 The clinical manifestations of generalized myasthenia gravis (GMG) are fluctuating skeletal weakness and fatigue intolerance, which aggravate after activity and alleviate after rest. Local or general weakness is the main symptom. In severe cases, it involves respiratory muscles and causes respiratory failure, which is potentially fatal. Zilbrysq® is a novel macrocyclic peptide of a C5 complement inhibitor that can be used to treat adult patients with generalized myasthenia gravis (gMG) with positive anti-acetylcholine receptor (AChR) antibodies.526 It is also the first gMG targeting C5 complement inhibitor that requires only a once-daily subcutaneous injection, providing a simpler dosing option for patients with generalized myasthenia gravis. Zilucoplan (Zilbrysq®), a cyclic peptide drug, was approved by the FDA in October for the treatment of AChR antibody-positive gMG that fails to respond to other immunosuppressive therapies.529,530 As a complement C5 inhibitor, zilucoplan inhibits the complement mediated neuromuscular junction injury by targeting the mechanism of action. Different from the monoclonal antibody C5 inhibitor, zilucoplan, as a peptide, can be used with intravenous immunoglobulin and plasma exchange at the same time without supplementary administration, which is a new type of peptide drug with extremely simple administration.

Short bowel syndrome (SBS)

Short bowel syndrome (SBS) is mostly caused by large area surgical resection of the small intestine or congenital bowel diseases. In this condition, the absorption capacity of small intestine is limited, which cannot meet the needs of normal growth and development of patients, and parenteral nutrition (PN) support is required.531,532 Clinically, the most common disorders leading to SBS in adults are Crohn’s disease, mesenteric ischemia, radiation enteritis, postoperative adhesions, and postoperative complications. The common causes of SBS in children include congenital diseases such as necrotizing enterocolitis, volvulus, gastroschisis and intestinal atresia.533 Nutritional therapy is a very important treatment for SBS, and parenteral nutrition is very important for the survival of infants and children with SBS. However, the related complications caused by long-term PN may endanger the life of SBS children, and more effective drugs are urgently needed to meet the clinical needs.534,535

GLP-2 is a specific growth factor with intestinal protective effect, which can enhance the transport of glucose and lipid in intestinal cells, enhance nutrient absorption and promote intestinal adaptation.536 Teduglutide is a GLP-2 analogue that was previously approved by EMV in for the treatment of short bowel syndrome and malabsorption. In February , Teduglutide received NMPA approval for the treatment of short bowel syndrome in adults and children aged 1 year and older. The substitution of alanine at position 2 by glycine makes teduglutide resistant to degradation by dipeptidyl peptidase-4 (DPP-4). Thus, tidulutide has a longer half-life than GLP-2. Tidulutide can reduce the requirement of PN in SBS children, increase the tolerance of enteral nutrition (EN), help some children achieve intestinal autonomy, and has good safety and tolerance, which provides a new treatment option for SBS children suffering from PN.537 In addition, the duration of PN is positively correlated with the incidence of catheter-related infections. The present study proves that GLP-2 plays a role in intestinal adaptation. GLP-2 is a peptide secreted by intestinal L cells after foodintake.538,539 It can prevent the apoptosis of intestinal epithelial cells, thereby promoting the growth and recovery of intestinal tissue. This process of growth and repair is essential for children with SBS to achieve intestinal autonomy, which provides a new hope for reducing the dependence on PN.540,541

In addition, a number of peptide drugs have been approved for the treatment of other rare diseases. For example, Etelcalcetide (Parsabiv®), a calicomimetic agent, was approved by the FDA in February for the treatment of secondary hyperparathyroidism in adults with chronic kidney disease (CKD) undergoing hemodialysis.542 Glatiramer was approved in for the treatment of multiple sclerosis (MS), an immune-mediated inflammatory demyelinating disease of the central nervous system.543,544 The common clinical manifestations are recurrent vision loss, diplopia, limb sensory disturbance, limb movement disorder, ataxia, bladder or rectal dysfunction, etc.). In June , glatiramer acetate (Copaxone®) was officially approved in China for the treatment of adult patients with relapsing multiple sclerosis (MS). In addition, glatiramer acetate (Copaxone®) can also be used for the treatment of Rett syndrome in a phase II clinical trial (NCT).545 Other therapies include pegcetacoplan (a C3 complement inhibitor, approved by the FDA in May ) for paroxysmal nocturnal hemoglobinuria, and terlipressin (an AVPR1A agonist, approved by the FDA in September ) for hepatorenal syndrome (HRS).546

Advance of peptide-based therapeutics in orthopedic metabolic diseases

The application of peptide drugs in orthopedics is mainly focused on the treatment of osteoporosis. Among them, osteoporotic fracture (OPF) is the most serious symptom of osteoporosis, which is difficult to heal, has a high disability rate and difficult to heal, and poses a serious threat to the health of middle-aged and elderly people. Osteoporosis can be treated in two ways. On one hand, it reduces bone loss by inhibiting bone resorption; on the other hand, it decreases bone loss by increasing the number or activity of osteoblasts. However, the use of anti-bone resorption drugs alone does not restore lost bone structure. Increasing the number or activity of osteoblasts may be a more attractive approach to enhancing bone formation and promoting bone regeneration.547

Nowadays, there are many drugs to promote bone formation, including monoclonal antibodies to sclerostin and recombinant human parathyroid hormone drugs, parathyroid hormone-related protein (PTHrP) analogs, such as Evenity® (romosozumab), Forteo® (teriparatide) and Tymlos® (abaloparatide).548,549,550,551 Evenity® has the potentialside effect of increasing the risk of heart attack, stroke or death due to cardiovascular disease. Parathyroid hormone drugs have drawbacks such as a two-year use limit and a potential risk of osteosarcoma. Therefore, new strategies and methods are urgently needed for the research and development of drugs for bone formation. YLL3 and YLL8 are “osteogenic specific” peptides discovered by Yao et al. through the OBOC (One Bead One Compound) library. YLL3 and YLL8 not only have high affinity for osteogenic progenitor cells, but also activate the phosphorylation of the pro-survival factor Akt. In vitro experiments confirmed that YLL3 and YLL8 can increase the differentiation and maturation of osteoblasts. Moreover, YLL3 and YLL8 can target endogenous osteogenic progenitor cells for bone regeneration therapy.547

In addition, Osteogenic Growth Peptide (OGP) is also very effective in preventing osteoporosis. OGP is a peptide composed of fourteen amino acid residues which is a promoter of systemic response to bone marrow injury. Osteogenic growth peptide and its C-terminal pentapeptide OGP (10-14) can up-regulate the mRNA expression of type I collagen, bone calcium and alkaline phosphatase in osteoblast-like cells.552 At the same time, it can significantly increase the content of collagen, osteocalcin, calcium and alkaline phosphatase activity of cells, promote bone formation and inhibit bone resorption, increase the number of osteoblasts, and reduce the number of osteoclasts, so as to prevent osteoporosis.

Advance of peptide-based therapeutics in migraine disease

Migraine is a periodic neurological disorder in which patients often experience unilateral, throbbing recurring headaches accompanied by nausea, vomiting, light sensitivity, sound sensitivity, and other symptoms that last from hours to days. Migraine seriously reduces the quality of life and work of patients, and it even has a significant economic and social impact. The International Headache Society categorizes migraine into three types: migraine without aura, migraine with aura and chronic migraine.553 Migraine without aura is the most common type, accounting for 70 to 90 percent of cases.

Treatment of migraine includes medication and non-medication. For acute migraine, medication is the main treatment. Commonly used medications include painkillers, tricyclic antidepressants, calcium channel blockers, etc., which are used to relieve headaches and control the frequency of attacks. Despite the wide range of medications available, their efficacy is not satisfactory, and many others are difficult for patients to adhere to because of side effects. This is related to the fact that the exact cause of migraine is still not fully understood. In recent years, clinical models have identified key signaling molecules involved in migraine, including calcitonin gene-related peptide (CGRP), pituitary adenylate cyclase-activating peptide 38 (PACAP-38), and nitric oxide, whose exposure significantly increases the risk of migraine attacks. Among these molecules, calcitonin gene-related peptide (CGRP) is a vasodilatory neuropeptide that plays a crucial role in the pathophysiology of migraine and is a promising target for migraine therapy.554

Monoclonal antibodies targeting CGRP have demonstrated migraine prevention in multiple Phase II and Phase III clinical trials, and their long-term effects in key regions provide stable CGRP blockade beyond existing prevention methods. Monoclonal antibodies targeting calcitonin gene-related peptide (CGRP) and its receptor have opened a new era in migraine prevention.555

We refer to those that antagonize CGRP receptors as gepants, which have a strong affinity for CGRP receptors and can prevent other molecules from binding to them, blocking signaling. The advantage over other types of drugs is that they do not constrict blood vessels.556 Early studies of generation gepants relieved migraine symptoms, although their preventive effect on the disease has yet to be studied. However, development of the first generation of gepants has also been halted for various reasons, such as the discontinuation of ocegepant due to low oral bioavailability, and the banning of tecapant telcagepant and MK- due to hepatotoxicity after frequent use.557,558

Despite these safety concerns in the initial studies of first-generation gepants, the efficacy of gepants has prompted further efforts to develop safe CGRP-blocking molecules. Three second-generation gepants continue to be in clinical development: Rimegepant, Ubrogepant, and Atogepant. Rimegepant passed the clinical phase 2b trial based on its superior efficacy in treating acute migraine. This was followed by a Phase 3 trial (NCT, NCT), which was randomized, double-blind and placebo-controlled, which provided preliminary evidence of the findings of Phase IIb trial.559 In addition, safety studies were conducted concurrently. In the Phase 2b/3 study of atogepant, the drug was also found to reduce the number of migraine days per month in patients compared to placebo.560

In December , the U.S. Food and Drug Administration, approved the first gepant drug, ubrogepant (Ubrelvy®), for the acute treatment of migraine in adults. Aquipta® (atogepant) was approved by the EMA in August . It is also the first and only once-daily oral calcitonin gene-related peptide (CGRP) receptor antagonist (gepant) therapy in the European Union for the prophylactic treatment of chronic and episodic migraine. As an oral small molecule CGRP receptor antagonist, it is competitive, highly selective, and highly effective. In addition, atogepant not only prevents vasodilation, but also relieves or prevents migraine by preventing CGRP-induced neurogenic inflammation, injurious transmission, and various functions of CGRP. In a phase 3 double-blind trial, researchers randomly assigned adults with 4-14 migraine days per month in a 1:1:1:1 ratio to receive either a once-daily oral antimigraine agent (10 mg, 30 mg, or 60 mg) or a placebo for 12 weeks,561 and the data demonstrated that the combination of the agent taken orally once daily was effective in reducing the number of migraine days and headache days.562

Launched in September , Nurtec ODT® (rimegepant) is approved in several countries, including the US and the EU, for the acute treatment of migraine with or without aura in adults and the prophylactic treatment of episodic migraine in adults. Structurally, it contains a cyclohepta[b]pyridine core, and due to the poor water solubility of the pre-developed BMS- ( < 2 μg/mL), the team fitted Rimegepant with a primary amine, which resulted in a better potency and greatly increased water solubility of this drug (50 μg/mL). In vitro, Rimegepant effectively antagonized both the CGRP receptor and the insulin 1 (AMY 1) receptor, but was about 30 times more effective at blocking the CGRP receptor.563 It is effective not only for acute migraine,564,565 but also for prophylactic treatment.566

Launched in March , zavegepant is a third-generation small-molecule calcitonin gene-related peptide (CGRP) receptor antagonist developed by Pfizer Roots for the prevention and treatment of chronic and episodic migraine. It is the first nasal CGRP antagonist.

By comprehensively analyzing journals and patents, the researchers summarized the common structural features of CGRP receptor antagonists and synthesized the new compound. Although this molecule effectively inhibited CYP3A4, it had poor solubility. A SAR study of the side chain of benzothiophene identified 7-methylindazole as capable of enhancing activity and moderately inhibiting CYP3A4. By adding a fluorine atom at the C-8 position of the quinazolinone, the molecule BMS- was synthesized, which showed a substantial increase in solubility, but low oral bioavailability. The researchers replaced the oxidation-sensitive benzylidene methylene group with an electron-deficient sp2 hypomethyl group. At the same time, a simple SAR of the piperidinopiperidine side chain to N-methylpiperidinyl-piperazine (with two protonatable nitrogens) yielded BMS- (i.e., the marketed drug zavegepant), with a polar surface area of 116.18 Å2, which further reduced binding to serum proteins. In addition, the crystalline compound was surprisingly water-soluble, sufficient to support nasal administration of the drug ultimately developing zavegepant.

Introduction 
How peptide therapy works
Key areas of peptide therapy research
Advantages of peptide therapy over traditional treatments
Challenges and limitations
Future directions in peptide therapy
What this means for healthcare and drug development
References
Further reading 

Peptide therapy consists of a unique pharmaceutical class of agents that are constructed of a range of organized amino acids, which usually have a molecular weight of 500-5,000 Da.1

Image Credit: raker/Shutterstock.com

Precision vs. Personalized Medicine

Introduction

Since , when the first therapeutic peptide, insulin, was synthesized, many peptide-related accomplishments have reverberated throughout the industry, with more than 80 peptide drugs being approved globally.1

Peptide-based drug development has been a popular area of pharmaceutical research due to many peptides having specific physiological activities in the human body, such as oxytocin, vasopressin, and gonadotrophin-releasing hormone.1

The advantages of peptide therapy consist of their deep penetration into tissues such as the skin and intestines, compared to larger biomolecules, including antibodies, which enable a faster entrance into the bloodstream.1

Additionally, their low immunogenicity and high target specificity are superior to small drug molecules, which only target 2-5% of the human genome. In contrast, peptides have higher selectivity for specific protein targets.2

How peptide therapy works

Peptide drugs can work in a versatile manner, acting as hormones, growth factors, ion channel ligands, neurotransmitters, or anti-infective agents. They bind to cell surface receptors and stimulate intracellular effects with both high specificity and affinity.1

The mode of action for therapeutic peptides is similar to biologics such as therapeutic antibodies and proteins, with the advantage of having less immunogenicity and production expenditure.1

This growing area of pharmaceutical development led to 26 peptides being approved as drugs between and by the Food and Drug Administration (FDA), with more than 315 new drugs also being approved in that same time period.

The growth in this area has translated to more than 200 peptides being in current clinical development and another 600 peptides in preclinical trials.3 G17DT is an example of a therapeutic peptide that is currently undergoing clinical trials and indicates various forms of cancer.1

Key areas of peptide therapy research

Oncology

Investigational peptide therapies in cancer and targeted drug delivery are essential, with direct drug delivery into tumor cells mitigating off-target effects. This key characteristic causes decreased quality of life in patients receiving chemotherapy.4

Peptide therapeutics are also being investigated for their capacity to disturb and disrupt critical tumor anti-apoptosis proteins, as well as their ability to inhibit tumor drug resistance mechanisms through targeting related protein-protein signaling pathways.4

An example of a well-researched related peptide is Bim BH3, which stimulates apoptosis using the regular protein Bcl-2. With cancer cells varying in their cell surface receptors, even within the same organ, precision targeting mechanisms for these are critical, with peptide therapies being the ideal solution due to the capacity to screen and synthesize specific peptide sequences fit for purpose.4

Metabolic Disorders

Metabolic homeostasis is critical for all life activities, which is enabled through several biological pathways. The disruption of metabolic homeostasis can lead to the development of metabolic diseases, which are slow-forming and challenging, with many complex etiologies.

Approximately 35% of adults and 50% of the aging population have metabolic diseases within the United States.5

The most common metabolic diseases include, but are not limited to, obesity, diabetes, non-alcoholic fatty liver disease, metabolic function-associated fatty liver disease, and cardiovascular diseases.5

The peptide liraglutide, known under Saxenda, as well as semaglutide, known more commonly under the name Ozempic, are two obesity peptide drugs. Liraglutide consists of an agonist for the GLP-1 receptor, which has physiological effects such as increasing insulin release, reducing glucagon release, and decreasing hunger sensations.5

Studies on incorporating liraglutide into a lifestyle intervention experiment found that this peptide therapeutic caused an average weight loss of 4-6 kg within one year. Semaglutide also has similar effects to liraglutide, impacting hypoglycemia and weight loss.5

Neurodegenerative Diseases

Within neurodegenerative diseases that are characterized by gradual neuron loss in the brain, resulting in death, peptide therapy research may be significant in advancing insufficient available treatments to manage these diseases.6

Approximately 25% of global deaths and disabilities are caused by brain-associated diseases, including Alzheimer’s disease, Parkinson’s disease, Huntington’s disease, and amyotrophic lateral sclerosis (ALS).6

Peptide therapy research can be used to investigate the properties of misfolded proteins/peptides within neurodegenerative disorders that lead to disrupted signaling in neurons and their eventual death.6

Peptide vaccines, which have been predominantly researched for fighting against influenza, also include UB-311 and were found to stimulate improvements in cognition in patients with early-stage Alzheimer’s dementia.6

Other peptide inhibitors being researched for neurodegenerative diseases include (i) neurotrophins, (ii) NAPVSIPQ, (iii) and vasoactive intestinal peptides (VIP).6

Regenerative Medicine

Finally, with the wound healing process consisting of complex and multi-stage steps, such as hemostasis, inflammation, proliferation, and remodeling, the use of peptide research may be crucial in advancing this process, especially for those suffering from chronic wounds secondary to diseases.7

Peptide-based wound dressings may provide a novel approach to wound management, with many natural peptides, such as RL-QN15, derived from secretions in the skin, that function to accelerate the wound healing process.

Additionally, other natural peptides, such as fibroblast growth factor and epidermal growth factor, function to improve wound healing processes, including proliferation and tissue remodeling.7

Advantages of peptide therapy over traditional treatments

As previously mentioned, the advantages of peptide therapy over conventional treatments and small molecules are deeply rooted in properties such as high specificity, high biological activity, high penetrative ability for membranes, and low cost.6

This may lead to faster development timelines compared to small-molecule drugs that go through significant trial and error.9

Additionally, the increased specificity of peptide therapy can also lead to reduced side effects, which is pertinent in cancer therapeutics, such as chemotherapy, that can cause systemic adverse effects.4

The potential for utilizing peptide therapies for personalized medicine applications is also vast, with specific peptide sequences being screened and synthesized for targeting mechanisms and various cell surface receptors in many different diseases and disorders. In this way, peptide-based drug development may be significant for the future of targeted treatment.4

What's Next for Semaglutide? Beyond Diabetes and Weight Loss

Challenges and limitations

Challenges of peptide-based drug development consist of stability, toxicity, and immunogenicity. However, strategies for enhancing peptide stability include integrating D-amino acids or α-aminoxy amino acids, which modifies the backbone chemistry and cyclization.

Additionally, incorporating these into the manufacturing process can decrease storage stabilities by enabling peptides to be more sensitive to both pH and temperature, which can cause easy degradation.6

Regulatory hurdles for peptide-based drugs are also a challenge, with only 4% of FDA-approved peptide/protein drugs utilizing oral administration, which is the delivery route with the highest patient adherence rate.

However, oral administration can be challenging in itself, with barriers such as the intestinal epithelial membrane barrier and mucus barrier, which may prevent drugs from penetrating and absorbing effectively.8

Future directions in peptide therapy

With novel technologies such as artificial intelligence (AI) and machine learning, along with peptide therapeutic research, the future of drug development may advance significantly, reducing costs, enhancing personalized, targeted disease treatment, and decreasing toxicity.9

Traditional peptide discovery methods are limited in their ability to explore the large chemical arena of potential peptide sequences, which are time-consuming, expensive, and inefficient at finding promising targets.

This process can be accelerated with AI through efficient analysis of diverse datasets, including but not limited to genomic and clinical data and protein structures.9

Interestingly, deep learning and AI methods have discovered novel functional and antimicrobial peptides (AMPs) from many sources, such as the human proteome and microbiome; this is significant for developing alternative antibacterial drugs that will be effective against the rapid growth of antibiotic resistance.9

Overall, the integration into the drug discovery process can exponentially reduce the time and cost of identifying and developing new peptide therapies.9

What this means for healthcare and drug development

The development of new drugs and strategies, such as peptide therapies, is critical for healthcare, with aims to identify and characterize substances that hold the potential for improving patient outcomes and addressing unmet medical needs for a number of disease areas.9

With personalized medicine being at the forefront of the future of targeted therapies, hundreds of peptides are currently being researched in preclinical and clinical trials. This area is expected to grow exponentially, attracting both investment and research efforts.9

In , peptide drug sale reached $20 billion, and this was expected to grow to more than $50 billion by ; the market size is also likely to increase from $25.3 billion in to $41.7 billion by , with a Compound Annual Growth Rate (CAGR) of more than 8%.9

How is AI Transforming Drug Discovery?

If you want to learn more, please visit our website Peptide Drug Delivery.

References

1. Wang L, Wang N, Zhang W, et al. Therapeutic Peptides: Current Applications and Future Directions. Signal Transduction and Targeted Therapy. ;7(1). doi:10./s-022--4.
2. Barman P, Joshi S, Sharma S, Preet S, Sharma S, Saini A. Strategic Approaches to Improvise Peptide Drugs as Next Generation Therapeutics. International Journal of Peptide Research and Therapeutics. ;29(4). doi:10./s-023--3.
3. Rossino G, Marchese E, Galli G, et al. Peptides As Therapeutic Agents: Challenges and Opportunities in the Green Transition Era. Molecules. ;28(20):. doi:10./molecules.
4. Warthen JL, Lueckheide MJ. Peptides as Targeting Agents and Therapeutics: A Brief Overview. Biomacromolecules. ;25(11):-. doi:10./acs.biomac.4c.
5. Teng B, Li J, Ren P. Peptide Drugs Application in Metabolic Diseases and Discovery Strategies. Journal of Holistic Integrative Pharmacy. ;3(1):24-31. doi:10./s-(23)-8.
6. Baig MH, Ahmad K, Saeed M, et al. Peptide Based Therapeutics and Their Use for the Treatment of Neurodegenerative and Other Diseases. Biomedicine & Pharmacotherapy. ;103:574-581. doi:10./j.biopha..04.025.
7. He X, Wu W, Hu Y, et al. Visualizing the Global Trends of Peptides in Wound Healing Through an In‐Depth Bibliometric Analysis. International Wound Journal. ;21(4). doi:10./iwj..
8. Wu J, Sahoo JK, Li Y, Xu Q, Kaplan DL. Challenges in Delivering Therapeutic Peptides and Proteins: A Silk-Based Solution. Journal of Controlled Release. ;345:176-189. doi:10./j.jconrel..02.011.
9. Hashemi S, Vosough P, Taghizadeh S, Savardashtaki A. Therapeutic peptide development revolutionized: Harnessing the power of Artificial Intelligence for Drug Discovery. Heliyon. ;10(22). doi:10./j.heliyon..e.

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