View source for Discovery and development of dipeptidyl peptidase-4 inhibitors/en

<languages />
[[Dipeptidyl peptidase-4 inhibitors]] (DPP-4 inhibitors) are [[enzyme inhibitor]]s that inhibit the [[enzyme]] [[dipeptidyl peptidase-4]] (DPP-4). They are used in the treatment of [[type 2 diabetes mellitus]]. Inhibition of the DPP-4 enzyme prolongs and enhances the activity of [[incretin]]s that play an important role in [[insulin]] secretion and [[blood glucose]] control regulation.
Type 2 diabetes mellitus is a chronic metabolic disease that results from inability of the [[Beta cell|β-cells]] in the pancreas to secrete sufficient amounts of [[insulin]] to meet the body's needs. Insulin resistance and increased [[hepatic]] glucose production can also play a role by increasing the body's demand for insulin. Current [[therapy|treatments]], other than insulin supplementation, are sometimes not sufficient to achieve control and may cause undesirable [[adverse effect|side effect]]s, such as [[weight gain]] and [[hypoglycemia]]. In recent years, new drugs have been developed, based on continuing research into the mechanism of insulin production and regulation of the metabolism of sugar in the body. The enzyme DPP-4 has been found to play a significant role.
{{TOC limit|3}}

==History==
Since its discovery in 1967, serine protease DPP-4 has been a popular subject of research. Inhibitors of DPP-4 have long been sought as tools to elucidate the functional significance of the [[enzyme]]. The first inhibitors were characterized in the late 1980s and 1990s. Each [[Enzyme inhibitor|inhibitor]] was important to establish an early [[structure activity relationship]] (SAR) for subsequent investigation. The inhibitors fall into two main classes, those that interact covalently with DPP-4 and those that do not. [[dipeptidyl peptidase-4|DPP-4]] is a [[dipeptidase]] that selectively binds [[Substrate (biochemistry)|substrate]]s that contain [[proline]] at the P1-position, thus many [[DPP-4 inhibitor]]s have 5-membered [[heterocyclic]] rings that [[mimic]] [[proline]], e.g. [[pyrrolidine]], cyanopyrrolidine, [[thiazolidine]] and cyanothiazolidine. These compounds commonly form covalent bonds to the [[catalytic]] residue Ser630.

In 1994, researchers from Zeria Pharmaceuticals unveiled cyanopyrrolidines with a [[nitrile]] [[function group]] that was assumed to form an [[imidate]] with the [[catalytic]] [[serine]]. Concurrently other DPP-4 inhibitors without a [[nitrile]] group were published but they contained other serine-interacting motifs, e.g. [[boronic acids]], [[phosphonate]]s or diacyl [[hydroxylamine]]s. These [[Chemical compound|compound]]s were not as potent because of the similarity of [[dipeptidyl peptidase-4|DPP-4]] and [[prolyl oligopeptidase]] (PEP) and also suffered from chemical [[instability]]. [[Ferring Pharmaceuticals]] filed for patent on two cyanopyrrolidine DPP-4 inhibitors, which they published in 1995. These compounds had excellent [[potency (pharmacology)|potency]] and improved chemical stability.

In 1995, Edwin B. Villhauer at [[Novartis]] started to explore N-substituted glycinyl-cyanopyrrolidines based on the fact that DPP-4 identifies N-methylglycine as an N-terminal [[amino acid]]. This group of new cyanopyrrolidines became extremely popular field of research in the following years. Some trials with dual inhibitors of DPP-4 and vasopeptidase have been represented, since vasopeptidase inhibition is believed to enhance the antidiabetic effect of [[dipeptidyl peptidase-4 inhibitors|DPP-4 inhibition]] by stimulating [[insulin]] secretion. Vasopeptidase-inhibiting motif is connected to the [[DPP-4 inhibitor]] at the N-substituent.

==DPP-4 mechanism==
[[File:Inhibition of DPP-4 enzyme.JPG|thumb|350px]]'''Fig.1:''' During a [[meal]], the [[incretin]]s [[glucagon-like peptide 1]] (GLP-1) and glucose-dependent [[gastric inhibitory polypeptide]] (GIP) are released by the [[small intestine]] into the blood stream. These [[hormone]]s regulate insulin secretion in a glucose-dependent manner. (GLP-1 has many roles in the [[human body]]. It stimulates insulin biosynthesis, inhibits glucagon secretion, slows gastric emptying, reduces appetite and stimulates regeneration of islet [[Beta cell|β-cell]]s.)

GLP-1 and GIP have extremely short plasma [[half-lives]] due to very rapid inactivation, catalyzed by the [[enzyme]] DPP-4. Inhibition of DPP-4 slows their inactivation, thereby potentiating their action, leading to lower plasma glucose levels, hence its utility in the treatment of type 2 diabetes. (''Figure 1'').

===DPP-4 distribution and function===
[[File:Teikning2 DPP-4.JPG|thumb|220px|left|'''Fig.2:''' DPP-4 cleaves two amino acids from the N-terminal end of peptides, such as GLP-1.]]DPP-4 is attached to the [[plasma membrane]] of the endothelium of almost every organ in the [[Human body|body]]. [[Tissue (biology)|Tissue]]s which strongly express DPP-4 include the [[exocrine pancreas]], [[sweat glands]], [[salivary gland|salivary]] and [[mammary glands]], [[thymus]], [[lymph nodes]], [[biliary tract]], [[kidney]], [[liver]], [[placenta]], [[uterus]], [[prostate]], [[skin]], and the [[capillary bed]] of the [[Gut (zoology)|gut]] [[mucosa]] (where most GLP-1 is inactivated locally). It is also present, in soluble form, in [[body fluids]], such as [[blood plasma]] and [[cerebrospinal fluid]]. (It also happens that DPP-4 is the CD26 [[T-cell]] activating [[antigen]].)

DPP-4 selectively cleaves two [[amino acid]]s from [[peptide]]s, such as GLP-1 and GIP, which have [[proline]] or [[alanine]] in the second position (''Figure 2''). At the [[active site]] where DPP-4 has its effect, there is a characteristic arrangement of three [[amino acid]]s, Asp-His-Ser. Since [[alanine]] and [[proline]] are crucial for the [[biological activity]] of GPL-1 and GIP, they are inactivated by cleaving away these [[amino acid]]s. Thus, preventing the degradation of the [[incretin]] [[hormone]]s GLP-1 and GIP by inhibition of DPP-4 has potential as a [[therapeutic]] strategy in the treatment of type 2 diabetes.

==DPP-4 characteristics==
Since DPP-4 is a [[protease]], it is not unexpected that inhibitors would likely have a [[peptide]] nature and this theme has carried through to contemporary research.

===Structure===

[[X-ray]] structures of [[dipeptidyl peptidase-4|DPP-4]] that have been published since 2003 give rather detailed information about the structural characteristics of the [[binding site]]. Many structurally diverse [[DPP-4 inhibitor]]s have been discovered and it is not that surprising considering the properties of the binding site:

1. A deep [[lipophilic]] pocket combined with several exposed [[aromatic]] [[side chain]]s for achieving high affinity small molecule binding.

2. A significant [[solvent]] access that makes it possible to tune the [[physico-chemical]] properties of the inhibitors that leads to better [[pharmacokinetic]] behavior.

DPP-4 is a 766-amino acid [[transmembrane protein|transmembrane glycoprotein]] that belongs to the [[Prolyl endopeptidase|prolyloligopeptidase]] family. It consists of three parts; a [[cytoplasmic]] tail, a transmembrane region and an [[extracellular]] part. The [[extracellular]] part is divided into a [[catalytic domain]] and an eight-bladed β-propeller domain. The latter contributes to the inhibitor binding site. The catalytic domain shows an α/β-hydrolase fold and contains the catalytic triad Ser630 - Asp708 - His740. The S1-pocket is very [[hydrophobic]] and is composed of the side chains: Tyr631, Val656, Trp662, Tyr666 and Val711. Existing [[X-ray]] structures show that there is not much difference in size and shape of the pocket that indicates that the S1-pocket has high specificity for [[proline]] residues

===Binding site===

[[File:The inhibitor ligand and the DPP-4 binding site.png|thumb|600px|'''Fig.3:''' The key interactions between the ligand and DPP-4 complex. The ligand's basic amine forms a hydrogen bonding network. The nitrile reacts with the catalytic active serine and forms an imidate adduct]]
[[DPP-4 inhibitors]] usually have an electrophilic group that can interact with the [[hydroxyl]] of the catalytic serine in the active binding site (''Figure 3''). Frequently that group is a [[nitrile]] group but can also be [[boronic acid]] or diphenyl [[phosphonate]]. This electrophilic group can bind to the [[imidate]] [[Multiprotein complex|complex]] with [[covalent bonds]] and slow, tight-binding kinetics but this group is also responsible for stability issues due to [[Chemical reaction|reaction]]s with the free amino group of the P2-amino acid. Therefore, inhibitors without the electrophilic group have also been developed, but these [[molecule]]s have shown [[toxicity]] due to affinity to other dipeptidyl peptidases, e.g. DPP-2, [[DPP8|DPP-8]] and [[DPP9|DPP-9]].

[[DPP-4 inhibitor]]s span diverse structural types. In 2007 few of the most [[potency (pharmacology)|potent]] compounds contain a [[proline]] [[mimetic]] cyanopyrrolidine P1 group. This group enhances the potency, probably due to a transient [[covalent]] trapping of the [[nitrile]] group by the active site Ser630 hydroxyl, leading to delayed dissociation and slow tight binding of certain inhibitors. When these potency enhancements were achieved, some chemical stability issues were noted and more advanced molecules had to be made. To avoid these stability issues, the possibility to exclude the [[nitrile]] group was investigated. Amino acids with [[aryl]] or [[Chemical polarity|polar]] [[side chains]] did not show appreciable [[dipeptidyl peptidase-4 inhibitors|DPP-4 inhibition]] and in fact, all compounds without the nitrile group in this research suffered a 20 to 50-fold loss of potency corresponding to the compounds containing the [[nitrile]] group.

==Discovery and development==

It is important to find a fast and accurate system to discover new [[DPP-4 inhibitor]]s with ideal [[therapeutic]] profiles. [[High throughput screening]] (HTS) usually gives low hit rates in identifying the inhibitors but [[virtual screening]] (VS) can give higher rates. [[virtual screening|VS]] has for example been used to screen for small primary [[aliphatic]] amines to identify [[wikt:part|fragments]] that could be placed in S1 and S2 sites of [[dipeptidyl peptidase-4|DPP-4]]. On the other hand, these fragments were not very potent and therefore identified as a starting point to design better ones.
[[3D modeling|Three-dimensional models]] can provide a useful tool for designing novel [[DPP-4 inhibitor]]s. [[Pharmacophore]] models have been made based on key [[chemical]] features of [[Chemical compound|compound]]s with [[DPP-4 inhibitor]]y activity. These models can provide a hypothetical picture of the primary [[chemical]] feature responsible for inhibitory activity.
The first DPP-4 inhibitors were reversible inhibitors and came with bad [[adverse effect|side effects]] because of low selectivity. Researchers suspected that inhibitors with short [[half-lives]] would be preferred in order to minimize possible [[adverse effect|side effects]]. However, since [[clinical trials]] showed the opposite, the latest DPP-4 inhibitors have a long-lasting effect. One of the first reported DPP-4 inhibitor was P32/98 from [[Merck & Co.|Merck]]. It used thiazolidide as the P1-substitute and was the first DPP-4 inhibitor that showed effects in both [[animal]]s and [[human]]s but it was not developed to a market [[drug]] due to [[adverse effect|side effects]]. Another old inhibitor is DPP-728 from [[Novartis]], where 2-cyanopyrrolidine is used as the P1-substitute. The addition of the cyano group generally increases the potency. Therefore, researchers' attention was directed to those compounds. Usually, DPP-4 inhibitors are either [[Substrate (biochemistry)|substrate]]-like or non-substrate-like.
[[File:Substrate-like inhibitors and binding to the DPP-4 complex.png|thumb|250px| '''Fig.4:''' A generic structure of a substrate-like inhibitor]]

===Substrate-like inhibitors===

Substrate-like inhibitors (''Figure 4'') are more common than the non-substrate-likes. They bind either [[covalently]] or non-covalently and have a basic structure where the P1-substituent occupies the S1-pocket and the P2-substituent occupies the S2-pocket. Usually they contain a [[proline]] [[mimetic]] that occupies the S1-pocket. Large substituents on the 2-cyanopyrrolidine ring are normally not tolerated since the S1-pocket is quite small.
Since [[dipeptidyl peptidase-4|DPP-4]] is identical with the [[T-cell]] activation marker [[CD26]] and [[DPP-4 inhibitor]]s are known to inhibit T-cell [[cell growth|proliferation]], these compounds were initially thought to be potential [[immunomodulator]]s. When the function against [[type 2 diabetes]] was discovered, the cyanopyrrolidines became a highly popular research material. A little later [[vildagliptin]] and [[saxagliptin]], which are the most developed cyanopyrrolidine [[DPP-4 inhibitor]]s to date, were discovered.

====Cyanopyrrolidines====

Cyanopyrrolidines have two key interactions to the DPP-4 complex:

1. [[Nitrile]] in the position of the [[scissile bond]] of the peptidic substrate that is important for high potency. The nitrile group forms reversible [[covalent bonds]] with the catalytically active [[serine]] hydroxyl (Ser630), i.e. cyanopyrrolidines are competitive inhibitors with slow dissociation kinetics.

2. [[Hydrogen bond]]ing network between the protonated amino group and a negatively [[electric charge|charged]] region of the [[protein]] surface, Glu205, Glu206 and Tyr662. All cyanopyrrolidines have basic, primary or secondary [[amine]], which makes this network possible but these compounds usually drop in potency if these amines are changed. Nonetheless, two patent applications unveil that the amino group can be changed, i.e. replaced by a [[hydrazine]], but it is claimed that these compounds do not only act ''via'' DPP-4 inhibition but also prevent diabetic vascular complications by acting as a [[Radical (chemistry)|radical]] scavenger.

=====Structure-activity relationship (SAR)=====

Important [[structure-activity relationship]]:

1. Strict steric constraint exists around the [[pyrrolidine]] ring of cyanopyrrolidine-based inhibitors, with only [[hydrogen]], [[fluoro]], [[acetylene]], [[nitrile]], or methano substitution permitted.

2. Presence of a [[nitrile]] moiety on the [[pyrrolidine]] ring is critical to achieving potent activity

Also, systematic SAR investigation has shown that the ring size and [[stereochemistry]] for the P2 position is quite conditioned. A 5-membered ring and L-configuration has shown better results than a 4-membered or 6-membered ring with D-configuration. Only minor changes on the [[pyrrolidine]] ring can be tolerated, since the good fit of the ring with the [[hydrophobic]] S1 pocket is very important for high affinity. Some trials have been made, e.g. by replacing the [[pyrrolidine]] with a [[thiazolidine|thiazoline]]. That led to improved potency but also loss of chemical stability. Efforts to improve chemical stability often led to loss of specificity because of interactions with [[DPP8|DPP-8]] and [[DPP9|DPP-9]]. These [[Drug interaction|interaction]]s have been connected with increased [[toxicity]] and [[Mortality rate|mortality]] in animals. There are strict limitations in the P1 position and hardly any changes are tolerated. On the other hand, a variety of changes can be made in the P2 position. In fact, [[substitution reaction|substitution]] with quite big branched side chains, e.g. ''tert''-butylglycin, normally increased activity and chemical stability, which could lead to longer-lasting inhibition of the DPP-4 enzyme. It has also been noted that [[biaryl]]-based [[side chains]] can also give highly active inhibitors. It was originally believed that only [[lipophilic]] substitution would be tolerated. Now it is stated that also the substitution of [[Chemical polarity|polar]] negatively charged side-chains as well as [[hydrophilic]] substitution can lead to excellent inhibitory activity.

=====Chemical stability=====

[[File:Intramolecular cyclization of cis-rotamer.svg|thumb|330px|left|'''Fig.5:''' ''Trans''-rotamers are more stable then ''cis''-rotamers. ''Cis''-rotamers undergo intramolecular cyclization.]]

In general, [[DPP-4 inhibitor]]s are not very stable compounds. Therefore, many researchers focus on enhancing the stability for cyanopyrrolidines. The most widespread technique to improve chemical stability is to incorporate a [[steric bulk]]. The two cyanopyrrolidines that have been most pronounced, [[vildagliptin]] and [[saxagliptin]], were created in this manner. K579 is a [[DPP-4 inhibitor]] discovered by researchers at Kyowa Hakko Kyogo. It had improved not only chemical stability but also a longer-lasting action. That long-lasting action was most likely due to slow dissociation of the enzyme-inhibitor complex and an active [[oxide]] [[metabolite]] that undergoes enterohepatic circulation. The discovery of the active [[oxide]] was in fact a big breakthrough as it led to the development of [[vildagliptin]] and [[saxagliptin]]. One major problem in [[DPP-4 inhibitor]] stability is [[Intramolecular reaction|intramolecular]] cyclization. The precondition for the intramolecular cyclization is the conversion of the ''trans''-[[rotamer]], which is the DPP-4 binding [[rotamer]] (''Figure 5''). Thus, preventing this conversion will increase stability. This prevention was successful when incorporating an amide group into a ring, creating a compound that kept the DPP-4 inhibitory activity that, did not undergo the intramolecular cyclization and was even more selective over different DPP enzymes. It has also been reported that a cyanoazetidine in the P1 position and a β-amino acid in the P2 position increased stability.

=====Vildagliptin=====

[[Vildagliptin]] (Galvus)(''Figure 6'') was first synthesized in May 1998 and was named after Edwin B. Villhauer. It was discovered when researchers at [[Novartis]] examined adamantyl [[Derivative (chemistry)|derivative]]s that had proven to be very [[potency (pharmacology)|potent]]. The [[adamantyl]] group worked as a [[steric bulk]] and slowed intramolecular cyclization while increasing chemical stability. Furthermore, the primary [[metabolite]]s were highly active. To avoid additional [[Chiral (chemistry)|chiral]] center a [[hydroxylation]] at the [[adamantyl]] ring was carried out (''Figure 6''). The product, [[vildagliptin]], was even more stable, undergoing intramolecular cyclization 30-times slower, and having high [[DPP-4 inhibitor]]y activity and longer-lasting [[pharmacodynamic]] effect.

=====Saxagliptin=====

[[File:Structure of cyanopyrrolidine DPP-4 inhibitors.svg|thumb|450px|'''Fig.6:''' The basic structure of cyanopyrrolidines compared with vildagliptin, saxagliptin, and denagliptin]]
Researchers at [[Bristol-Myers Squibb]] found that increased [[steric bulk]] of the ''N''-terminal [[amino acid]] [[Side chain|side-chain]] led to increased stability. To additionally increase stability the ''trans''-[[rotamer]] was stabilized with a ''cis''-4,5-methano substitution of the [[pyrrolidine]] ring, resulting in an [[Intramolecular force|intramolecular]] [[van der Waals force|van-der-Waals]] interaction, thus preventing [[Intramolecular reaction|intramolecular]] cyclisation. Because of that increased stability, the researchers continued their investigation on ''cis''-4,5-methano cyanopyrrolidines and came across with a new [[adamantyl]] [[Derivative (chemistry)|derivative]], which showed extraordinary ''ex vivo'' DPP-4 inhibition in rat plasma. Also noted, high [[microsomal]] turnover rate which indicated that the derivative was quickly converted to an active [[metabolite]]. After [[hydroxylation]] on the [[adamantyl]] group they had a product with better [[microsomal]] stability and improved chemical stability. That product was named [[saxagliptin]] (Onglyza) (''Figure 6''). In June 2008 [[AstraZeneca]] and [[Bristol-Myers Squibb]] submitted a new drug application for Onglyza in the [[United States]] and a marketing authorization application in [[Europe]]. Approval was granted in the [[United States]] by the FDA in July 2009 for Onglyza 5&nbsp;mg and Onglyza 2.5&nbsp;mg. This was later combined with extended-release [[metformin]] (taken once daily) and approved by the FDA in January 2011 under the trade name Kombiglyze XR.

=====Denagliptin=====

Denagliptin (''Figure 6'') is an advanced [[Chemical compound|compound]] with a branched [[Side chain|side-chain]] at the P2 position, but also has (4''S'')-fluoro substitution on the cyanopyrrolidine ring. It is a well-known [[DPP-4 inhibitor]] developed by [[GlaxoSmithKline]] (GSK). Biological evaluations have shown that the ''S''-configuration of the [[amino acid]] portion is essential for the inhibitory activity since the ''R''-configuration showed reluctantly inhibition. These findings will be useful in future designs and synthesis of [[DPP-4 inhibitor]]s. GSK suspended Phase III clinical trials in October 2008.

====Azetidine based compounds====

Informations for this group of inhibitors are quite restricted. [[Azetidine]]-based [[DPP-4 inhibitor]]s can roughly be grouped into three main subcategories: 2-cyanoazetidines, 3-fluoroazetidines, and 2-ketoazetidines. The most potent ketoazetidines and cyanoazetidines have large [[hydrophobic]] [[amino acid]] groups bound to the [[azetidine]] [[nitrogen]] and are active below 100nM.

===Non-substrate-like inhibitors===

Non-substrate-like [[Enzyme inhibitor|inhibitors]] do not take after dipeptidic nature of DPP-4 substrates. They are [[Enzyme inhibitor#Reversible inhibitors|non-covalent inhibitors]] and usually have an [[aromatic ring]] that occupies the S1-pocket, instead of the proline mimetic.

In 1999, [[Merck & Co.|Merck]] started a [[drug development]] program on DPP-4 inhibitors. When they started internal screening and [[medicinal chemistry]] program, two [[DPP-4 inhibitor]]s were already in [[clinical trials]], isoleucyl thiazolidide (P32/38) and NVP-DPP728 from [[Novartis]]. [[Merck & Co.|Merck]] in-licensed L-''threo''-isoleucyl [[thiazolidine|thiazolidide]] and its ''allo'' [[stereoisomer]]. In animal studies, they found that both [[isomers]] had similar affinity for DPP-4, similar ''in vivo'' efficacy, similar [[pharmacokinetic]] and metabolic profiles. Nevertheless, the ''allo'' isomer was 10-fold more [[toxic]]. The researchers found out that this difference in [[toxicity]] was due to the ''allo'' isomer's greater inhibition of [[DPP8|DPP-8]] and [[DPP9|DPP-9]] but not because of selective DPP-4 inhibition. More research also supported that DPP-4 inhibition would not cause compromised immune function. Once this link between affinity for DPP-8/DPP-9 and [[toxicity]] was discovered, [[Merck & Co.|Merck]] decided on identifying an inhibitor with more than a thousandfold affinity for DPP-4 over the other dipeptidases. For this purpose, they used [[positional scanning libraries]]. From scanning these libraries, the researchers discovered that both DPP-4 and [[DPP8|DPP-8]] showed a strong preference for breaking down [[peptide]]s with a [[proline]] at the P1 position but they found a great difference at the P2 site; i.e., they found that acidic functionality at the P2 position could provide a greater affinity for DPP-4 over [[DPP8|DPP-8]]. [[Merck & Co.|Merck]] kept up doing even more research and screening. They stopped working on compounds from the α-amino acid series related to isoleucyl [[thiazolidine|thiazolidide]] due to lack of selectivity but instead they discovered a very selective β-[[amino acid]] [[piperazine]] series through [[structure-activity relationship|SAR]] studies on two screening leads. When trying to stabilize the [[piperazine]] [[moiety (chemistry)|moiety]], a group of [[bicyclic]] derivatives were made, which led to the identification of a [[potency (pharmacology)|potent]] and selective triazolopiperazine series. Most of these [[Structural analog|analogs]] showed excellent [[pharmacokinetic]] properties in [[preclinical]] species. Optimization of these compounds finally led to the discovery of [[sitagliptin]].

====Sitagliptin====

[[File:Sitagliptin.svg|thumb|250px|right|'''Fig.7:''' The structure of sitagliptin]]
[[Sitagliptin]] (Januvia) has a novel structure with β-amino amide [[Derivative (chemistry)|derivatives]] (''Figure 7''). Since [[sitagliptin]] has shown excellent selectivity and ''in vivo'' efficacy it urged researchers to inspect the new structure of [[DPP-4 inhibitor]]s with appended β-[[amino acid]] moiety. Further studies are being developed to optimize these compounds for the treatment of [[diabetes]].
In October 2006 sitagliptin became the first DPP-4 inhibitor that got FDA approval for the treatment of [[type 2 diabetes]]. [[Crystallographic]] structure of [[sitagliptin]] along with [[molecular modeling]] has been used to continue the search for structurally diverse inhibitors. A new [[Potency (pharmacology)|potent]], selective and orally [[bioavailable]] DPP-4 inhibitor was discovered by replacing the central [[cyclohexylamine]] in [[sitagliptin]] with 3-aminopiperidine. A 2-pyridyl substitution was the initial SAR breakthrough since that group plays a significant role in potency and selectivity for DPP-4.

It has been shown with an [[X-ray]] [[crystallography]] how [[sitagliptin]] binds to the DPP-4 complex:

1. The trifluorophenyl group occupies the S1-pocket

2. The trifluoromethyl group interacts with the side chains of residues Arg358 and Ser209.

3. The [[amino group]] forms a [[salt bridge]] with Tyr662 and the carboxylated groups of the two glutamate residues, Glu205 and Glu206.

4. The triazolopiperazine group collides with the [[phenyl group]] of residue Phe357

[[File:ABT-341.svg|thumb|250px|right|'''Fig.8:''' The structure of ABT-341]]

====Constrained phenylethylamine compounds====

Researchers at [[Abbott Laboratories]] identified three novel series of DPP-4 inhibitors using HTS. After more research and optimization ABT-341 was discovered (''Figure 8''). It is a potent and selective DPP-4 inhibitor with a 2D-structure very similar to [[sitagliptin]]. However, the 3D-structure is quite different. ABT-341 also has a trifluorophenyl group that occupies the S1-pocket and the free [[amino group]], but the two carbonyl groups are orientated 180° away from each other. ABT-341 is also believed to interact with the Tyr547, probably because of [[steric hindrance]] between the cyclohexenyl ring and the [[tyrosine]] side-chain. [[Omarigliptin]] is one of such compound which is in Phase-III development by [[Merck & Co]].

====Pyrrolidine compounds====

The [[pyrrolidine]] type of [[DPP-4 inhibitor]]s was first discovered after [[High throughput screening|HTS]]. Research showed that the [[pyrrolidine]] rings were the part of the compounds that fit into the [[binding site]]. Further development has led to [[fluoro]] substituted pyrrolidines that show superior activity, as well as [[pyrrolidine]]s with fused cyclopropylrings that are highly active.

===Xanthine-based compounds===

This is a different class of inhibitors that was identified with HTS. [[Aromatic]] [[heterocyclic]]-based [[DPP-4 inhibitor]]s have gained increased attention recently. The first patents describing [[xanthine]]s (''Figure 10'') as [[DPP-4 inhibitor]]s came from [[Boehringer-Ingelheim]](BI) and [[Novo Nordisk]].
When [[xanthine]] based DPP-4 inhibitors are compared with [[sitagliptin]] and [[vildagliptin]] it has shown a superior profile. [[Xanthines]] are believed to have higher [[potency (pharmacology)|potency]], longer-lasting inhibition and longer-lasting improvement of [[glucose tolerance]].

====Alogliptin====
[[File:Quinazolinone based structure developed to alogliptin.png|thumb|350px|'''Fig.9:''' Quinazolinone structure and alogliptin]]
[[Alogliptin]] (''Figure 9'') is a novel [[DPP-4 inhibitor]] developed by the [[Takeda Pharmaceutical Company]]. Researchers hypothesized that a [[quinazolinone]] based structure (''Figure 9'') would have the necessary groups to interact with the [[active site]] on the DPP-4 [[Multiprotein complex|complex]]. [[Quinazolinone]] based compounds interacted effectively with the DPP-4 complex, but suffered from low metabolic [[half-life]]. It was found that when replacing the [[quinazolinone]] with a pyrimidinedione, the metabolic stability was increased and the result was a [[Potency (pharmacology)|potent]], selective, [[bioavailable]] [[DPP-4 inhibitor]] named [[alogliptin]]. The quinazoline based compounds showed potent inhibition and excellent selectivity over related [[protease]], [[DPP8|DPP-8]]. However, short [[metabolic]] [[half-life]] due to [[oxidation]] of the A-ring [[phenyl]] group was problematic. At first, the researchers tried to make a [[fluorinated]] derivative. The derivative showed improved metabolic stability and excellent inhibition of the DPP-4 enzyme. However, it was also found to inhibit [[CYP 450]] 3A4 and block the [[hERG]] channel. The solution to this problem was to replace the [[quinazolinone]] with other [[heterocycle]]s, but the [[quinazolinone]] could be replaced without any loss of DPP-4 inhibition. [[Alogliptin]] was discovered when quinazolinone was replaced with a [[pyrimidinedione]]. [[Alogliptin]] has shown excellent inhibition of DPP-4 and extraordinary selectivity, greater than 10.000 fold over the closely related serine proteases [[DPP8|DPP-8]] and [[DPP9|DPP-9]]. Also, it does not inhibit the [[CYP 450]] enzymes nor block the [[hERG]] channel at concentration up to 30 µM. Based on this data, [[alogliptin]] was chosen for preclinical evaluation. In January 2007 [[alogliptin]] was undergoing the phase III [[clinical trial]] and in October 2008 it was being reviewed by the [[U.S. Food and Drug Administration]].

====Linagliptin====
[[File:Structure of xanthine type DPP-4 inhibitors.svg|thumb|250px|right|'''Fig.10:''' The structure of xanthine type inhibitors '''(TOP)''' and linagliptin '''(BOTTOM)''']]

Researchers at BI discovered that using a buty-2-nyl group resulted in a potent candidate, called BI-1356 (''Figure 10''). In 2008 BI-1356 was undergoing [[Phases of clinical research#Phase III|phase III clinical trials]]; it was released as [[linagliptin]] in May 2011. [[X-ray crystallography]] has shown that that [[xanthine]] type binds the DPP-4 complex in a different way than other [[Enzyme inhibitor|inhibitors]]:

1. The amino group also interacts with the Glu205, Glu206 and Tyr662

2. The buty-2-nyl group occupies the S1-pocket

3. The uracil group undergoes a π-stacking interaction with the Tyr547 residue

4. The quinazoline group undergoes a π-stacking interaction with the Trp629 residue

==Pharmacology==

{| class="wikitable" align="center"
|+ Comparative pharmacology of sitagliptin and vildagliptin.
|-
! Drug !!Absorption !! Bioavailability (%) !!IC<sub>50</sub> (nM) !!Mean volume of distribution (L) !!Protein binding (%) !!Half-life (hours,100&nbsp;mg dose) !!Metabolism !!Excretion
|-
| '''Sitagliptin''' ||Rapidly absorbed with peak concentration at 1–4 hours ||align="center"|87 ||align="center"|18 ||align="center"|198 ||align="center"|38 ||align="center"|12.4 ||Small fraction undergoes hepatic metabolism ''via'' CYP 450 3A4 and 2C8 ||Excreted in an unchanged form in the urine (79%)
|-
| '''Vildagliptin''' ||Rapidly absorbed with peak concentration at 1–2 hours ||align="center"|85 ||align="center"|3 ||align="center"|70.5 ||align="center"|9.3 ||1.68 (once a day) and 2.54 (twice a day) ||Hydrolysis resulting in a pharmacologically inactive metabolite. A fraction (22%) is also excreted unchanged by the kidneys ||Excretion of the metabolite is carried out through urine (85%) and feces (15%)
|}

The [[pharmacokinetic]] properties of sitagliptin and vildagliptin appear unaffected by age, sex or [[Body mass index|BMI]]. [[Clinical research]]es have shown that [[sitagliptin]] and [[vildagliptin]] do not have the [[adverse effect|side effects]] that tend to follow [[type 2 diabetes]] treatment, e.g. [[weight gain]] and [[hyperglycemia]], but however, other side effects have been observed, including [[upper respiratory tract]] [[infection]]s, [[sore throat]] and [[diarrhea]].

==See also==
* [[Dipeptidyl peptidase-4]]
* [[Dipeptidyl peptidase-4 inhibitors]]
* [[Linagliptin]]
* [[Vildagliptin]]
* [[Sitagliptin]]
* [[Saxagliptin]]
* [[Berberine]]
* [[teneligliptin]]
* [[gosogliptin]]

{{Drug design}}

{{二次利用|date=8 August 2023}}
{{DEFAULTSORT:Development Of Dipeptidyl Peptidase-4 Inhibitors}}
[[Category:Dipeptidyl peptidase-4 inhibitors]]
[[Category:Drug discovery|Dipeptidyl peptidase-4 inhibitors]]