Perspectives in Diabetes
`Inhibition of the Activity of Dipeptidyl-Peptidase IV
`as a Deatment for Q p e 2 Diabetes
`
`Jens J. Holst and Carolyn F. Deacon
`
`The insulinotropic hormone, glucagon-like peptide 1
`(GLP-I), which has been proposed as a new treatment for
`type 2 diabetes, is metabolized extremely rapidly by the
`ubiquitous enzyme, dipeptidyl peptidase IV (DPP-IV),
`resulting in the formation of a metabolite, which may act
`as an antagonist at the GLP-1 receptor. Because of this,
`the effects of single injections of GLP-1 are short-lasting,
`and for full demonstration of its antidiabetogenic
`effects, continuous intravenous infusion is required. To
`exploit the therapeutic potential of GLP-1 clinically, we
`here propose the use of specific inhibitors of DPP-IV.
`We have demonstrated that the administration of such
`inhibitors may completely protect exogenous GLP-1
`from DPP-IV-mediated degradation, thereby greatly
`enhancing its insulinotropic effect, and provided evi-
`dence that endogenous GLP-l may be equally pro-
`tected. Preliminary studies by others in glucose-intol-
`erant experimental animals have shown that DPP-IV
`inhibition greatly ameliorates the condition. GLP-1 has
`multifaceted actions, which include stimulation of
`insulin gene expression, trophic effects on the p-cells,
`inhibition of glucagon secretion, promotion of satiety,
`inhibition of food intake, and slowing of gastric empty-
`ing, all of which contribute to normalizing elevated glu-
`cose levels. Because of this, we predict that inhibition of
`DPP-IV, which will elevate the levels of active GLP-1 and
`reduce the levels of the antagonistic metabolite, may be
`useful to treat impaired glucose tolerance and perhaps
`prevent transition to type 2 diabetes. The actions of
`DPP-IV, other than degradation of GLP-1, particularly in
`the immune system are discussed, but it is concluded that
`side effects of inhibition therapy are likely to be mild.
`Thus, DPP-IV inhibition may be an effective supplement
`to diet and exercise treatment in attempts to prevent the
`deterioration of glucose metabolism associated with the
`Western lifestyle. Diabetes 47:1663-1670, 1998
`
`From the Department of Medical Physiology, University of Copenhagen,
`Copenhagen, Denmark.
`Address correspondence and reprint requests to J.J. Holst, MD, Depart-
`ment of Medical Physiology, the Panum Institute. Blegdamsvej 3, DK-2200
`Copenhagen N, Denmark. E-mail: holst@mfi.ku.dk.
`Received for publication 19 May 1998 and accepted in revised form 16
`July 1999.
`DPP, dipeptidyl peptidase; GIP, gastric inhibitory polypeptide; GLP,
`glucagon-like peptide; GRH, growth hormone-releasing hormone; NPY,
`neuropeptide Y; PP, pancreatic polypeptide; PYY, peptide YY.
`
`DIABETES, VOL. 47, NOVEMBER 1998
`
`I n a study of the susceptibility of a number of regulatory
`
`peptides to the amino-dipeptidase activity of the
`enzyme, dipeptidyl peptidase-N (DPP-TV), Mentlein
`et al. (1) in 1993 found the two incretin hormones,
`glucagon-like peptide 1 (GLP-1) and gastric inhibitory
`polypeptide (GIP), to be substrates for this enzyme. At about
`the same time, in a published abstract, Buckley and
`Lundquist described degradation of GLP-1 by plasma and
`the resulting formation of an inactive metabolite truncated by
`the two NH,-terminal residues (2). The latter authors also
`mentioned the use of bacitracin to prevent the degradation
`and the finding that analogs substituted with D-amino acids
`in positions 7 and 8 were resistant to the degradation. In our
`laboratory, the importance of these findings was quickly real-
`ized, and a thorough investigation of DPP-IV-mediated
`metabolism of GLP-1 in humans was initiated. We found that
`endogenous as well as exogenous GLP-1 was extensively
`degraded to the metabolite GLP-l(9-36) amide (3). An
`abstract by Grandt et al. (4) and subsequent studies by
`Bjerre-Knudsen and Pridal (5) and our own group (6) sug-
`gested that GLP-l(9-36) amide might, in fact, act as a GLP-1
`receptor antagonist,, acting not only on the pancreatic GLP-
`1 receptor, but also antagonizing the gastrointestinal effects
`of GLP-1 (6). In further studies (7), we demonstrated the
`extensive degradation of exogenous GLP-1 given intra-
`venously or subcutaneously to patients with type 2 diabetes
`as well as control subjects. The degradation was particularly
`dramatic for subcutaneously administered GLP-1 (up to 90%;
`Fig. 1). Not only did this observation explain the ineffective-
`ness (short duration of action) of subcutaneously mected
`GLP-1 to normalize blood glucose in patients with type 2 dia-
`betes (8), but it also formed the basis for the proposition (7)
`that "inhibition (of DPP-IV) may prove useful.. .in the man-
`agement of type 2 diabetes, as has been the case for the
`development of angiotensin-converting enzyme inhibitors to
`treat hypertension and the suggested use of neutral endopep
`tidase inhibitors to enhance endogenous atrial natriuretic
`peptide activity in the treatment of heart failure. Intubition of
`GLP-l(7-36) amide degradation would not only increase the
`availability of the biologically active peptide but would also
`reduce the effect of feedback antagonism at the level of the
`receptor." The use of GLP-1 analogs resistant to NH,-termi-
`nal degradation was also suggested (7).
`In further studies, we investigated the metabolism of GLP-
`1 in vivo in pigs and found that the half-life of the conversion
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`Time (min)
`
`FIG. 1. Increase in plasma concentrations of total (0) and intact ( 0 ) GLP-1 after subtraction of endogenous levels and after subcutaneous
`administration of GLP-l(7-36) amide (1.5 nmollkg) in type 2 diabetic subjects (n = 8). *P c 0.05; **P c 0.001.
`
`of intact GLP-1 to its NH,-terminally truncated metabolite was
`between 1 and 1.5 min (9) (whereas the half-life on incuba-
`tion with plasma was 2030 min). In fact, the clearance of
`GLP-1 exceeded cardiac output by a factor of 2. Thus, the pep
`tide may be degraded before it reaches its presumed target
`organs. The metabolite, in turn, was eliminated from the cir-
`culation with a half-life of 4-5 rnin, mainly as a result of renal
`extraction. This value corresponds closely to the half-lives
`determined in previous investigations of GLP-1 metabolism,
`in which the NH,-terminal metabolism was disregarded (10).
`We next demonstrated that analogs substituted at position 8
`of GLP-l(7-36) amide (corresponding to position 2 of the
`actual peptide) could be designed to be largely resistant to the
`actions of DPP-IV, but retained a high affinity for the GLP-1
`receptor; such analogs may therefore be useful clinically
`(11). We, finally, investigated in anesthetized pigs, the effects
`of inhibition of DPP-IV on the effects of infused GLP-1 using
`the specific inhibitor, val-pyrrolidide (12). With this inhibitor,
`it was possible to completely inhibit the NH,-terminal degra-
`dation of GLP-1 (Fig. 2); simultaneously, the insulin response
`to glucose was greatly augmented. It was concluded that
`DPP-IV inhibition may "be a viable approach to the manage-
`ment of diabetes" (12). A number of recently published pre-
`liminary reports support this conclusion: Pauly et al. (13)
`reported that inhibition of DPP-IV by Ile-thiazolidide in rats
`augmented insulin responses to glucose and enhanced glu-
`cose clearance; Balkan and colleagues (14,15) made similar
`findings using a Sandoz inhibitor, SDZ 272-070, and also
`showed improved glucose tolerance in obese Zucker rats
`and in rats rendered insulin resistant by fat-enriched diets.
`
`The purpose of this review is to discuss the potential clin-
`ical use of DPP-IV inhibition in view of the evidence available
`at present. In the following 11 points, the pros and cons of this
`approach will be discussed.
`1. Side effects of DPP-IV inhibition. The side effects of DPP-
`IV inhibition are of paramount importance. DPP-IV is reported
`to act not only to degrade regulatory peptides with Pro or Ala
`in position 2 (I), but also to play an important role in the
`immune system. Thus in addition to its enzymatic actions,
`DPP-IV, as a membrane-associated molecule on the surface of
`T-cells (where it is also h o w n as CD26), has a function in the
`immune system by contributing to T-cell activation and pro-
`liferation (16). Here, its role in transduction of activation sig-
`nals is dependent on its interaction with other membrane-
`expressed antigens such as CD45 (17). Whether this function
`of DPP-IVlCD26 is dependent on its enzymatic activity has not
`yet been conclusively demonstrated. In studies using specific
`competitive and irreversible inhibitors, which block up to 95%
`of the enzymatic activity (18) or mutant CD26 molecules
`devoid of enzymatic activity (19), T-cell activation was unim-
`paired, suggesting that the enzymatic activity of the molecule
`was not required. However, another study using DPP-IV
`inhibitors indicated that the enzymatic activity was involved in
`the signal transduction cascade (20). Studies using mutant
`DPP-IVlCD26 molecules have indicated a role for the enzymatic
`activity in modulating the responsiveness of T-cells (2 l), while
`others have indicated that it is important but not essential for
`its co-stimulatory activity (22), and suggested that DPP-
`IVICD26 functions to augment the cellular responses. It there-
`fore appears that the immune functions of DPP-IV are largely
`
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`4
`
`I
`
`GLP-1 infusion
`C-------------,
`
`Glucose
`u
`
`GLP-1 infusion - Glucose
`
`u
`
`Time (min)
`
`FIG. 2. Plasma concentrations of total (0) and intact (@) GLP-1 in blood sampled from the carotid artery of anesthetized pigs during intra-
`venous infusions of GLP-l(7-36) amide (5 pmol kg1 . min-I). Intravenous glucose (0.2 glkg) was given during 21 min and 30 min of each GLP-1
`infusion; a DPP-IV inhibitor (val-pyrrolidide; 300 pnol/kg) was given at 100 min.
`
`independent of the catalytic site of its dipeptidase activity
`(23), so that blockade of this site with small molecule
`inhibitors should not compromise its immune functions. In
`agreement with this view, the already known DPP-IV
`inhibitors differ greatly with respect to their potencies as DPP-
`IV inhibitors and their effects on lymphocytes. Thus val-pyrro-
`lidide, the inhibitor employed in some of the studies men-
`tioned above, was reported to inhibit DPP-IV with an IC, (the
`concenlration causing 50% inhibition) of 6 pmol/l, whereas it
`caused <50% inhibition of mitogen-induced DNA synthesis in
`lymphocytes at the highest tested dose (5 X 104mol/I) (24). To
`further support that DPP-IV does not have vital or irreplace
`able functions, Fischer rats devoid of DPP-IV have been found
`to be completely viable and have a normal phenotype (also
`with respect to glucose tolerance [25]), and from studies of
`their T-cell activation (26), it was concluded that the pres-
`ence of DPP-IVlCD26 was unnecessary. Upregulation of com-
`pensatory mechanisms caused by the inherent lack of DPP-IV
`in these animals cannot, however, be excluded. Thus, it will be
`necessary to conduct studies of prolonged DPP-IV inhibition,
`e.g., with already available compounds (several are reported
`in the literature) to investigate this. Notably, a compound with
`a half-life of 8 days (presumably the half-life of the enzyme) has
`been described (27). This compound may be particularly suit-
`able for the study of long-term "side" effects.
`As mentioned, a number of other regulatory peptides,
`including the duodenal incretin hormone, GIP, and two mem-
`bers of the pancreatic polypeptide (PP) family, peptide YY
`(PYY) and neuropeptide Y (NPY), are also substrates for
`DPP-IV (but not PP itself) (1,29). No studies have been con-
`ducted so far in which the protection of these hormones was
`determined. In DPP-IV-deficient Fischer rats, GIP levels
`
`were reported to be reduced and pancreatic sensitivity to GIP
`decreased, perhaps as compensatory measures (25). Most
`likely, levels of intact GIP will increase on DPP-IV inhibition.
`Elevated levels of GIP may contribute to enhanced glucose
`tolerance (although presumably not in human type 2 dia-
`betes, see below), and this "side effect" therefore, must be
`considered expedient.
`On digestion with DPP-IV, the 36 amino acid peptides P W
`and NPY generate NH,-terminally truncated 3-36 metabolites
`(29). NPY is a neuropeptide and probably plays a limited role
`as a circulating peptide (see below). PYY, however, is a gut hor-
`mone, produced in the L-cells, the same cells that produce GLP-
`1. While GLP-1 stored in the Lcell almost exclusively con-
`sists of intact GLP-1 (28), about 40% of stored PYY is
`accounted for by PYY 336 (30), indicating that part of the
`truncated form found in plasma (30) is not generated by DPP-
`IV digestion in the circulation. And in contrast to the metabe
`lite of GLP-1, PYY 336 is highly active, retaining full activity
`toward the Y2 receptors, but losing its effects on Y1 receptors
`(31). Some of the peripheral actions of PYY, which are mainly
`related to its functions as one of the hormones of the "ileal
`brake mechanism" (inhibition of upper gastrointestinal func-
`tions elicited by the presence of food in the distal small i n k s
`tine [32-34]) seem to be mediated via Y2 receptors (35) but Y1
`receptors may also be involved (36). Thus, the extent to which
`inhibition of DPP-IV increases the ratio of intact-tetruncated
`PYY in the circulation is difficult to predict, but will, if it
`occurs, cause a change toward activation of more Y1 and
`fewer Y2 receptors. However, the consequences of this
`change are also difficult to predict. Presumably, PYY-medi-
`ated regulation of gastrointestinal functions will be marginally
`affected, but perhaps other, mainly Y1 receptor-regulated
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`functions such as blood flow regulation, could be affected
`(34). In initial human studies of DPP-IV inhibition, careful
`blood pressure control will be required.
`The hypothalamic peptide, growth hormone-releasing hor-
`mone (GRH), which is structurally related to GLP-1, is also a
`substrate for DPP-IV and is inactivated by digestion (37).
`However, DPP-IV-resistant analogs of GRH are rapidly
`degraded by enzymes other than DPP-IV (38), and it is uncer-
`tain whether inhibition of DPP-IV will affect the actions of
`GRH released to the pituitary portal circulation.
`2. Long-term efficacy of the compound. Indeed, the very
`fact that the DPP-IV-deficient Fischer rats (25) seem com-
`pletely unaffected might suggest that compensatory mech-
`anisms may take over in DPP-IV4eficient animals. Similarly,
`other routes of GLP-1 degradation might be uncovered or
`induced during continuing DPP-IV inhibition. Thus, first,
`will GLP-1 degradation remain inhibited on long-term
`inhibitor administration? Second, will the effects of DPP-IV
`inhibition themselves show tachyphylaxis (i.e., will there be
`tachyphylaxis to the effects of an increased level of intact
`GLP-I)?
`These questions cannot be answered presently, but must be
`investigated in long-term studies of DPP-IV inhibition. Again,
`such studies could be conducted in experimental animals
`using the available inhibitors such as val-pyrrolidide. With
`respect to tachyphylaxis to GLP-1, this important question has
`been addressed in a few studies of GLP-1 administration. In
`two studies in which GLP-1 was infused continuously for 7
`days to patients with type 2 diabetes, there was no sign of
`tachyphylaxis with respect to its effects on glucose metabo-
`lism (39,40), and in a recent study the antidiabetic effect of
`a GLP-1 analog delivered by engineered cells transplanted into
`glucose-intolerant mice was preserved for the duration of
`the experiment (1 month) (41). However, no studies have
`addressed the question directly. In addition, there is evidence
`that the gastrointestinal effects of GLP-1 (see below) may
`show tachyphylaxis (42). It must be borne in mind, however,
`that development of tachyphylaxis may depend on the
`dosage scheme. The continued presence of elevated levels of
`(active) GLP-1 might promote tachyphylaxis as opposed to
`discontinuous therapy, although the relatively small
`increases in GLP-1 levels that may be obtained by DPP-IV
`inhibition may be less prone to cause tachyphylaxis.
`3. Is it possible to protect endogenous GLP-1 from
`degradation? This essential question has not been
`addressed so far. DPP-IV-mediated degradation of GLP-1 is
`extensive, reflecting the widespread distribution of the
`enzyme, and occurs not only in plasma but also in numerous
`tissues, with, for example, the liver being one of the maor
`sites for inactivation of the circulating peptide (9). In a recent
`study, we showed that, although GLP-1 in the gut is stored
`entirely in the intact form, 50% of the newly secreted peptide
`released from isolated perfused preparations of pig ileum
`was already degraded by the time it reached the local venous
`drainage (28). This degradation could be completely pre-
`vented by intraluminal or intravascular val-pyrrolidide (28).
`In our study of administration of the same compound to pigs
`in vivo (12), we found that degradation of GLP-1, secreted in
`the basal state, was greatly reduced (determined by com-
`parison of levels of intact and total [intact + metabolite] GLP-
`1 with and without inhibitor; Fig. 2). However, the conse-
`quences for GLP-1 secretion in relation to meals have not been
`
`investigated. Our prediction is that it will be possible to pro-
`tect endogenous GLP-1 extensively from degradation.
`4. Is full protection of endogenous GLP-1 enough to
`have a significant effect in type 2 diabetes? In our orig-
`inal study (3), levels of total GLP-1 increased from - 12.6 to
`22.3 pmoYl postprandially. The levels of intact GLP-1
`increased from 3.3 to 9.9 pmoH; the difference was reason-
`ably accounted for by the concentration of the metabolite.
`With a DPP-IV inhibitor, it could be predicted that all of the
`12.6 and 22.3 pmolfl would occur as intact, biologically active
`peptide, a two- to fourfold increase. In addition, there would
`be no antagonist (9-36 amide) to antagonize the actions of
`GLP-1. In our experiments conducted in patients with type 2
`diabetes, full normalization of blood glucose levels was
`obtained during intravenous infusion of GLP-1 at a rate of 1.2
`pmol . kg1 . min-' (43). This infusion rate increases levels of
`intact GLP-1 to 15-20 pmoYl(7), and on top of this, there is
`a concentration of 80-100 pmoYl of the antagonistic metabo-
`lite. Thus, one would predict that DPP-IV inhibition might pro-
`duce plasma levels of intact GLP-1 that would be large
`enough to significantly and unopposedly affect the target
`organs for GLP-1. The effect would be largest postprandialls:
`but inspection of the 24-h profile for plasma GLP-1 (44)
`reveals that although there are clear meal-related increases,
`GLP levels remain elevated throughout the day, once the
`digestive processes are initiated by breakfast ingestion. In
`addition, there is evidence that even fasting subjects may
`have a small but significant secretion of GLP-1([3] and stud-
`ies by Toft-Nielsen et al. [45], in which it was shown that the
`basal levels could actually be significantly suppressed by
`somatostatin infusion). The preliminary studies (13-15) cited
`earlier reveal that administration of DPP-IV inhibitors to glu-
`cose-intolerant rodents does in fact improve glucose tolerance
`(but, clearly, in these studies the responsible mechanism
`could not be deduced). Our prediction is that DPP-IV inhibi-
`tion will have a significant effect.
`5. What is the rationale for treating type 2 diabetes with
`increased availability of GLP-l? Or, in other words, why
`is it that the GLP-1 these patients produce cannot help their
`p-cells keep up insulin production? The answer to this ques
`tion is twofold. First, there is now evidence that the secretion
`of GLP-1 is impaired in type 2 diabetes (M.-B. Toft-Nielsen, S.
`Madsbad, J.J.H., unpublished studies of meal-induced GLP-1
`secretion in 55 patients with type 2 diabetes). The initial meal-
`induced increase seems to be the same, but the duration of the
`increase is markedly shorter. However, the deficient response
`does not seem to be responsible for diabetes, but rather to be
`a consequence of diabetes. This is because GLP-1 secretion
`is only slightly impaired in patients with impaired glucose tol-
`erance and with a high probability for transition to overt dia-
`betes (J. Lindqvist, J. Pigon, J.J.H., S. Efendic, unpublished
`observations) (if the GLP-1 deficiency had been a primary
`cause, one would have expected a similar impairment of
`secretion in prediabetes); indeed, in identical twins discordant
`for type 2 diabetes (at the time of investigation), GLP-1 secre-
`tion was impaired in the diabetic but less so in the glucose-
`tolerant twin (47). Thus, the decreased postprandial GLP-1
`response in type 2 diabetes may aggravate the disease but does
`not cause the disease. More importantly, however, it seems
`that an important and perhaps primary defect in type 2 dia-
`betes may be an impaired incretin function (i.e., little aug-
`mentation of insulin secretion after oral as compared with
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`intravenous glucose administration [48,49]). Furthermore, in
`the (rather few) patients with type 2 diabetes so far inveshgated
`for this, all had a greatly decreased or absent insulin response
`to the "other" incretin hormone, namely, GIP from the upper
`gut (49,50). The impaired incretin function is not due to
`impaired secretion of GIF? I€ anythng, GIP levels are elevated
`in these patients (47,48), but it has been proposed that there
`may be defective expression of the GIP receptor (49). This is
`important because GIP is the "kt-in-line" incretin, because
`GIP signaling is defective, meal-induced insulin secretion is also
`defective. This cannot be overcome with endogenous or
`exogenous GIP because the patients are insensitive to GIP, but
`it may be compensated for with GLP-1 (50). Thus GLP-1 treat-
`ment may be considered a substitution therapy that restores
`the defective incretin effect. This is probably the reason why
`GLP-1 treatment in itself is effective and shows little or no
`tachyphylaxiq a mere overdosing with one of the compounds
`that take part in the regulation of blood glucose presumably
`would have activated downregulatory mechanisms. At any
`rate, the defective GIP sign-
`in type 2 diabetes provides the
`rationale for a "substitution therapy" with GLP-1.
`6. Will the antidiabetogenic effect of DPP-IV inhibition
`be similar to that of GLP-1 infused intravenously? As dis-
`cussed above, the maximum levels of intact GLP-1 achieved
`by DPP-N inhibition will correspond to those normally
`observed for "total GLP-1" (i.e., 20-30 pmoV1). On top of this
`comes absence of the antagonistic metabolite. However, it can-
`not be excluded that the higher levels of biologically active
`GLP-1 may negatively feed back on the GLP-1 cells to inhibit
`endogenous secretion (there is evidence that this may occur
`[51]). In patients with long-standing type 2 diabetes, intra-
`venous GLP-1 is remarkably effective, in the sense that it is pos
`sible to completely normalize blood glucose even in patients
`with high fasting blood glucose values and little residual
`insulin secretory capacity, but in such patients the predomi-
`nant mechanism of action is likely to be inhibition of hepatic
`glucose production, secondary to inhibition of glucagon
`secretion (43,52,53), and a similar glucose-lowering effect
`may be observed in C-peptide negative patients with type 1 dia-
`betes (54). In such type 2 patients, the glucose-lowering effect
`of GLP-1 is a slow process, proceeding with a rate constant of
`-0.27% per minute (as compared to 0.44% per minute in
`patients with greater insulin secretion) (53). These figures
`presumably reflect the fact that GLP-1 in itself has little effect
`on peripheral glucose disposal (45). Possibly, with low doses
`of GLP-1, this mechanism, although capable of lowering
`blood glucose in the fasting patients, may not be sufficient to
`also effectively dispose of dietary carbohydrates, resulting in
`inadequate blood glucose control on an average. With larger
`doses of GLP-1, it is likely that glucagon inhibition will be more
`extensive, and that gastrointestinal and satiety effects are
`more pronounced (see below). At any rate, with an infusion
`rate of 2.4 pmol . kg-' . min-', a remarkable improvement of
`glycemic control may be achieved for up to 7 days in poorly
`controlled patients (40). With DPP-N inhibition, the levels of
`intact GLP-1 that result from this infusion rate may not be
`attainable. Possibly, therefore, this therapeutic principle may
`be best applied to patients with some insulin secretory
`reserve, in whom the glucose-lowering effect results from a
`combination of reduced hepatic glucose production plus
`increased insulin-mediated glucose disposal. As noted above,
`in acute, preliminary studies, DPP-N inhibition effectively
`
`DIABETES, VOL. 47, NOVEMBER 1998
`
`improved (oral) glucose tolerance in glucose-intolerant ani-
`mals (with considerable insulin capacity [13-151). This raises
`the question of whether all of the effect is, in fact, due to pro-
`tection of endogenous GLP-1 (because oral glucose is a rather
`weak stimulus for GLP-1 secretion). A s noted above, the other
`incretin hormone, GIP, may also be protected, and its effects
`are therefore enhanced; in addition, elevated levels of the
`"ileal-brake" hormone, PYY, may dampen postprandial glucose
`excursions. However, in this context, ths lack of specificity of
`DPP-N inhibition must be considered expedient.
`7. Is the compound intrinsically safe? With this, we think
`of the toxicology of the compound (i.e., side effects apart
`from those due to DPP-N inhibition). We will have to consider
`that the patients likely to benefit from a DPP-IV inhibitor will
`have to take the drug every day for seve1-d decades of their life
`time. Any side effect will probably seriously limit the useful-
`ness of the compound. On the other hand, if a nontoxic com-
`pound can be developed, it is likely to have a vast applicabil-
`ity. In view of the fact that efficient and apparently nontoxic
`inhibitors already exist, such compounds would seem fairly
`easy to develop. In fact, a nontoxic, orally active compound
`with reasonable pharmacokinetics (see below) might actu-
`ally make one of the greatest dreams of the diabetologist to
`come true: it might prevent the transition from impaired glu-
`cose tolelance to overt type 2 diabetes. This is because elevated
`levels of active GLP-1 would be expected to restore the (mild)
`incretin deficiency (discussed above) and normalize com-
`pletely glucose levels (as shown in the first animal studies). The
`lowered glucose levels would then remove the demand on the
`P-cell thereby reducing insulin secretion, which together
`would lead to normalization of insulin sensitivity. All of these
`plus perhaps specific direct effects of GLP-1 might result in pro-
`moted growth and survival of the p-cells (55-57). It could be
`envisaged that the drug could be given to patients discovered
`by population screening in the 40-70 age-group, having bor-
`derline elevations of fasting blood glucose (perhaps as low as
`5.9 rnmoV1). Oral glucose tolerance tests may also be carried
`out on a screening basis, leading to the identification of indi-
`viduals with impaired glucose tolerance and perhaps a family
`history of type 2 diabetes. Since the treatment can be pre-
`dicted to have little effect unless blood glucose is m y elevated
`(GLP-1 has very little effect in subjects with normal glucose lev-
`els regardless of dose because its actions are glucose depen-
`dent [58]) and because it should be nontoxic, over-treatment
`would be expected to have inconspicuous negative effects
`(except perhaps for hypothetically causing an increased ten-
`dency to postprandial reactive hypoglycemia, see below). On
`the contrary, treatment might still have beneficial effects on
`body weight (see below).
`This new principle of diabetes treatment should be viewed
`under the perspective that -5W of patients with type 2 dia-
`betes have irrepamble complications at the time of diagnosis.
`It is generally assumed that the complications are due to
`long-stmding disturbances of glucose metabolism. If it were
`possible to improve glucose metabolism at an earlier stage,
`it might be possible to reduce the prevalence of complications.
`In addition, the existence of an efficient treatment of early
`impaired glucose metabolism with few or no side effects
`would be an incentive for general practitioners to try to iden-
`tlfy such cases among their patients at a much earlier stage.
`Thus with DPP-N inhibitor, it may be possible to prevent or
`delay type 2 diabetes and its complications.
`
`SAXA-DEF-00013
`
`Page 5 of 8
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`
`8. Gastrointestinal and satiating effects. GLP-1 is known
`to potently inhibit gastrointestinal motility and secretion (the
`"ileal brake" effect [33]). Thus, GLP-1 is a hormone signaling
`the presence of an abundancy of nutrients in the distal small
`intestine, and which acts to limit further transfer to and
`digestion of foodstuff in the intestine until the load already
`present has been absorbed. Possibly, these effects will be
`enhanced during chronic therapy with a DPP-IV inhibitor.
`To what extent that this is good or bad cannot be settled
`presently. Judging from experience with infusion of GLP-1 into
`diabetic patients, the gastrointestinal effects will be to even
`out meal-induced glucose excursions (39,40,59,60). GLP-1
`secretion is greatly exaggerated in patients with accelerated
`gastric emptymg (61,62), and it has been shown recently that
`the exaggerated secretion is sufficient to explain reactive
`hypoglycemia in these patients (63). Thus it must be consid-
`ered whether or not DPP-IV inhibition may be associated
`with an increased risk of reactive hypoglycemia. In insulin-
`resistant patients, the risk is likely to be small (it has proven
`difficult to bring about reactive hypoglycemia with GLP-1 in
`diabetic patients, in contrast to subjects with normal glu-
`cose tolerance and insulin sensitivity (T. Vilsb~ll, T. Krarup,
`J.J.H., unpublished observations), but in subjects with faster-
`than-average gastric emptying and normal insulin sensitivity!
`this risk must be taken into account.
`GLP-1 has also been shown to promote satiety and mildly
`reduce caloric intake in humans (64), including obese subjects
`(65). Presumably, GLP-1 functions as one of the physiologi-
`cal intestinal satiety signals (33). To what extent that the
`satiating effect is related to its effects on gastrointestinal
`motility is not known. Because of this effect, it may be pre-
`dicted that DPP-IV inhibition would reduce food intake. Of
`come, this is a desirable effect, given the fact that obesity is
`both a causative and an aggravating factor for type 2 diabetes,
`insulin resistance, and glucose intolerance. It is impossible at
`present to predict how effective DPP-IV inhibition will be in
`this respect. High doses of exogenous GLP-1 can provoke nau-
`sea and even vomiting (39,66). These doses result in levels of
`intact GLP-1 that greatly exceed those that result from DPP-
`IV inhibition. As mentioned above, the latter levels may be
`obtained with the usual "therapeutic" infusion rates of -1-1.2
`pmol . kg-' . rnin-'. Such infusion rates, even when extended
`for 7 continuous days, have never been noted to have side
`effects (39,52). I11 effects because of the satiating actions
`would, therefore, not be expected.
`9. Effects on GLP-2 secretion. Like GLP-1, glucagon-like
`peptide-2 (GLP-2) is a product of intestinal processing of
`proglucagon, and is secreted in parallel with GLP-1 (33).
`Until recently, its biological function was not known, but
`studies by Drucker et al. (67) suggest that it acts as an intesti-
`nal growth factor. Further studies revealed that both endoge
`nous and exogenous GLP-2 are substrates for DPP-IV (68).
`Accordmgly, in recent (unpublished) studies in our laboratory,
`the growth-promoting effect on the small intestinal mucosa
`of subcutaneous hjections of GLP-2 was greatly enhanced by
`the simultaneous administration of the specific DPP-IV
`inhibitor, val-pyrrolidide, and, similarly, a DPP-IV-resistant
`analog wa