E m e r g i n g T r e a t m e n t s a n d T e c h n o l o g i e s
`O R I G I N A L
`A R T I C L E
`
`Improved Glycemic Control With No
`Weight Increase in Patients With Type 2
`Diabetes After Once-Daily Treatment
`With the Long-Acting Glucagon-Like
`Peptide 1 Analog Liraglutide (NN2211)
`A 12-week, double-blind, randomized, controlled trial
`
`Downloaded from http://diabetesjournals.org/care/article-pdf/27/6/1335/645887/zdc00604001335.pdf by guest on 13 January 2024
`
`CONCLUSIONS — A once-daily dose of
`liraglutide provides efficacious glycemic con-
`trol and is not associated with weight gain.
`Adverse events with the drug are mild and
`transient, and the risk of hypoglycemia is
`negligible.
`
`Diabetes Care 27:1335–1342, 2004
`
`T ype 2 diabetes is characterized by
`
`insulin resistance and defective
`␤-cell function and is associated
`with hyperglucagonemia, increased he-
`patic glucose production, and obesity (1).
`In addition, patients with type 2 diabetes
`experience a subnormal secretion of the
`incretin hormone glucagon-like peptide 1
`(GLP-1) during meals (2,3). Sulfonyl-
`ureas, although efficient in stimulating
`insulin secretion and lowering blood glu-
`cose, pose the disadvantages of weight
`gain and risk of hypoglycemia (4). Studies
`have demonstrated that GLP-1 stimulates
`insulin secretion, inhibits glucagon secre-
`tion in a glucose-dependent manner, and
`delays gastric emptying (2,5–7). In addi-
`tion, several studies have shown GLP-1 to
`have an appetite-reducing effect (8 –10),
`and one study has demonstrated weight
`loss after treatment with GLP-1 (11).
`These mechanisms make this hormone an
`attractive candidate for the treatment of
`type 2 diabetes. However, native GLP-1
`has a very short half-life (1 min) (12), be-
`ing rapidly metabolized by the enzyme
`dipeptidyl peptidase IV (13). It has been
`shown that GLP-1 must be present con-
`tinuously in the blood stream to exert its
`actions (14).
`Liraglutide is a long-acting, acylated
`GLP-1 analog, acting as a full agonist to-
`ward the GLP-1 receptor (15). Studies in
`animals and humans have demonstrated
`promising blood glucose⫺lowering ef-
`
`1
`
`STEN MADSBAD, MD, DMSC
`2
`OLE SCHMITZ, MD, DMSC
`3
`JONAS RANSTAM, PHD
`GRETHE JAKOBSEN, MSCPHARM
`
`3
`
`4
`DAVID R. MATTHEWS, DPHIL, FRCP
`ON BEHALF OF THE NN2211-1310
`INTERNATIONAL STUDY GROUP
`
`OBJECTIVE — Liraglutide is a long-acting glucagon-like peptide 1 analog designed for once
`daily injection. This study assessed the efficacy and safety of liraglutide after 12 weeks of
`treatment in type 2 diabetic patients.
`
`RESEARCH DESIGN AND METHODS — A double-blind, randomized, parallel-
`group, placebo-controlled trial with an open-label comparator arm was conducted among 193
`outpatients with type 2 diabetes. The mean age was 56.6 years and the mean HbA1c was 7.6%
`across the treatment groups. Patients were randomly assigned to one of five fixed-dosage groups
`of liraglutide (0.045, 0.225, 0.45, 0.60, or 0.75 mg), placebo, or open-label sulfonylurea
`(glimepiride, 1– 4 mg). The primary end point was HbA1c after 12 weeks; secondary end points
`were fasting serum glucose, fasting C-peptide, fasting glucagon, fasting insulin, ␤-cell function,
`body weight, adverse events, and hypoglycemic episodes.
`
`RESULTS — A total of 190 patients were included in the intention-to-treat (ITT) analysis.
`HbA1c decreased in all but the lowest liraglutide dosage group. In the 0.75-mg liraglutide group,
`HbA1c decreased by 0.75 percentage points (P ⬍ 0.0001) and fasting glucose decreased by 1.8
`mmol/l (P ⫽ 0.0003) compared with placebo. Improvement in glycemic control was evident
`after 1 week. Body weight decreased by 1.2 kg in the 0.45-mg liraglutide group (P ⫽ 0.0184)
`compared with placebo. The proinsulin-to-insulin ratio decreased in the 0.75-mg liraglutide
`group (⫺0.18; P ⫽ 0.0244) compared with placebo. Patients treated with glimepiride had
`decreased HbA1c and fasting glucose, but slightly increased body weight. No safety issues were
`raised for liraglutide; observed adverse events were mild and transient.
`
`● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ●
`
`From the 1Hvidovre Hospital, University of Copenhagen, Denmark; the 2University Hospital of Aarhus,
`Aarhus, Denmark; 3Novo Nordisk A/S, Bagsvaerd, Denmark; and the 4Oxford Centre for Diabetes, Endo-
`crinology & Metabolism, University of Oxford, Oxford, U.K.
`Address correspondence and reprint requests to Dr. Sten Madsbad, Department of Endocrinology,
`Hvidovre Hospital, Kettegaards Alle´ 30, DK-2650, Hvidovre, Denmark. E-mail: sten.madsbad@hh.hosp.dk.
`Received for publication 21 October 2003 and accepted in revised form 7 March 2004.
`D.R.M. has received consulting fees from Novo Nordisk, and the Oxford Centre for Diabetes, Endocri-
`nology & Metabolism is supported by Novo Nordisk in an academic partnership.
`Abbreviations: GLP-1, glucagon-like peptide 1; HOMA, homeostasis model assessment; ITT, intention
`to treat; OHA, oral hypoglycemic agent.
`A table elsewhere in this issue shows conventional and Syste`me International (SI) units and conversion
`factors for many substances.
`© 2004 by the American Diabetes Association.
`
`DIABETES CARE, VOLUME 27, NUMBER 6, JUNE 2004
`
`1335
`
`Novo Nordisk Exhibit 2075
`Mylan Pharms. Inc. v. Novo Nordisk A/S
`IPR2023-00724
`Page 00001
`
`

`

`Improved glycemic control with liraglutide (NN2211)
`
`Downloaded from http://diabetesjournals.org/care/article-pdf/27/6/1335/645887/zdc00604001335.pdf by guest on 13 January 2024
`
`fects as well as a favorable safety profile.
`The half-life of liraglutide is ⬃12 h in
`both healthy subjects and type 2 diabetic
`patients after single and multiple dosing
`(16 –18). The dosing regimen is a once
`daily injection. This trial investigated the
`efficacy and safety of liraglutide after 12
`weeks of treatment in type 2 diabetic pa-
`tients.
`
`RESEARCH DESIGN AND
`METHODS —This trial was a 12-week,
`multicenter, double-blind, randomized,
`parallel-group, dosage-response trial in
`six treatment arms. In addition, an open-
`label sulfonylurea (glimepiride) arm was
`included as a reference group. Patients
`were instructed to maintain their diet, ex-
`ercise program, and daily routines during
`the course of the trial. After a 4-week
`wash-out period during which current
`oral hypoglycemic agent (OHA) treat-
`ment was discontinued, patients were
`equally randomized to double-blind
`treatment of one of six arms (one of five
`dosages of liraglutide or placebo) or to the
`open-label reference group (glimepiride).
`The trial was conducted between Decem-
`ber 2000 and October 2001 in Scandina-
`via and the U.K. The trial was conducted
`in accordance with the Helsinki Declara-
`tion (19), and the protocol was approved
`by each institution’s independent ethics
`committee. The trial was explained to all
`patients, and their written informed con-
`sent was obtained before any trial-related
`procedures were initiated.
`Eligible patients were men and
`women age ⱖ30 years who had a type 2
`diabetes diagnosis (according to Ameri-
`can Diabetes Association criteria) (20),
`had BMI ⱕ40 kg/m2, were being treated
`ⱕ
`with diet or an OHA, and had an HbA1c
`9.5% (OHA) or 7.5–10.0% (diet). For pa-
`tient safety, an upper limit for HbA1c was
`defined as a 4-week wash-out period and
`a placebo arm was included in the trial.
`Patients were excluded if any of the fol-
`lowing were present: liver or renal dis-
`ease, heart failure (New York Heart
`Association class III and IV) (21), unsta-
`ble angina pectoris, myocardial infarction
`within the previous 12 months, concom-
`itant treatment with thiazolidinediones or
`other investigational drugs, or other sig-
`nificant conditions likely to affect a pa-
`tient’s diabetes and/or ability to complete
`the trial. Women who were pregnant,
`breast-feeding, or not using an adequate
`method of contraception were also ex-
`
`cluded. At randomization, fasting blood
`glucose had to be 6 –13 mmol/l. Patients
`were withdrawn from the trial if fasting
`plasma glucose was ⬎15 mmol/l.
`In total, 311 patients were screened
`and 193 patients were randomized to a
`treatment group (Table 1). The reasons
`for failing screening were not meeting any
`inclusion criteria or meeting any exclu-
`sion criteria (n ⫽ 86), not meeting ran-
`domization criteria (n ⫽ 7), withdrawn
`consent (n ⫽ 6), and other (n ⫽ 19). The
`majority of patients (n ⫽ 158) were being
`treated with OHA(s) and the remaining
`35 patients were being treated with diet
`only. Most patients were on monotherapy
`with either metformin (65 patients) or
`sulfonylureas (55 patients); 22 patients
`received a combination of metformin and
`sulfonylureas, 14 patients received repa-
`glinide, and 2 patients received acarbose
`treatment. In all, 190 patients were ex-
`posed to the experimental protocol; 3 pa-
`tients withdrew consent before receiving
`randomized treatment.
`Patients were recruited from the par-
`ticipating investigators’ outpatient clinics
`or by local advertisements. Each patient
`was seen on seven occasions: screening;
`baseline; after 1, 4, 8, and 12 weeks of
`treatment; and follow-up. Current treat-
`ment with oral antidiabetic medication
`was discontinued at screening. Patients
`were supplied with a blood glucose meter
`(One Touch Profile Glucometer) and in-
`structed in its use. Fasting blood glucose
`was measured every morning. HbA1c,
`fasting serum glucose, insulin, C-peptide,
`and glucagon were measured every 4
`weeks. Fasting serum glucose was also
`measured after the first week of treatment.
`Proinsulin was assessed in a subset of pa-
`tients (n ⫽ 74, equally distributed among
`the groups). Fasting samples were ob-
`tained before administration of the trial
`drug. Safety parameters (adverse events,
`hypoglycemic episodes, weight, standard
`hematology and biochemistry profile, vi-
`tal signs, and electrocardiogram) were
`assessed at each visit. Liraglutide anti-
`bodies were measured before and after
`treatment.
`The trial was double blind for the five
`dosage levels of liraglutide and placebo,
`and open label for glimepiride. The blind-
`ing was kept until database release.
`Liraglutide and placebo (Novo Nor-
`disk A/S, Bagsvaerd, Denmark) were ad-
`ministered as a once-daily injection
`(subcutaneously) in the morning before
`
`breakfast. The five dosages of liraglutide
`administered were 0.045, 0.225, 0.45,
`0.60, and 0.75 mg. These dosages were
`chosen based on observations from previ-
`ous trials (16 –18). In one of those trials, a
`single dose of 10 ␮g/kg (corresponding to
`⬃0.80 mg) showed significant effects on
`glycemia, but 2 of 11 patients reported
`nausea (18). Further, once-daily dosing
`for 7 days showed a 40% increase in Cmax
`in the steady state (17). Therefore, to
`avoid causing unacceptable side effects,
`the highest dosage included in this trial
`was 0.75 mg, corresponding to 7.5 ␮g/kg
`(equivalent to 10.5 ␮g/kg in steady state)
`for a person weighing 100 kg. The lowest
`dosage of 0.045 mg was considered too
`low to have any significant effect on gly-
`cemic control. Glimepiride (Amaryl;
`Aventis Pharma, Frankfurt, Germany)
`was supplied as 1- and 2-mg tablets for
`oral use, with the dosage adjusted accord-
`ing to glycemic control during the first 4
`weeks, with the aim to achieve a fasting
`plasma glucose level ⬍7 mmol/l. The
`mean glimepiride dosage during the trial
`was 2.7 mg. Compliance was assessed by
`drug accountability and plasma concen-
`tration measurements.
`
`Analytic methods
`HbA1c was analyzed with a Unimate
`HbA1c assay (Roche Diagnostics; normal
`range 4.5–5.7%). Liraglutide antibodies
`were determined by a radioimmunoassay
`developed by the Department of Immu-
`nochemistry at Novo Nordisk A/S. The
`serum concentrations of insulin, C-
`peptide, and proinsulin were analyzed by
`enzyme-linked immunosorbent assay
`methods. Plasma glucagon was analyzed
`by MDS Pharma Services (Wangen, Swit-
`zerland) using a radioimmunoassay
`(Linco Research, St. Charles, MO). Stan-
`dard laboratory analyses were performed
`by a central laboratory (Novo Medical
`Medi-Lab, Clinical Trials Lab, Copenha-
`gen, Denmark, if not otherwise stated).
`
`Statistical analysis
`Pretrial calculation showed that a two-
`sided test of the highest dosage versus the
`placebo group required 30 patients per
`group to detect a difference in mean
`HbA1c of at least 1% unit with 5% signif-
`icance and 95% power.
`The primary end point HbA1c, as well
`as secondary end points (fasting serum
`glucose, fasting C-peptide, glucagon, in-
`sulin, homeostasis model assessment
`
`1336
`
`DIABETES CARE, VOLUME 27, NUMBER 6, JUNE 2004
`
`Novo Nordisk Exhibit 2075
`Mylan Pharms. Inc. v. Novo Nordisk A/S
`IPR2023-00724
`Page 00002
`
`

`

`Downloaded from http://diabetesjournals.org/care/article-pdf/27/6/1335/645887/zdc00604001335.pdf by guest on 13 January 2024
`
`Table 1—Patient disposition and characteristics
`
`Placebo
`
`0.045 mg
`
`0.225 mg
`
`0.45 mg
`
`0.60 mg
`
`0.75 mg
`
`Glimepiride Total
`
`Liraglutide
`
`Madsbad and Associates
`
`—
`—
`—
`
`—
`
`—
`—
`29
`29
`5
`0
`3
`2
`24
`29
`
`—
`—
`—
`
`—
`
`—
`—
`26
`26
`3
`0
`2
`1
`23
`26
`
`—
`—
`—
`
`—
`
`—
`—
`25
`24
`3
`1
`1
`1
`21
`24
`
`—
`—
`—
`
`—
`
`—
`—
`27
`27
`7
`0
`4
`3
`20
`27
`
`—
`—
`—
`
`—
`
`—
`—
`30
`30
`2
`1
`1
`0
`28
`30
`
`—
`—
`—
`
`—
`
`—
`—
`29
`28
`2
`1
`1
`0
`26
`28
`
`—
`—
`—
`
`—
`
`—
`—
`27
`26
`0
`0
`0
`0
`26
`26
`
`311
`118
`86
`
`7
`
`6
`19
`193
`190
`22
`3
`12
`7
`168
`190
`
`Patient disposition
`Screened
`Screening failures
`Nonfulfillment of any
`inclusion/exclusion
`criteria
`Not meeting randomization
`criterion
`Withdrew consent
`Other
`Randomized
`Exposed
`Withdrawals
`Adverse events
`Ineffective therapy
`Other
`Completed
`Included in ITT population
`Baseline characteristics
`Age (years)
`Sex
`Male
`Female
`BMI (kg/m2)
`Previous diabetes treatment
`Diet
`OHA
`Duration of diabetes (years)
`HbA1c (%)
`Fructosamine (mmol/l)
`Fasting serum glucose
`(mmol/l)
`0.9 ⫾ 0.5
`1.0 ⫾ 0.4
`1.1 ⫾ 0.6
`1.1 ⫾ 0.5
`1.0 ⫾ 0.5
`1.0 ⫾ 0.5
`1.1 ⫾ 0.4
`C-peptide (mmol/l)
`72.5 ⫾ 55.3 71.5 ⫾ 42.0 64.1 ⫾ 40.4
`73.8 ⫾ 51.0
`84.7 ⫾ 43.6 74.1 ⫾ 40.1 81.2 ⫾ 73.0
`Insulin (mmol/l)
`99 ⫾ 31.1
`91 ⫾ 23.4
`101 ⫾ 23.6
`110 ⫾ 33.0
`110 ⫾ 37.7
`111 ⫾ 47.6
`131 ⫾ 140.0
`Glucagon (mmol/l)
`0.21 ⫾ 0.11 0.25 ⫾ 0.15 0.28 ⫾ 0.19
`0.30 ⫾ 0.20
`0.23 ⫾ 0.15 0.23 ⫾ 0.16 0.26 ⫾ 0.17
`Proinsulin-to-insulin ratio
`44.06 ⫾ 26.3 34.65 ⫾ 23.1 31.83 ⫾ 22.6 31.95 ⫾ 31.7 34.40 ⫾ 37.3 33.75 ⫾ 21.7 26.78 ⫾ 18.2
`␤-Cell function (%)
`Data are means ⫾ SD. Demographics were obtained at visit 1 (screening) and diabetes characteristics at visit 2 (baseline). Of the 193 patients randomized, 3 were
`never exposed. Preferred terms for adverse event withdrawals include sinusitis and fever (0.225 mg), aggressive reaction (0.60 mg), and abnormal hepatic function
`(0.75 mg). Pro-insulin was measured only in a subset of patients (n ⫽ 74).
`
`57 ⫾ 9.4
`
`53 ⫾ 9.0
`
`58 ⫾ 7.5
`
`57 ⫾ 11.3
`
`57 ⫾ 7.7
`
`58 ⫾ 9.7
`
`57 ⫾ 9.2
`
`20
`9
`30.3 ⫾ 4.2
`
`22
`4
`30.2 ⫾ 5.4
`
`15
`9
`32.0 ⫾ 5.3
`
`18
`9
`30.1 ⫾ 5.0
`
`20
`10
`30.4 ⫾ 4.8
`
`16
`12
`31.9 ⫾ 4.3
`
`16
`10
`30.2 ⫾ 4.6
`
`4
`25
`3.4 ⫾ 2.9
`7.4 ⫾ 1.2
`335 ⫾ 72.5
`9.7 ⫾ 2.9
`
`2
`24
`4.1 ⫾ 3.7
`7.4 ⫾ 0.8
`344 ⫾ 77.9
`10.2 ⫾ 2.2
`
`11
`13
`4.4 ⫾ 4.0
`7.9 ⫾ 0.8
`358 ⫾ 57.7
`10.9 ⫾ 3.6
`
`2
`25
`4.5 ⫾ 4.6
`7.7 ⫾ 1.0
`374 ⫾ 104.9
`11.2 ⫾ 2.8
`
`4
`26
`4.6 ⫾ 4.6
`7.4 ⫾ 1.2
`352 ⫾ 74.2
`10.8 ⫾ 2.8
`
`4
`24
`6.1 ⫾ 7.9
`7.4 ⫾ 0.9
`344 ⫾ 73.7
`9.9 ⫾ 2.3
`
`7
`19
`3.8 ⫾ 3.4
`7.8 ⫾ 0.9
`364 ⫾ 69.5
`10.6 ⫾ 2.4
`
`[HOMA], proinsulin-to-insulin ratio, and
`weight) were analyzed in a mixed-effects
`model with treatment, visit, and center as
`fixed effects and patient as the random
`effect. The interaction term, baseline
`HbA1c by visit, was included in the model
`as a covariate. Adjusted end point levels at
`12-week follow-up were calculated for
`each treatment group by means of this
`model.
`The HOMA(S) (␤-cell function) was
`determined as ␤-cell function (%) ⫽ 20 ⫻
`insulin/(glucose ⫺ 3.5). HOMA(R) (insu-
`lin resistance) was determined as resis-
`tance ⫽ insulin/(22.5e⫺ln[glucose]) (22).
`All analyses were performed for the
`
`intention-to-treat (ITT) population (i.e.,
`all patients who received at least one dose
`of a trial drug). Statistical analyses were
`performed using SAS software (version 8;
`SAS Institute, Cary, NC).
`
`RESULTS
`
`Enrollment
`The 193 patients randomized in this trial
`were evenly distributed across treatment
`groups (Table 1). Baseline clinical charac-
`teristics of the 190 patients exposed to the
`experimental protocol are given in Table
`1. No apparent differences were seen
`
`among the treatment groups with regard
`to baseline characteristics.
`
`Effect on glycemic control
`After 12 weeks of treatment, HbA1c was
`decreased in all groups except the lowest
`liraglutide dosage group (Fig. 1A). Treat-
`ment with the two highest dosages re-
`duced HbA1c significantly more than
`placebo (Table 2). At these dosage levels,
`the effect of liraglutide was comparable
`with that of glimepiride with respect to
`effect on HbA1c. The effect of liraglutide
`increased with duration of treatment. The
`largest decreases in HbA1c levels were ob-
`served at the end of the 12-week treat-
`
`DIABETES CARE, VOLUME 27, NUMBER 6, JUNE 2004
`
`1337
`
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`IPR2023-00724
`Page 00003
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`

`Improved glycemic control with liraglutide (NN2211)
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`Downloaded from http://diabetesjournals.org/care/article-pdf/27/6/1335/645887/zdc00604001335.pdf by guest on 13 January 2024
`
`Figure 1—Glycemic control. Mean difference in HbA1c compared with placebo (95% CI) after 12
`weeks of treatment (A), mean HbA1c (%) during the 12-week treatment period (B), mean differ-
`ence in fasting serum glucose compared with placebo (95% CI) after 12 weeks of treatment (C),
`and mean fasting serum glucose (mmol/l) during the 12-week treatment period in the ITT popu-
`lation (D). A and C: *P ⬍ 0.05, **P ⬍ 0.01, ***P ⬍ 0.001 vs. placebo. Glim, glimepiride. B and
`D: F, 0.045 mg liraglutide; f, 0.225 mg liraglutide; Œ, 0.45 mg liraglutide; , 0.60 mg lira-
`glutide; }, 0.75 mg liraglutide; [gray circles], glimepiride; 䡺, placebo.
`
`protocol as blood glucose ⬍2.8 mmol/l)
`and 7 reported symptoms of hypoglyce-
`mia only. These incidences seemed lower
`than in the glimepiride group (n ⫽ 26),
`where four patients experienced minor
`hypoglycemia and five patients reported
`symptoms of hypoglycemia.
`The number of patients with adverse
`events (based on spontaneous adverse
`event reporting) was comparable across
`the liraglutide groups and the placebo
`
`group (60% [81 of 135]) vs. 55% [16 of
`29] of patients, respectively) and was
`lower in the open-label reference group
`(35% [9 of 26] of patients). For gastroin-
`testinal events, the incidence seemed to
`increase with increasing doses of lira-
`glutide; nausea was reported by 1–2 pa-
`tients in each of the lowest dose groups
`and by 5 of 28 patients in the highest dose
`group (in total, reported by 10 of 135 pa-
`tients exposed to liraglutide) compared
`
`ment period for the highest dosages (Fig.
`1B), and it seemed that HbA1c levels were
`still decreasing at the end of the treatment
`period at these dosages. We observed that
`59% of patients completing the trial in the
`two highest liraglutide dosage groups
`ⱕ7% after 12 weeks.
`achieved HbA1c
`As was seen with HbA1c, fasting se-
`rum glucose levels decreased in most
`treatment groups during the trial, with
`the decreases being statistically significant
`for the 0.225-, 0.60-, and 0.75-mg lira-
`glutide dosage groups compared with
`placebo (Table 2). In accordance with the
`observations for HbA1c, the effect of the
`highest dosages of liraglutide was compa-
`rable with that of glimepiride with respect
`to fasting serum glucose (Fig. 1C). In the
`liraglutide groups, maximal effect on fast-
`ing serum glucose was evident after the
`first week of treatment (Fig. 1D).
`
`Effect on body weight
`Treatment with liraglutide did not in-
`crease body weight (Fig. 2). Furthermore,
`for the 0.45-mg liraglutide dosage group
`(Table 2), a statistically significant de-
`crease, compared with placebo, was ob-
`served.
`
`Effect on islet cell function
`HOMA (22) was used to assess ␤-cell
`function and insulin resistance after 12
`weeks of liraglutide treatment. Mean
`␤-cell function (derived from fasting in-
`sulin and glucose) was significantly
`higher in the 0.75-mg liraglutide group
`after 12 weeks than in the placebo group
`(Table 2). The effect obtained with the
`highest liraglutide dosage was similar to
`that observed with glimepiride. For insu-
`lin resistance, no differences were seen
`among the three treatments (liraglutide,
`glimepiride, and placebo; data not
`shown). Further, the proinsulin-to-
`insulin ratio decrease was statistically sig-
`nificantly after treatment with 0.75 mg
`liraglutide compared with placebo (Table
`2). No change in the proinsulin-to-insulin
`ratio was demonstrated after treatment
`with glimepiride. There were no statisti-
`cally significant differences between the
`liraglutide groups and the placebo group
`for fasting insulin, C-peptide, and gluca-
`gon (Table 2).
`
`Safety evaluation
`Of the 135 patients exposed to liraglutide,
`1 (in the 0.60-mg group) experienced mi-
`nor hypoglycemia (defined in the study
`
`1338
`
`DIABETES CARE, VOLUME 27, NUMBER 6, JUNE 2004
`
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`IPR2023-00724
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`
`Madsbad and Associates
`
`with 1 of 29 patients in the placebo group.
`Other gastrointestinal events included di-
`arrhea (5 of 135 liraglutide-treated pa-
`tients), vomiting (3 of 135 patients), and
`constipation (3 of 135 patients). None of
`these events were reported in the placebo
`or glimepiride groups, with the exception
`of one patient experiencing vomiting in
`the glimepiride group. Approximately
`two-thirds of the gastrointestinal events
`reported during treatment with lira-
`glutide were resolved within 1–3 days.
`None of the patients withdrew due to gas-
`trointestinal events (Table 1). Overall, the
`most frequent adverse events were head-
`ache and nausea, which were also the
`events most often assessed as related to
`the trial product. Generally, adverse
`events were mild or moderate and com-
`pletely resolved. Only 1 of the 135 pa-
`tients exposed to liraglutide experienced
`two mild injection site reactions, de-
`scribed as an “urticarial reaction at injec-
`tion place.” No antibody formation
`against liraglutide could be detected in
`this trial.
`No safety issues with regard to vital
`signs, electrocardiogram, or laboratory
`analysis were raised by this trial.
`
`CONCLUSIONS — This trial was the
`first to demonstrate a sustained improve-
`ment in glycemic control after long-term
`treatment with the GLP-1 analog lira-
`glutide, administered once daily. After 12
`weeks of treatment with 0.60 or 0.75 mg
`liraglutide, both HbA1c and fasting serum
`glucose levels were significantly lower
`than with placebo. Glycemic control was
`maintained throughout the treatment pe-
`riod and was as effective at the highest
`liraglutide dosages as that provided by the
`open-label reference therapy glimepiride
`
`Figure 1—Continued
`
`Table 2—Repeated measures analysis after 12 weeks, comparison with placebo in ITT population
`
`Liraglutide (mg)
`
`0.045
`
`0.225
`
`0.45
`
`0.60
`
`0.75
`
`Glimepiride
`
`26
`28
`30
`27
`24
`26
`0.25 (0.1905) ⫺0.34 (0.0877) ⫺0.30 (0.1131) ⫺0.70 (0.0002) ⫺0.75 (⬍0.0001) ⫺0.74 (0.0001)
`0.74 (0.1499) ⫺1.37 (0.0090) ⫺0.35 (0.4976) ⫺2.14 (⬍0.0001) ⫺1.82 (0.0003)
`⫺2.60 (⬍0.0001)
`⫺0.03 (0.9602) ⫺0.74 (0.1544) ⫺1.20 (0.0184)
`0.27 (0.5838) ⫺0.39 (0.4391)
`0.94 (0.0622)
`⫺0.04 (0.6468) ⫺0.12 (0.1314) ⫺0.04 (0.6502) ⫺0.12 (0.1775) ⫺0.18 (0.0244) ⫺0.003 (0.9635)
`⫺6.07 (0.3546)
`7.55 (0.2514)
`8.55 (0.1978)
`7.33 (0.2409)
`23.56 (0.0002)
`24.57 (0.0002)
`⫺2.44 (0.8425) ⫺4.15 (0.7381) ⫺0.50 (0.9679) ⫺7.47 (0.5269)
`8.57 (0.4708)
`16.79 (0.1597)
`⫺0.09 (0.4033) ⫺0.01 (0.9449)
`0.05 (0.6374) ⫺0.08 (0.4308)
`0.11 (0.2921)
`0.08 (0.4636)
`12.74 (0.3619) ⫺15.84 (0.2710) ⫺6.77 (0.6277) ⫺7.68 (0.5625) ⫺9.33 (0.4980)
`⫺7.21 (0.5970)
`
`n
`HbAlc (%)
`Fasting serum glucose (mmol/l)
`Weight (kg)
`Proinsulin-to-insulin ratio
`␤-Cell function (%)
`Fasting insulin (mmol/l)
`Fasting C-peptide (mmol/l)
`Fasting glucagon (mmol/l)
`P value given in parentheses.
`
`DIABETES CARE, VOLUME 27, NUMBER 6, JUNE 2004
`
`1339
`
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`IPR2023-00724
`Page 00005
`
`

`

`Improved glycemic control with liraglutide (NN2211)
`
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`
`tected after 12 weeks of treatment. The
`sensitivity of the assay has been found to
`be 30 ng/ml (data on file) using a mono-
`clonal antibody. The liraglutide antibody
`assay is essentially similar to a previously
`published assay used to assess insulin and
`insulin aspart antibodies with a similar
`and acceptable sensitivity (33).
`Another approach for investigating
`the potential of GLP-1 includes adminis-
`tering native GLP-1 by means of continu-
`ous infusion (11,14). Continuous
`intravenous infusion of GLP-1 for 1 week
`resulted in an instant decrease in blood
`glucose levels in patients in whom sulfo-
`nylurea therapy failed to be effective (14).
`A 6-week trial with GLP-1 administered
`as continuous subcutaneous infusion via
`an insulin pump demonstrated significant
`effects on fasting and 8-h plasma glucose
`and HbA1c, as well as an inhibition of gas-
`tric emptying and body weight loss (11).
`The latter trial was conducted in patients
`with poorly controlled HbA1c levels, indi-
`cating that the GLP-1 treatment concept
`may be applied to a wide range of glyce-
`mia in type 2 diabetes. Also, the peptide
`exenatide (synthetic exendin-4) has dem-
`onstrated effects similar to those of GLP-1
`(34). In this 28-day study, adding ex-
`enatide to the existing treatment two or
`three times daily showed a significant ef-
`fect in controlling patients’ HbA1c levels,
`which were previously not satisfactorily
`controlled with diet and/or OHAs.
`In conclusion, the long-acting GLP-1
`analog liraglutide shows considerable
`promise as a once-daily therapy in type 2
`diabetes for lowering blood glucose with-
`out weight gain or substantial risk of
`hypoglycemia.
`
`Acknowledgments — This trial was spon-
`sored by Novo Nordisk A/S, which established
`the protocol in conjunction with an academic
`committee that was responsible for data re-
`view and this article. The decision to submit
`the manuscript was that of the named authors.
`We are indebted to the following principal
`investigators and their staff, who participated
`in the NN2211-1310 international study
`group. Norway: H. Ulrichsen, Harstad; A.
`Tandberg, Bekkestua; M. Wangestad, Oslo; S.
`Madsbu, Elverum. Sweden: B. Eliasson, Gote-
`borg; A. Gamstedt, Orebro; E. Bjork, Uppsala;
`B. Polhem, Goteborg. Denmark: S. Madsbad,
`Hvidovre; O. Schmitz, Aarhus; K. Koelendorf,
`Koege; H. Perrild, Copenhagen; A. Prange,
`Kolding; S. Nistrup Holmegaard, Thisted.
`
`Figure 2—Mean weight change from baseline (kg), ITT population. F, 0.045 mg liraglutide; f,
`0.225 mg liraglutide; Œ, 0.45 mg liraglutide; , 0.60 mg liraglutide; }, 0.75 mg liraglutide; [gray
`circles], glimepiride; 䡺, placebo.
`
`at an average dosage of 2.7 mg (a near-
`maximal dosage) (23), with trends toward
`less hypoglycemia and weight loss. At the
`highest dosage levels, there was a trend
`toward improved HbA1c with duration of
`treatment. This suggests that further im-
`provements in HbA1c levels may be seen
`with extended (⬎12 weeks) treatment.
`More than half the patients in the two
`highest dosage liraglutide groups had
`ⱕ 7% after 12 weeks of treatment.
`HbA1c
`Furthermore, treatment with liraglutide
`caused no weight increase, and, at the
`highest dosage, the difference with
`glimepiride was statistically significant.
`This observation was in accordance with
`findings that GLP-1 suppresses energy in-
`take in humans (8).
`The full dosage-response curve for li-
`raglutide remains to be elucidated, and
`future trials will investigate higher dos-
`ages of drug, but the current trial con-
`firmed the proposed once-daily dosing
`regimen for liraglutide.
`The proinsulin-to-insulin ratio de-
`creased during 12 weeks’ treatment with
`liraglutide. This ratio is known to be ele-
`vated in type 2 diabetes and is considered
`an index of insulin secretory dysfunction
`(24,25). It is believed that the hyperpro-
`insulinemia is caused by an increased
`demand placed on the ␤-cell by hypergly-
`cemia and insulin resistance, which in
`turn leads to secretion of an increased
`amount of incompletely processed gran-
`ules containing proinsulin. This is also
`
`known as the “overworked ␤-cell” hy-
`pothesis (26). An alternative hypothesis is
`that the increased proinsulin concentra-
`tion is related to an intrinsic ␤-cell defect
`in type 2 diabetes (24). In both cases, im-
`provement of the proinsulin-to-insulin
`ratio, and thus of ␤-cell function, would
`be a desirable effect of a potential new
`treatment for type 2 diabetes. ␤-Cell func-
`tion, as determined by HOMA analysis,
`was significantly improved at the highest
`liraglutide dosage compared with pla-
`cebo. There was no effect on the ␣-cells in
`the fasting state, in so far as the glucagon
`concentrations were unaltered.
`The safety profile of liraglutide was
`favorable, with the expected very low risk
`of hypoglycemic episodes as anticipated
`with the glucose-dependent mode of ac-
`tion of native GLP-1 (27). Adverse events
`were isolated, with the most frequent be-
`ing headache and nausea. Usually the
`events were transient and of mild or mod-
`erate severity and resolved without inter-
`vention. Although both nausea and
`vomiting were reported, neither of these
`was of a severity that caused the patient to
`withdraw. Most gastrointestinal events
`were transient (i.e., they resolved within
`1–3 days). Side effects within the gastro-
`intestinal system are known to occur after
`GLP-1 treatment and seem to be dosage
`related (14,28,29). These effects probably
`result from the inhibition of gastric emp-
`tying by GLP-1 (9,30 –32). No formation
`of antibodies toward liraglutide was de-
`
`1340
`
`DIABETES CARE, VOLUME 27, NUMBER 6, JUNE 2004
`
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`Mylan Pharms. Inc. v. Novo Nordisk A/S
`IPR2023-00724
`Page 00006
`
`

`

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`
`Madsbad and Associates
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`U.K.: R. Blandford, Worksop; C. Fox, North-
`hampton; B. Frier, Edinburgh; R. Gregory,
`Leicester; S. Heller, Sheffield; D. Hepburn,
`Hull; A. Jaap, Kilmarnock; P. Mansell, Not-
`tingham; D. Matthews, Oxford; B. Millward,
`Plymouth; P. O’Hare, Rugby; M. Page, Llant-
`risant; J. Vora, Liverpool; A. Ogilvie, Enfield.
`The study results were presented in an oral
`presentation at the American Diabetes Associ-
`ation 62nd Annu