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Journal of Clinical Endocrinology & Metabolism , doi:10.1210/jc.2006-1740
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The Journal of Clinical Endocrinology & Metabolism Vol. 92, No. 2 527-531
Copyright © 2007 by The Endocrine Society

Lack of an Effect of a Novel ß3-Adrenoceptor Agonist, TAK-677, on Energy Metabolism in Obese Individuals: A Double-Blind, Placebo-Controlled Randomized Study

Leanne M. Redman, Lilian de Jonge, Xiaobing Fang, Betsy Gamlin, David Recker, Frank L. Greenway, Steven R. Smith and Eric Ravussin

Pennington Biomedical Research Center (L.M.R., L.d.J., X.F., F.L.G., S.R.S., E.R.), Baton Rouge, Louisiana 70808; and Takeda Pharmaceuticals North America, Inc. (B.G., D.R.), Lincolnshire, Illinois 60059

Address all correspondence and requests for reprints to: Eric Ravussin, 6400 Perkins Road, Baton Rouge, Louisiana 70808. E-mail: ravusse{at}pbrc.edu.


   Abstract
Top
 Abstract
Introduction
 Subjects and Methods
 Results
 Discussion
 References
 
Objective: Our objective was to test the safety and metabolic effects of a novel ß3-adrenoreceptor agonist (TAK-677) in humans.

Design, Setting, and Participants: Sixty-five obese (body mass index = 33.9 ± 2.1 kg/m2, mean ± SE) men and women (31.4 ± 0.9 yr) participated in a double-blind placebo-controlled study at an institutional research center.

Intervention: Participants were randomized to 0.1 mg TAK-677 twice daily (BID) (n = 21), 0.5 mg TAK-677 BID (n = 22), or placebo BID (n = 22) for 29 d.

Outcomes: Drug safety, 24-h respiratory quotient (RQ), 24-h energy expenditure (EE), body composition, fat distribution, and fasting plasma concentration of substrates and hormones were assessed. An acute-response study was also conducted.

Results: The drug was well tolerated by all participants; however, heart rate was elevated (9 ± 2 beats per minute) with the 0.5-mg BID dose. After 28 d of treatment and when compared with placebo, there was no change in 24-h RQ with either 0.1-mg BID (P = 0.1) or 0.5-mg BID (P = 1.0) doses of TAK-677. However, TAK, 0.5 mg BID, resulted in a small increase in 24-h EE that was significantly different from placebo [change from baseline, 13 ± 17 (0.5 mg BID) vs.–39 ± 18 (placebo) kcal/d, P < 0.05]. Changes in weight, fat-free mass, and abdominal fat depots (visceral or sc) were not different between the three groups, nor were changes in fasting insulin, free fatty acid, or glucose concentrations.

Conclusion: TAK-677 has no effect on 24-h RQ or fat oxidation but does slightly increase 24-h EE at the highest dose (0.5 mg BID). The acute studies showed large interindividual variability in plasma concentrations of TAK-677 indicating some possible problems with bioavailability and therefore efficacy.


   Introduction
Top
 Abstract
 Introduction
Subjects and Methods
 Results
 Discussion
 References
 
FOR THE PAST 20 yr, the adrenoceptor (AR) has been the focus of extensive research as a potential drug target for the treatment of obesity (1). Specifically, the ß3-AR subtype identified in both white (2) and brown (3, 4) adipose tissue and potentially in skeletal muscle (5) has been demonstrated to significantly decrease obesity and improve insulin-sensitizing actions in rodents (6, 7). TAK-677, a hydroxyethyl aminopropyl derivative ([3-[(2R)-[[(2R)-(3-chlorophenyl)-2-hydroxyethyl]amino]propyl]-1H-indol-7-yloxy]-acetic acid), is a novel ß3-AR agonist (8).

The first studies of TAK-677 (also reported as AJ-9677) in rodents have demonstrated potential antiobesity and antidiabetic outcomes (9, 10, 11). Administration of AJ-9677 (0.1 mg/kg·d) to genetically obese and diabetic mice (KK-Ay/Ta) inhibited weight gain (9) and decreased white adipose tissue mass independent of food intake, suggesting increased energy expenditure (EE). Interestingly, the fat loss was associated with the conversion of large adipocytes into smaller ones and a 20- to 80-fold increase in UCP-1, both suggestive of increased brown adipose tissue and increased fat oxidation.

This study was undertaken to assess the safety of TAK-677 and its effect on 24-h EE, substrate oxidation, body composition, and fat distribution in obese individuals before and after 4 wk (29 d) administration. The acute effects of TAK-677 on EE and fasting biochemical markers of metabolism were also assessed at the first exposure and after 4 wk of treatment.


   Subjects and Methods
Top
 Abstract
 Introduction
 Subjects and Methods
Results
 Discussion
 References
 
Subjects

Sixty-five obese (body mass index, 33.9 ± 2.1 kg/m2) but otherwise healthy men and women (31.4 ± 0.9 yr) enrolled in a placebo-controlled double-blind study. This study was approved by the Pennington Biomedical Research Center Institutional Review Board, and participants provided their written informed consent. Sample size calculations indicated that 20 subjects per treatment group were needed to detect a change in 24-h respiratory quotient (RQ) of 0.03 between the 0.5-mg TAK-677 group and placebo ({alpha} < 0.05 and 80% power). Such change in RQ would result in an increase of 15 g fat oxidation per day, a change similar to what was previously observed with CL316,243, a highly selective ß3-AR agonist (12).

Design

Participants were assigned to one of three treatment groups: 0.1 mg TAK-677, 0.5 mg TAK-677, or placebo in random order. For 29 d, participants were required to ingest TAK-677 or placebo, orally twice a day (BID), once in the morning (at least 1 h before breakfast) and once in the evening (at least 5 h after lunch and 1 h before dinner). At baseline (before treatment) and after 28 d of treatment, participants were admitted to the inpatient unit where assessments for body weight and composition, 24-h energy metabolism, and fasting plasma concentration of substrates and hormones were determined. The acute (2-h) effects of TAK-677 administration were also performed after exiting the chamber (d 1 and 29) during these visits. Drug safety (vital signs) and compliance were monitored throughout the study. Females commenced treatment between d 3–14 of their menstrual cycle. An Investigational New Drug approval was obtained for TAK-677.

Diet

Throughout the study, individuals followed a preplanned isocaloric diet. All meals prepared by the metabolic kitchen were provided for 72 h before the inpatient visits at baseline and d 28.

Body weight and body composition

At baseline and on d 28, the following measurements were performed: body mass, percent fat (dual-energy x-ray absorptiometry; Hologics QDR 4500A, Bedford, MA) and abdominal fat mass (multislice computed tomography; GE Medical Systems, Fairfield, CT) as previously described (13).

Energy metabolism

The 24-h RQ, 24-h EE, and sleeping metabolic rate were measured in a metabolic chamber (14). Subjects entered the chamber at 0745 h after an overnight fast and remained therein until 0700 h the following morning. During the chamber stay, subjects refrained from exercise and received an isocaloric diet provided in three meals. Energy intake was adjusted to match EE to maintain energy balance (15), and the same energy intake was provided during both chamber visits. Twenty-four-hour urine was collected for quantification of nitrogen, epinephrine, and norepinephrine excretion.

Acute assessment of TAK-677

Immediately after exiting the chamber at baseline (d 1, data not shown) and d 29, an iv catheter was inserted into a forearm vein and a fasting blood sample collected for glucose, free fatty acids (FFA), insulin, and TAK-677. Resting metabolic rate (RMR) was then measured for 30 min (Deltatrac II metabolic cart; Datex-Ohmeda, Helsinki, Finland). After the RMR, participants consumed the morning dose of their study medication and RMR measurements continued for another 2 h. Blood samples were collected at 1 and 2 h, and heart rate was monitored continually by a three-channel Holter monitor (Mortara Instrument Inc., Milwaukee, WI).

Biochemical analysis

Serum insulin was measured using an immunoassay (DPC 2000; Diagnostic Products Corp., Los Angeles, CA), glucose using a glucose oxidase electrode (Syncron CX7; Beckman, Brea, CA), and 24-h urine epinephrine and norepinephrine by HPLC (Bio-Rad, Hercules, CA). TAK-677 concentrations were measured in heparinized plasma samples by sensitive and specific liquid chromatography-mass spectrometry at MDS Pharma Services (Lincoln, NE). The minimal detectible concentration was 10.00 pg/ml, and the mean coefficient of variation across 15 assay runs was less than 9.65%.

Statistics

Statistical analyses were performed with the SAS procedures version 9.1.3 (SAS Institute, Cary, NC), and the level of significance for all statistical tests was set at P < 0.05. Data are expressed as means ± SEM. To test for treatment effects (placebo, 0.1 mg TAK-677, and 0.5 mg TAK-677), the change from baseline to d 28 or d 29 was computed and an ANOVA was performed with the corresponding baseline value included in the model as a covariate. Longitudinal effects of the study medication in the pharmacokinetic studies were tested by ANOVA incorporating repeated measures with treatment, time, and the interaction term included in the model. The change in TAK-677 concentration and the change in RMR, RQ, FFA, and heart rate were tested using Pearson correlation coefficient.


   Results
Top
 Abstract
 Introduction
 Subjects and Methods
 Results
Discussion
 References
 
Subjects

Overall, TAK-677 was well tolerated by all participants. A summary of the adverse events is reported in Table 1Go. Nine participants withdrew from the study: four voluntarily, two after study protocol violation, and three because of adverse events, including palpitations, insomnia (placebo), nausea/vomiting (0.1 mg TAK-677), and alcoholism (0.5 mg TAK-677).


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  		TABLE 1. Effect of TAK-677 administration on 24-h EE and substrate oxidation measured in a metabolic chamber and drug safety

 
Longitudinal effects of TAK-677

There was no change in body weight, fat-free mass, abdominal fat, or fasting plasma concentrations of glucose, insulin, or FFA, and homeostasis model assessment for insulin resistance. Contrary to expectation, 0.5 mg TAK-677 caused a small increase in fat mass (P = 0.03), which was different from placebo (P = 0.01) and 0.1 mg TAK-677 (P < 0.04).

The 24-h energy metabolism. At baseline, 24-h RQ, 24-h EE, and sleep metabolic rate were similar across all three groups (Table 1Go). TAK-677 0.5 mg resulted in a small increase in 24-h EE that was significantly different from the placebo-treated group (change from baseline, 0.5 mg BID 13 ± 17 vs. placebo –39 ± 18 kcal/d, P < 0.05). There was no treatment effect on 24-h RQ, sleeping metabolic rate, or urinary epinephrine and norepinephrine excretion.

Vital signs. TAK-677 treatment did not affect oral temperature (P = 0.69), resting blood pressures (P > 0.50), or electrocardiogram (PR, QRS, or QT). Resting heart rate, however, was significantly increased from baseline in all groups (placebo 4.9 ± 2.7, 0.1 mg TAK 2.6 ± 1.4, and 0.5 mg TAK 8.7 ± 2.1 beats/min, P < 0.05 for all), but no treatment effect (P = 0.16) was observed. No participant reported hand or other body tremors.

Acute effects of TAK-677 on d 29

There was no difference in the acute response of TAK-677 on any parameter between d 1 and 29. After an acute dose, plasma TAK-677 concentration increased several-fold above baseline (mean 2-h change, 0.1 mg BID 2110 ± 1821 vs. 0.50 mg BID 9608 ± 5332 pg/ml) in both of the TAK-677-treated groups (Fig. 1AGo).


Figure 1
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  		FIG. 1. A–C, Acute effects of a single oral dose of TAK-677 on TAK-677 concentrations (A), FFA concentrations (B), and RMR (C). The 2-h mean changes are also shown (inset). D, The relationship between TAK-677 concentrations and RMR to an acute dose of TAK-677. *, Significant change from placebo group; {dagger}, significant change from 0.1 mg TAK-677 group. Black, Placebo; light gray, 0.1 mg TAK-677; dark gray, 0.5 mg TAK-677.

 
Energy metabolism. RMR increased progressively with TAK-677, reaching approximately 7% greater above baseline with 0.5 mg BID and approximately 3% with 0.1 mg BID (Fig. 1CGo). The mean change in 2-h RMR (Fig. 1CGo, inset) was positively associated with the mean change in TAK-677 (Fig. 1DGo). There was no difference reported between groups for RQ.

Fasting concentrations of glucose, insulin, and FFA. Fasting FFA concentrations increased significantly in all three groups (Fig. 1BGo) with the mean change greatest with 0.5 mg TAK-677 and significantly different from placebo (P < 0.01). The mean change in FFA was positively correlated with the mean change in TAK-677 (r = 0.34; P < 0.01). Fasting insulin concentrations decreased in all three groups (data not shown), and the reduction was different between 0.5 mg BID and placebo (P = 0.01). Fasting glucose concentrations were not altered by the study medication in any group.

Heart rate. Heart rate did not change in the placebo or 0.1-mg TAK-677-treated groups but increased by 8 beats/min (~11%) in the 0.5-mg TAK-677-treated group 2 h after dose. The 2-h mean heart rate response was positively associated with the mean change in TAK-677 (r = 0.57; P < 0.001).


   Discussion
Top
 Abstract
 Introduction
 Subjects and Methods
 Results
 Discussion
References
 
The purpose of the present study was to test for the first time in humans the acute and longer-term effects of a novel ß3-AR agonist TAK-677 on energy metabolism. When compared with placebo, the highest dose of TAK-677 (0.5 mg BID) resulted in a very small but statistically significant increase in 24-h EE after 28 d of treatment. There was little or no change in 24-h RQ and no change in fasting concentrations of glucose, FFA, and insulin or 24-h catecholamine excretion. The acute study indicated that plasma concentrations of TAK-677 vary considerably between individuals, indicating some issues with bioavailability. Acutely, the changes in RMR, RQ, FFA, and heart rate were related in a plasma concentration-dependent manner to the exposure of the compound (changes in TAK-677 concentration).

In rodents, TAK-677 proved very potent, increasing the expression of UCP-1 in white and brown adipose tissue, causing conversion of large to smaller adipocytes (9). However, our study in humans indicates that TAK-677 was unable to sufficiently stimulate EE and fat oxidation. The lack of thermogenic efficacy of TAK-677 in humans may be explained by species variation of the ß3-AR or by the low numbers of biologically active ß3 receptors in tissues that can mediate a significant thermogenic response. Another explanation for the lack of an effect on 24-h EE and substrate oxidation could be that the plasma concentrations of TAK-677 were too low or too highly variable. Indeed, at the high dose, fasting concentrations were 835 ± 346 pg/ml but ranged from 0–5711 pg/ml, and as expected, concentrations were lower in the 0.1-mg-treated group but again very variable (0–4545 pg/ml). Although it is tempting to speculate that the lack of a chronic effect is because of a down-regulation of the ß3-AR, studies indicate that the ß3-AR, unlike its ß1 and ß2 counterparts, cannot undergo desensitization because it holds no sites for phosphorylation by protein kinase A or ß-AR kinase (16), and moreover, studies of TAK-677 in rodent suggest that the ß3-AR may even be up-regulated during long-term exposure (11).

Because many of the previously tested compounds have shared selectivity with the ß1- and ß2-ARs, it is important to ascertain whether the small change in energy metabolism is mediated by ß3 activation alone or via synergistic effects on ß1- and ß2-AR. In vitro, TAK-677 has an affinity for the ß3-AR that is 100-fold greater than the ß1 and 200-fold greater than ß2 (9). No participant reported muscular tremors, suggesting a clear separation with the ß2 receptor; however, with regard to ß1, the pharmacokinetic study showed that heart rate was significantly elevated by treatment with the 0.5-mg dose. Furthermore, plasma TAK-677 concentrations were positively correlated to heart rate responses, indicating some possible affinity with the ß1-AR.

The second generation ß3-AR agonist, TAK-677, a potent and highly selective agonist of the human ß3-AR, failed to induce significant metabolic effects in humans. At the highest tested dose, TAK-677 induced a slight increase in thermogenesis coupled with an increase in heart rate and no evidence of tremor. Plasma concentrations of TAK-677 were highly variable. The results indicated that the compound probably failed to stimulate the low number of ß3-AR in humans. Unfortunately TAK-677 is added to the growing list of ß3 compounds tested that have failed to induce any significant metabolic effects in humans, raising questions about the future of ß3-AR agonists in the treatment of obesity and type 2 diabetes.


   Acknowledgments
 
We thank the clinic staff and the subjects for their valued participation in this trial.


   Footnotes
 
This work was funded by Takeda Pharmaceuticals of North America. L.M.R. is supported by a Neil Hamilton-Fairley Training Fellowship awarded by the National Health and Medical Research Council of Australia (ID 349553).

Disclosure Summary: L.M.R., L.d.J., X.F., F.L.G., and S.R.S. have nothing to declare. B.G. and D.R. are employed by and have equity interests in Takeda Pharmaceuticals. E.R. received funds to conduct this study from Takeda Pharmaceuticals.

First Published Online November 21, 2006

Abbreviations: AR, Adrenoceptor; BID, twice a day; EE, energy expenditure; FFA, free fatty acids; RMR, resting metabolic rate; RQ, respiratory quotient.

Received August 11, 2006.

Accepted November 15, 2006.


   References
Top
 Abstract
 Introduction
 Subjects and Methods
 Results
 Discussion
 References
 

  1. Arch JR 2002 ß3-Adrenoceptor agonists: potential, pitfalls and progress. Eur J Pharmacol 440:99–107[CrossRef][Medline]
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  5. Astrup A, Bulow J, Madsen J, Christensen NJ 1985 Contribution of BAT and skeletal muscle to thermogenesis induced by ephedrine in man. Am J Physiol 248:E507–E515
  6. Arch JR, Ainsworth AT, Ellis RD, Piercy V, Thody VE, Thurlby PL, Wilson C, Wilson S, Young P 1984 Treatment of obesity with thermogenic ß-adrenoceptor agonists: studies on BRL 26830A in rodents. Int J Obes 8(Suppl 1):1–11
  7. Cawthorne MA, Carroll MJ, Levy AL, Lister CA, Sennitt MV, Smith SA, Young P 1984 Effects of novel ß-adrenoceptor agonists on carbohydrate metabolism: relevance for the treatment of non-insulin-dependent diabetes. Int J Obes 8(Suppl 1):93–102
  8. Francke S 2002 TAK-677 (Dainippon/Takeda). Curr Opin Invest Drugs 3:1624–1628[Medline]
  9. Harada H, Hirokawa Y, Suzuki K, Hiyama Y, Oue M, Kawashima H, Kato H, Yoshida N, Furutani Y, Kato S 2005 Discovery of a novel and potent human and rat ß3-adrenergic receptor agonist, [3-[(2R)-[[(2R)-(3-chlorophenyl)-2-hydroxyethyl]amino]propyl]-1H-indol-7-y loxy]acetic acid. Chem Pharm Bull (Tokyo) 53:184–198[Medline]
  10. Kato H, Ohue M, Kato K, Nomura A, Toyosawa K, Furutani Y, Kimura S, Kadowaki T 2001 Mechanism of amelioration of insulin resistance by ß3-adrenoceptor agonist AJ-9677 in the KK-Ay/Ta diabetic obese mouse model. Diabetes 50:113–122[Abstract/Free Full Text]
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