Tryptophan hydroxylase
In vitro activity: Telotristat (formerly known as LP-778902) is the active metabolite of LX1606 (Telotristat etiprate) which is an orally bioavailable, small-molecule, tryptophan hydroxylase (TPH) inhibitor with potential an in vivo IC50 of 0.028 μM and with antiserotonergic activity. Telotristat has activity in controlling diarrhea associated with carcinoid syndrome. Telotristat acts by inhibiting the enzyme tryptophan hydoxylase (TPH) and reduces serotonin production both inside and outside the GI tract without affecting brain serotonin levels. Blocking peripheral serotonin synthesis by telotristat reduces severity of both chemical- and infection-induced intestinal inflammation.

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Kinase Assay: Telotristat (formerly known as LP-778902) is the active metabolite of LX1606 (Telotristat etiprate) which is an orally bioavailable, small-molecule, tryptophan hydroxylase (TPH) inhibitor with potential an in vivo IC50 of 0.028 μM and with antiserotonergic activity.

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Cell Assay: BON CBA cells are grown in equal volume of DMEM and F12K with 5% bovine serum for 3-4 hours (20 K cell/well) and telotristat is added at a concentration range of 0.07 to 50 μM. The cells are incubated at 37°C overnight. 50 μM of the culture supernatant is then taken for 5HTP measurement. The supernatant is mixed with equal volume of 1M TCA, then filtered through glass fiber. The filtrate is loaded on reverse phase HPLC for 5HTP concentration measurement. The cell viability is measured by treating the remaining cells with Celltiter-Glo Luminescent Cell Viability Assay.

In vitro activity: Telotristat (formerly known as LP-778902) is the active metabolite of LX1606 (Telotristat etiprate) which is an orally bioavailable, small-molecule, tryptophan hydroxylase (TPH) inhibitor with potential an in vivo IC50 of 0.028 μM and with antiserotonergic activity. Telotristat has activity in controlling diarrhea associated with carcinoid syndrome. Telotristat acts by inhibiting the enzyme tryptophan hydoxylase (TPH) and reduces serotonin production both inside and outside the GI tract without affecting brain serotonin levels. Blocking peripheral serotonin synthesis by telotristat reduces severity of both chemical- and infection-induced intestinal inflammation.

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Kinase Assay: Telotristat (formerly known as LP-778902) is the active metabolite of LX1606 (Telotristat etiprate) which is an orally bioavailable, small-molecule, tryptophan hydroxylase (TPH) inhibitor with potential an in vivo IC50 of 0.028 μM and with antiserotonergic activity.

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Cell Assay: BON CBA cells are grown in equal volume of DMEM and F12K with 5% bovine serum for 3-4 hours (20 K cell/well) and telotristat is added at a concentration range of 0.07 to 50 μM. The cells are incubated at 37°C overnight. 50 μM of the culture supernatant is then taken for 5HTP measurement. The supernatant is mixed with equal volume of 1M TCA, then filtered through glass fiber. The filtrate is loaded on reverse phase HPLC for 5HTP concentration measurement. The cell viability is measured by treating the remaining cells with Celltiter-Glo Luminescent Cell Viability Assay.

The peripheral but not the brain of the mice is less serotonin-containing when telotristat ethyl (15, 50, 150, 300 mg/kg, po, qd) is administered. Telotristat ethyl (200 mg/kg po, qd) significantly protects against TNBS-induced elevation in blood neutrophil counts in a rat model of inflammatory bowel disease. The mouse IBD model is protected by telotristat ethyl (200 mg/kg po, qd), as shown by histopathological evaluation[1]. In the jejunum, telotristat ethyl (15, 50, 150, and 300 mg/kg) depletes 5-HT, but not in the brain. However, neither the constitutive gastrointestinal motility in mice nor the depletion of enteric neuronal serotonin (5-HT) are caused by telotristat ethyl (200 mg/kg, po). The severity of colitis caused by trinitrobenzene sulfonic acid (TNBS) is lessened by telotristat ethyl (200 mg/kg)[2].

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In mice treated orally with LX1606 at 200 mg/kg once daily for 6 days, the concentration of serotonin (5-HT) in whole blood was significantly reduced compared to vehicle-treated controls (p<10⁻⁶).
In a TNBS-induced inflammatory bowel disease (IBD) mouse model, pretreatment and continued treatment with oral LX1606 (200 mg/kg, qd) starting 6 days before TNBS challenge resulted in a significant reduction in body weight loss compared to the vehicle group (p<0.03).
LX1606 treatment significantly lowered the histopathology score (p=0.04) and reduced the percentage of animals with a pathology score ≥10.
It also prevented the increase in blood neutrophil counts that was observed in the vehicle group after TNBS challenge.
Quantitative PCR analysis of distal colon tissues from LX1606-treated mice showed a trend toward decreased expression of proinflammatory cytokines IL-1β, IL-6, and IL-17α after TNBS challenge.
Treatment with LX1606 (200 mg/kg) reduced serotonin content in the small intestine in a dose-dependent manner but did not affect serotonin levels in the brain. [1]

Dissolved in 15% cyclodextrin or 0.25% methylcellulose; 300 mg/kg; p.o.Male C57BL/6 mice and male C57 albino mice. In normal mice, telotristat etiprate (administered once daily for 4 days at doses of 15–300 mg/kg/day) was found to reduce serotonin levels throughout the gastrointestinal tract. These reductions occurred in a dose dependent fashion with maximal effects observed with doses of telotristat etiprate ≥150 mg/kg. No significant change in brain serotonin or 5-hydroxyindoleacetic acid (5-HIAA, a serotonin metabolite) was observed. Similar findings were seen in Sprague-Dawley rats. Gastrointestinal motility studies were conducted in rats using the charcoal meal test. There was a significant dose-related delay in both gastrointestinal transit and gastric emptying, associated with a reduction in blood serotonin levels and proximal colon serotonin. A quantitative whole-body autoradiography study was conducted to assess the absorption, distribution and excretion of radioactivity in rats following a single oral dose of telotristat etiprate labeled with carbon 14. Rats were administered either 30 mg/kg or 100 mg/kg of the compound. The distribution of radioactivity was limited to tissues of the hepatic and renal system and the contents of the GI tract. There was no measurable radioactivity in the brain at any dose tested.

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C57Bl/6brd x 129SvEv F1 hybrid mice were used in all experiments.
For the TNBS-IBD model, animals were challenged via intra-rectal administration with 2% TNBS solution. Naïve control mice were left untreated.
LX1606 and the reference drug sulfasalazine were formulated in 0.25% methylcellulose and administered to mice once daily via oral gavage.
Dosing began 6 days before TNBS challenge and continued throughout the challenge period.
In the efficacy study, mice (n=10 per group initially) were treated with vehicle, LX1606 (200 mg/kg, po, qd), or sulfasalazine (100 mg/kg, po, qd). Body weight was monitored.
For serotonin measurement, separate groups of mice (n=5 per group) were treated with indicated doses of LX1606 for 6 days before analysis.
Blood was collected for neutrophil counts via retro-orbital bleeding into EDTA tubes, and complete cell counts were measured.
At the endpoint, proximal and distal colon and cecum were collected, fixed in formalin, sectioned, and stained with hematoxylin and eosin for histopathological scoring using a modified scoring system. [1]

Absorption, Distribution and Excretion
Following a single oral administration of terlotrexate ethyl ester to healthy subjects, terlotrexate ethyl ester is absorbed and metabolized to the active metabolite terlotrexate. Peak plasma concentrations of terlotrexate ethyl ester are reached within 0.5 to 2 hours, while peak plasma concentrations of terlotrexate are reached within 1 to 3 hours. Thereafter, plasma concentrations decline in a biphasic manner. In healthy subjects, after a single fasting administration of 500 mg terlotrexate ethyl ester (twice the recommended dose), the mean Cmax and AUC0-inf of terlotrexate ethyl ester were 4.4 ng/mL and 6.23 ng•hr/mL, respectively. The mean Cmax and AUC0-inf of terlotrexate were 610 ng/mL and 2320 ng•hr/mL, respectively. In a fasting state, after a single dose of 100 mg (0.4 times the lowest recommended dose to 1000 mg [4 times the highest recommended dose]) of telotrelstat ethyl ester, the peak plasma concentrations and AUC of both telotrelstat ethyl ester and telotrelstat appeared to be dose-proportional. No significant steady-state accumulation of either telotrelstat ethyl ester or telotrelstat was observed after three daily doses of 500 mg telotrelstat ethyl ester. In patients with metastatic neuroendocrine tumors complicated by carcinoid syndrome diarrhea treated with somatostatin analogues (SSA), the median time to peak concentration (Tmax) of telotrelstat ethyl ester and telotrelstat was approximately 1 hour and 2 hours, respectively. After patients took 500 mg telotrelstat ethyl ester three times daily with food, the mean Cmax and AUC0–6hr of telotrelstat ethyl ester were approximately 7 ng/mL and 22 ng•hr/mL, respectively. The mean Cmax and AUC0-6hr of telotrexat were approximately 900 ng/mL and 3000 ng•hr/mL, respectively. The pharmacokinetic parameters of both telotrexat ethyl ester and telotrexat were highly variable, with a coefficient of variation of approximately 55%. Following a single oral dose of 500 mg 14C-telotrexat ethyl ester, 93.2% of the dose was recovered within 240 hours: 92.8% was recovered in feces, and less than 0.4% was recovered in urine. Based on population pharmacokinetic models, the apparent total volume of distribution (VDC) of the active metabolite was estimated to be 428.1 L in typical healthy, fasting subjects and 348.7 L in patients with carcinoid syndrome.
In healthy subjects, oral administration of 500 mg telotralstat ethyl ester three times daily for 14 days (twice the recommended dose) resulted in steady-state apparent total clearance (CL/Fss) of 2.7 L/hr and 152 L/hr, respectively. These refer to telotralstat ethyl ester and telotralstat, respectively.
Metabolism/Metabolites

After oral administration, telotralstat ethyl ester is hydrolyzed by carboxylesterases to its active metabolite, telotralstat. Telotralstat is further metabolized. Among the metabolites of telotralstat, the systemic exposure of the acidic metabolite of oxidative deamination and decarboxylation of telotralstat is approximately 35% of that of telotralstat. In vitro data indicate that telotralstat ethyl ester and telotralstat are not substrates of CYP enzymes.

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Biological Half-Life
After a single oral dose of 500 mg telotristat ethyl ester in healthy subjects, the apparent half-life of telotristat ethyl ester is approximately 0.6 hours, and the apparent half-life of telotristat is approximately 5 hours.

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Oral administration of telotristat ethyl ester (LX1606) significantly reduces 5-HT levels in the gut and blood, but has no effect on 5-HT levels in brain tissue, indicating that the compound can be absorbed and exerts its effects peripherally, but cannot cross the blood-brain barrier.
The manuscript notes that peripheral TPH inhibitors such as telotristat ethyl ester (LX1606) cannot cross the blood-enteric plexus barrier, as evidenced by the maintenance of 5-HT immunoreactivity in enteric neurons.
Specific Pharmacokinetic Parameters The half-life, oral bioavailability, Cmax, AUC, and other parameters of Telotristat Etiprate (LX1606) are not provided in this paper. [2]

Protein Binding
Both telotristat ethyl and telotristat bind to human plasma proteins at rates greater than 99%. In the TNBS-IBD study, 10 mice were used in each group. Four mice died at the end of the experiment in the control group (carrier group), while no deaths occurred in the LX1606 treatment group or the sulfasalazine treatment group. The cause of death in the carrier group may be related to severe IBD. [1]

[1]. LX1606 (aka LX1032), a Novel Inhibitor of Serotonin Synthesis, Alleviates Development of Inflammatory Bowel Disease in a Preclinical Model.

[2]. Pharmacological reduction of mucosal but not neuronal serotonin opposes inflammation in mouse intestine. Gut, 2013.

Tiloblastal ethyl ester, a prodrug of telotraststat, was approved by the FDA in March 2017 under the brand name Xermelo. It was previously known as telotraststat acetate, the hippurate form; however, the FDA recommended using the name of the neutral form rather than the salt name. Currently, telotraststat ethyl ester is used to treat carcinoid syndrome diarrhea caused by neuroendocrine tumors that are poorly controlled with short-acting somatostatin analogues (SSAs). Neuroendocrine cells are cells that secrete regulatory peptides and biogenic amines in response to chemical, neural, or other types of stimulation. Therefore, neuroendocrine tumors (NETs) originating from these cells can secrete chemical mediators into the bloodstream, causing side effects at distant sites—a phenomenon known as carcinoid syndrome. The most common peptides and amines secreted by neuroendocrine tumors (NETs) include histamine, tachykinin, kallikrein, and serotonin. Excessive serotonin can lead to severe diarrhea, one of the main clinical symptoms of carcinoid syndrome. Serotonin is metabolized and excreted in the urine as the metabolite 5-hydroxyindoleacetic acid (u5-HIAA), and high levels of u5-HIAA are associated with poor survival in patients with carcinoid syndrome. First-line treatment for diarrhea in carcinoid syndrome is somatostatin analogues (SSA), but symptoms can recur during the course of the disease. Telotristat Ethyl is the ethyl ester form of telotristat, a tryptophan hydroxylase (TPH) inhibitor with potential antiserotonin activity. After administration, telotristat binds to TPH and inhibits its activity. This may lead to reduced peripheral serotonin (5-HT) production and improve serotonin-mediated gastrointestinal effects, such as severe diarrhea. Tryptophan hydroxylase (TPH) is the rate-limiting enzyme in serotonin biosynthesis and is overexpressed in carcinoid tumor cells. See also: terlotrexate (with the active fraction).

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Drug Indications
Xermelo is indicated for the treatment of adult cancer-like syndrome diarrhea in combination with somatostatin analogues (SSA), particularly in patients who have not responded well to SSA therapy.
Xermelo is indicated for the treatment of adult cancer-like syndrome diarrhea in combination with somatostatin analogues (SSA), particularly in patients who have not responded well to SSA therapy.

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Mechanism of Action
Telotristat is the active metabolite of telotristat ethyl ester, an inhibitor of tryptophan hydroxylase, which mediates the rate-limiting step in serotonin biosynthesis. In vitro studies have shown that telotristat is 29 times more potent in inhibiting tryptophan hydroxylase than telotristat ethyl ester. Serotonin plays a role in regulating gastrointestinal secretion, motility, inflammation, and sensation, and is overproduced in patients with carcinoid syndrome.
Tilolostat and tilolostat ethyl ester reduce peripheral serotonin production and decrease the frequency of carcinoid syndrome diarrhea by inhibiting tryptophan hydroxylase.

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Pharmacodynamics
In normal mice, tilolostat ethyl ester (once daily for 4 days, at a dose of 15–300 mg/kg/day) reduced serotonin levels throughout the gastrointestinal tract. These reductions were dose-dependent, with the maximum effect observed at tilolostat ethyl ester doses ≥150 mg/kg. No significant changes in brain serotonin or 5-hydroxyindoleacetic acid (5-HIAA, a serotonin metabolite) were observed. Similar results were observed in Sprague-Dawley rats. Gastrointestinal motility studies in rats were performed using an activated charcoal meal test. Results showed significant dose-dependent delays in both gastrointestinal transit and gastric emptying, accompanied by decreases in blood and proximal colonic serotonin levels. To evaluate the absorption, distribution, and excretion of radioactive material after a single oral administration of carbon-14 labeled telotristat etepeptate in rats, a quantitative whole-body autoradiography study was conducted. Rats were administered the compound at doses of 30 mg/kg or 100 mg/kg. The distribution of radioactive material was limited to the liver and kidney tissues and gastrointestinal contents. No radioactivity was detected in brain tissue at any of the tested doses.

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LX1606 (also known as LX1032, later renamed Telotristat ethyl) is a novel oral tryptophan hydroxylase inhibitor designed to reduce peripheral serotonin production without affecting central nervous system levels.
It is considered a potential new approach for treating serotonin-mediated symptoms in diseases such as inflammatory bowel disease.
Its mechanism of action is to inhibit TPH1, thereby reducing tryptophan-mediated serotonin synthesis in intestinal chromaffin cells. [1]

C27H26CLF3N6O3

574.99

574.17

C, 56.40; H, 4.56; Cl, 6.17; F, 9.91; N, 14.62; O, 8.35

1033805-22-9

Telotristat etiprate;1137608-69-5;Telotristat;1033805-28-5; 1033805-22-9 (ethyl); 1374745-52-4 (besilate)

25181577

Typically exists as Light yellow to yellow solids at room temperature

1.4±0.1 g/cm3

704.7±70.0 °C at 760 mmHg

380.0±35.7 °C

0.0±2.2 mmHg at 25°C

1.621

5.47

2

11

10

40

821

2

N[C@@H](CC1=CC=C(C2=NC(N)=NC(O[C@H](C3=CC=C(Cl)C=C3N4N=C(C)C=C4)C(F)(F)F)=C2)C=C1)C(OCC)=O

MDSQOJYHHZBZKA-GBXCKJPGSA-N

InChI=1S/C27H26ClF3N6O3/c1-3-39-25(38)20(32)12-16-4-6-17(7-5-16)21-14-23(35-26(33)34-21)40-24(27(29,30)31)19-9-8-18(28)13-22(19)37-11-10-15(2)36-37/h4-11,13-14,20,24H,3,12,32H2,1-2H3,(H2,33,34,35)/t20-,24+/m0/s1

ethyl (2S)-2-amino-3-[4-[2-amino-6-[(1R)-1-[4-chloro-2-(3-methylpyrazol-1-yl)phenyl]-2,2,2-trifluoroethoxy]pyrimidin-4-yl]phenyl]propanoate

LX 1032; LX 1606; LX-1032; LX1606; LX1032; LX-1606; LX1606; LX 1606; Xermelo; Telotristat ethyl [USAN]; trade name: Xermelo

2934.99.9001

Powder      -20°C    3 years

                     4°C     2 years

In solvent   -80°C    6 months

                  -20°C    1 month

Room temperature (This product is stable at ambient temperature for a few days during ordinary shipping and time spent in Customs)

Solubility in Formulation 1: ≥ 2.5 mg/mL (4.35 mM) (saturation unknown) in 10% DMSO + 40% PEG300 + 5% Tween80 + 45% Saline (add these co-solvents sequentially from left to right, and one by one), clear solution.
For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 25.0 mg/mL clear DMSO stock solution to 400 μL PEG300 and mix evenly; then add 50 μL Tween-80 to the above solution and mix evenly; then add 450 μL normal saline to adjust the volume to 1 mL.
Preparation of saline: Dissolve 0.9 g of sodium chloride in 100 mL ddH₂ O to obtain a clear solution.

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Solubility in Formulation 2: ≥ 2.5 mg/mL (4.35 mM) (saturation unknown) in 10% DMSO + 90% Corn Oil (add these co-solvents sequentially from left to right, and one by one), clear solution.
For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 25.0 mg/mL clear DMSO stock solution to 900 μL of corn oil and mix evenly.

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 (Please use freshly prepared in vivo formulations for optimal results.)