Neuroblastoma (NB) cells are cytotoxic to tolcapone; its IC50 values range from 32.27 μM for SMS-KCNR cells to 219.8 μM for primary MGT9-102-08 cells[3]. In NB cells, tolcapone (25, 50, 75, and 100 μM) treatment triggers subsequent apoptotic processes. In neuroblastoma, tolcapone triggers caspase-mediated apoptosis[3].

Oral tolcapone (125 mg/kg) suppresses the growth of tumors and increases in vivo survival. The mice's weight or behavior have not changed, nor have any unfavorable events been reported[3].

Cell Viability Assay[3]
Cell Types: BE(2)-C, SMS-KCNR, CHLA-90, SH-SY5Y, MGT-015 -08 and MGT9-102-08
Tested Concentrations: 1.5625~400 μM
Incubation Duration: 48 hrs (hours)
Experimental Results: IC50s of 32.27, 72.31, 80.29, 109.4, 174.6, 219.8 μM for SMS-KCNR, SH-SY5Y, BE(2)-C, CHLA-90, MGT-015-08 and MGT9-102-08, respectively.

---

Cell Viability Assay[3]
Cell Types: NB
Cell Types: BE(2)-C, SMS-KCNR, CHLA-90, SH-SY5Y, MGT-015-08 and MGT9-102-08
Tested Concentrations: 25, 50, 75, 100 μM
Incubation Duration:
Experimental Results: A dose-dependent increase in cleaved caspase-3 and cleaved PARP protein in all six NB cell lines and a subsequent decrease in whole caspase-3 and whole PARP protein.

Animal/Disease Models: 4weeks old female nude mice (nu /nu) bearing SMS‐KCNR xenograft models [3]
Doses: 125 mg/kg
Route of Administration: Treated orally every 24 h for 35 days
Experimental Results: diminished tumor volume compared to control. Resulted in a smaller average tumor of 490±310 mm3 compared to control tumors of 1100±450 mm3.

Absorption, Distribution and Excretion
Rapidly absorbed (absolute bioavailability is about 65%)
Tolcapone is almost completely metabolized prior to excretion, with only a very small amount (0.5% of dose) found unchanged in urine. The glucuronide conjugate of tolcapone is mainly excreted in the urine but is also excreted in the bile.
9 L
7 L/h

---

Metabolism / Metabolites
The main metabolic pathway of tolcapone is glucuronidation
Tolcapone has known human metabolites that include (2S,3S,4S,5R)-3,4,5-trihydroxy-6-[2-hydroxy-4-(4-methylbenzoyl)-6-nitrophenoxy]oxane-2-carboxylic acid.
The main metabolic pathway of tolcapone is glucuronidation.
Route of Elimination: Tolcapone is almost completely metabolized prior to excretion, with only a very small amount (0.5% of dose) found unchanged in urine. The glucuronide conjugate of tolcapone is mainly excreted in the urine but is also excreted in the bile.
Half Life: 2-3.5 hours

---

Biological Half-Life
2-3.5 hours

Toxicity Summary
Tolcapone hepatoxicity can be attributed to elevated levels of transminases, but studies have shown that minimal risk exists for those without preexisting liver conditions when their enzyme levels were being monitored. No clear mechanism is implicated in tolcapone induced liver toxicity, but it has been hypothesized that it has something to do with abnormal mitochondrial respiration due to the uncoupling of oxidative phosphorylation. Dyskinesia occurs because the administration of Tolcapone results in the accumulation of the biological methyl donor S-adenosyl-L-methionine (SAM) in the striatum that works to induce symptoms of Parkinson's disease. (Wikipedia)

---

Hepatotoxicity
Tolcapone has been reported to cause serum aminotransferase elevations above 3 times the upper limit of normal in 1% to 5% of patients. While these abnormalities are usually asymptomatic and self-limiting, some persist if therapy is continued and resolved only with stopping tolcapone. More importantly, tolcapone has been implicated in several cases of severe, clinically apparent acute liver injury and at least three cases of death from acute liver failure. The onset of injury was insidious, arising 1 to 5 months after starting treatment. The pattern of serum enzyme elevations was hepatocellular and the clinical phenotype was similar to acute viral hepatitis. Immunoallergic manifestations were not present, but some patients had autoantibodies of unclear significance. Because of these reports, regular monitoring of serum aminotransferase levels has been mandated (every 2 to 4 weeks for the first 6 months of treatment and as clinically indicated thereafter) during tolcapone therapy, and treatment should be promptly discontinued if ALT or AST levels rise above twice the upper limit of the normal range or if signs or symptoms of liver injury are present.
Likelihood score: C (probable cause of clinically apparent liver injury).

---

Effects During Pregnancy and Lactation
◉ Summary of Use during Lactation
No information is available on the use of tolcapone during breastfeeding. An alternate drug may be preferred, especially while nursing a newborn or preterm infant.
◉ Effects in Breastfed Infants
Relevant published information was not found as of the revision date.
◉ Effects on Lactation and Breastmilk
Relevant published information was not found as of the revision date.

---

Protein Binding
> 99.9% (to serum albumin)

---

Toxicity Data
LD50: 1600 mg/kg (Oral, Rats) (A308)

[1]. Eur J Clin Pharmacol. 1998 May;54(3):215-9.

[2]. Entacapone and tolcapone, two catechol O-methyltransferase inhibitors, block fibril formation of alpha-synuclein and beta-amyloid and protect against amyloid-induced toxicity. J Biol Chem. 2010 May 14;285(20):14941-14954.

[3]. Tolcapone induces oxidative stress leading to apoptosis and inhibition of tumor growth in Neuroblastoma.Cancer Med. 2017 Jun;6(6):1341-1352.

Pharmacodynamics
Tolcapone is a potent, selective, and reversible inhibitor of catechol-O-methyltransferase (COMT). In humans, COMT is distributed throughout various organs. COMT catalyzes the transfer of the methyl group of S-adenosyl-L-methionine to the phenolic group of substrates that contain a catechol structure. Physiological substrates of COMT include dopa, catecholamines (dopamine, norepinephrine, epinephrine) and their hydroxylated metabolites. The function of COMT is the elimination of biologically active catechols and some other hydroxylated metabolites. COMT is responsible for the elimination of biologically active catechols and some other hydroxylated metabolites. In the presence of a decarboxylase inhibitor, COMT becomes the major metabolizing enzyme for levodopa catalyzing it to 3-methoxy-4-hydroxy-L-phenylalanine (3-OMD) in the brain and periphery. When tolcapone is given in conjunction with levodopa and an aromatic amino acid decarboxylase inhibitor, such as carbidopa, plasma levels of levodopa are more sustained than after administration of levodopa and an aromatic amino acid decarboxylase inhibitor alone. It is believed that these sustained plasma levels of levodopa result in more constant dopaminergic stimulation in the brain, leading to greater effects on the signs and symptoms of Parkinson's disease in patients as well as increased levodopa adverse effects, sometimes requiring a decrease in the dose of levodopa.

C14H11NO5

273.24

273.063

134308-13-7

Tolcapone-d7;Tolcapone-d4;1246816-93-2

4659569

Light yellow to yellow solid powder

1.4±0.1 g/cm3

485.6±45.0 °C at 760 mmHg

126-128ºC

205.7±17.2 °C

0.0±1.3 mmHg at 25°C

1.661

4.07

2

5

2

20

372

0

MIQPIUSUKVNLNT-UHFFFAOYSA-N

InChI=1S/C14H11NO5/c1-8-2-4-9(5-3-8)13(17)10-6-11(15(19)20)14(18)12(16)7-10/h2-7,16,18H,1H3

(3,4-dihydroxy-5-nitrophenyl)-(4-methylphenyl)methanone

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 (9.15 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.

---

Solubility in Formulation 2: ≥ 2.5 mg/mL (9.15 mM) (saturation unknown) in 10% DMSO + 90% (20% SBE-β-CD in 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 900 μL of 20% SBE-β-CD physiological saline solution and mix evenly.
Preparation of 20% SBE-β-CD in Saline (4°C，1 week): Dissolve 2 g SBE-β-CD in 10 mL saline to obtain a clear solution.

---

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