Thymidylate synthase

Adrucil is frequently used to treat a variety of cancers, such as breast and colorectal cancers. [1] 100 mg/kg Adrucil dramatically inhibits the growth of murine colon cancer tumors. Tumor-doubling time (TD), growth-delay factor (GDF), and T/C values of 26.5 days, 4.4, and 14% were observed in colon 38. [5]

Adrucil treatment for seven days in 96-well plates (4000 HT-29 cells/well in RPMI 1640 medium with 10% dialyzed fetal bovine serum) results in growth inhibition measurements; increasing Adrucil concentrations are added after allowing for cell attachment for an overnight period. Cells are washed five times with deionized water, fixed with 10% trichloroacetic acid for 60 minutes at 4 °C, and stained with 0.4% sulforhoda-mine B solution for 15 minutes at room temperature after three rounds of rinsing with phosphate-buffered saline (pH 7.4). Rinsing with 1% glacial acetic acid eliminates unstained sulforhodamine B. After that, dried and dissolved in 10 mM Tris-HCl are the stained cell proteins. Using a detector with a wavelength of 540 nm, the optical density value is determined.

Absorption, Distribution and Excretion
28-100%
Seven percent to 20% of the parent drug is excreted unchanged in the urine in 6 hours; of this over 90% is excreted in the first hour. The remaining percentage of the administered dose is metabolized, primarily in the liver.
Given by continuous iv infusion for 24 hr, plasma concn in range of 0.5 to 3.0 uM are obtained and urinary excretion of fluorouracil is only 4%.
Fluorouracil readily enters cerebrospinal fluid, and concn of about 7 uM are reached within 30 min after iv admin; values are sustained for approx 3 hr and subside slowly during period of 9 hr.
Fluorouracil crosses the placenta in rats.
Following iv administration of fluorouracil, no intact drug is detected in plasma after 3 hours.
For more Absorption, Distribution and Excretion (Complete) data for FLUOROURACIL (7 total), please visit the HSDB record page.

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Metabolism / Metabolites
Hepatic. The catabolic metabolism of fluorouracil results in degradation products ( e.g., CO2, urea and α-fluoro-ß-alanine) which are inactive.
A small portion of fluorouracil is anabolized in the tissues to 5-fluoro-2'-deoxyuridine and then to 5-fluoro-2'-deoxyuridine-5'-monophosphate, the active metabolite of the drug. The major portion of the drug is degraded in the liver. The metabolites are excreted as respiratory carbon dioxide and as urea, alpha-fluoro-beta-alanine, alpha-fluoro-beta-guanidopropionic acid, and alpha-fluoro-beta-ureidopropionic acid in urine. Following a single iv dose of fluorouracil, approximately 15% of the dose is excreted in urine as intact drug within 6 hours; over 90% of this is excreted in the first hour.
... Dihydropyrimidine dehydrogenase /is/ an NADPH-requiring homodimeric protein (Mr ~210 kDa) containing FMN/FAD, and an iron-sulfur cluster in each subunit. The enzyme is located mainly in liver cytosol, where it catalyzes the reduction of 5-fluorouracil and related pyrimidines ...
... Several routes are available for the formation of the 5'-monophosphate nucleotide (F-UMP) in animal cells. 5-FU may be converted to fluorouridine by uridine phosphorylase and then to F-UMP by uridine kinase, or it may react directly with 5-phosphoribosyl-1-pyrophosphate (PRPP), in a reaction catalyzed by ... orotate phosphoribosyl transferase, to form F-UMP. Many metabolic pathways are available to F-UMP, including incorporation into RNA. A reaction sequence crucial for antineoplastic activity involves reduction of the diphosphate nucleotide by the enzyme ribonucleoside diphosphate reductase to the deoxynucleotide level and the eventual formation of 5-fluoro-2'-deoxyuridine-5'-phosphate (F-dUMP). 5-FU also may be converted directly to the deoxyriboside 5-FUdR by the enzyme thymidine phosphorylase and further to F-dUMP, a potent inhibitor of thymidylate synthesis, by thymidine kinase ... The folate cofactor, 5,10-methylenetetrahydrofolate, and F-dUMP form a covalently bound ternary complex with the enzyme /thymidylate synthase/ ...
... Metabolic degradation /of 5-FU and floxuridine/ occurs in many tissues, particularly in liver. Floxuridine is converted by thymidine or deoxyuridine phosphorylases into 5-FU. 5-FU is inactivated by reduction of the pyrimidine ring; this reaction is carried out by dihydropyrimidine dehydrogenase (DPD), which is found in liver, intestinal mucosa, tumor cells, and other tissues ... Its metabolite, 5-fluoro-5,6-dihydrouracil ... is ultimately degraded to alpha-fluoro-beta-alanine ... Although the liver contains high concn of DPD, dosage does not have to be modified in patients with hepatic dysfunction, presumably because of degradation of the drug at extrahepatic sites or by vast excess of this enzyme in the liver ...
5-Fluorouracil is a known human metabolite of Tegafur.
Hepatic. The catabolic metabolism of fluorouracil results in degradation products ( e.g., CO2, urea and alpha-fluoro-beta-alanine) which are inactive.
Route of Elimination: Seven percent to 20% of the parent drug is excreted unchanged in the urine in 6 hours; of this over 90% is excreted in the first hour. The remaining percentage of the administered dose is metabolized, primarily in the liver.
Half Life: 10-20 minutes

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Biological Half-Life
10-20 minutes
Following iv administration, the plasma elimination half-life averages about 16 minutes (range: 8-20 minutes) and is dose dependent.
Rapid iv admin of 5-FU produces plasma concn of 0.1 to 1.0 mM; plasma clearance is rapid (half-life 10 to 20 min) ...

Toxicity Summary
The precise mechanism of action has not been fully determined, but the main mechanism of fluorouracil is thought to be the binding of the deoxyribonucleotide of the drug (FdUMP) and the folate cofactor, N5дус10-methylenetetrahydrofolate, to thymidylate synthase (TS) to form a covalently bound ternary complex. This results in the inhibition of the formation of thymidylate from uracil, which leads to the inhibition of DNA and RNA synthesis and cell death. Fluorouracil can also be incorporated into RNA in place of uridine triphosphate (UTP), producing a fraudulent RNA and interfering with RNA processing and protein synthesis.

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Toxicity Data
LD₅₀=230mg/kg (orally in mice)

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Interactions
To incr the complete response rate of patients with locally advanced head and neck cancer after 3 cycles of neoadjuvant chemotherapy, sequential methotrexate was added to the combination of cis-platin and continuous infusion fluorouracil. The feasibility of administering 3 additional cycles of the same regimen as adjuvant chemotherapy was examined. Thirty eight patients were treated; the median age was 53 yr and 36 patients had stage IV disease. Chemotherapy consisted of methotrexate 120 mg/sq m followed 24 hr later by cis-platin 100 mg/sq m and a 5 day continuous infusion of fluorouracil at 1000 mg/sq m/day. Of 34 patients evaluable for response to neoadjuvant chemotherapy, 9 had a complete response, 21 a partial response, 2 a minimal response, and 1 patient each stable disease and no response. Of 31 patients who received local therapy, 15 were treated with surgery and radiotherapy and 16 with radiotherapy alone. Of 25 patients eligible to receive adjuvant chemotherapy only 10 received all 3 intended cycles, while 15 received less or no adjuvant chemotherapy because of patient refusal, cumulative toxicity, or early disease progression. With a median follow up time of 39 mo, the median survival is estimated to be 20 mo. Of 8 patients with nasopharyngeal or paranasal sinus cancer, none had disease recurrence. Patients with good initial performance status and low N-stage also had a significant survival advantage. Chemotherapy-related toxicities consisted mainly of mucositis, requiring fluorouracil dose reduction in the majority of patients; similar toxicities were exacerbated in the adjuvant setting. The addition of methotrexate did not incr the complete response rate over what has been reported for the combination of cis-platin and fluorouracil alone.
Leukopenic and/or thrombocytopenic effects fluorouracil may be increased with concurrent or recent therapy with blood dyscrasia causing medications.
Concurrent use /with leucovorin/ may increase the therapeutic and toxic effects of fluorouracil.
Because normal defense mechanisms may be suppressed by fluorouracil therapy the patient's antibody responses to vaccine (killed virus) may be decreased.
For more Interactions (Complete) data for FLUOROURACIL (12 total), please visit the HSDB record page.

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Non-Human Toxicity Values
LD50 Dog oral 30 mg/kg
LD50 Mouse oral 115 mg/kg
LD50 Mouse iv 81 mg/kg
LD50 Mouse sc 169 mg/kg
For more Non-Human Toxicity Values (Complete) data for FLUOROURACIL (9 total), please visit the HSDB record page.

[1]. Nat Rev Cancer . 2003 May;3(5):330-8.

[2]. Biochem Biophys Res Commun . 2008 Jul 4;371(3):531-5.

[3]. Oncology . 2002;62(4):354-62.

[4]. J Biol Chem . 1995 Aug 11;270(32):19073-7.

[5]. Cancer Chemother Pharmacol . 1996;39(1-2):79-89.

Therapeutic Uses
Antimetabolites; Antimetabolites, Antineoplastic; Immunosuppressive Agents
Fluorouracil is indicated for palliative treatment of carcinoma of the colon, rectum, breast, stomach, and pancreas in patients considered to be incurable by surgery or other means. /Included in US product update/
Fluorouracil is also indicated for treatment of bladder carcinoma, prostatic carcinoma, epithelial ovarian carcinoma, cervical carcinoma, endometrial carcinoma, anal carcinoma, esophageal carcinoma,metastatic tumors of skin carcinoma, and hepatoblastoma, and is used by intra-arterial injection for treatment of hepatic tumors and head and neck tumors. /Not included in US product label/
Fluorouracil, in combination therapy, is reasonable medical therapy at some point in the management of adrenocortical carcinoma, vulvar carcinoma, penile carcinoma and carcinoid tumors (gastrointestinal and neuroendocrine tumors). /Not included in US product label/
For more Therapeutic Uses (Complete) data for FLUOROURACIL (12 total), please visit the HSDB record page.

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Drug Warnings
Anorexia and nausea are common adverse effects of fluorouracil, and vomiting occurs frequently. These reactions generally occur during the first week of therapy, can often be alleviated by antiemetics, and generally subside within 2 or 3 days following therapy. Stomatitis is one of the most common and often the earliest sign of specific toxicity, appearing as early as the fourth day but more commonly on the fifth to eighth day of therapy. Diarrhea, which also occurs frequently, usually appears slightly later than stomatitis, but may occur concurrently or even in the absence of stomatitis. Esophagitis, proctitis, and GI ulceration and bleeding have been reported, and paralytic ileus occurred in two patients who received excessive dosage. Patients must be closely monitored for adverse GI effects.
Leukopenia, predominantly of the granulocytopenic type, thrombocytopenia, and anemia occur commonly with fluorouracil therapy; leukopenia usually occurs after an adequate course of fluorouracil therapy. Pancytopenia and agranulocytosis also have occurred. The patient's hematologic status must be carefully monitored. The nadir of the white blood cell count usually occurs from the ninth to the fourteenth day after therapy is initiated but may occur as late as the 25th day after the first dose of fluorouracil. Maximum thrombocytopenia has been reported to occur from the seventh to seventeenth day of therapy. Hematopoietic recovery is usually rapid and by the thirtieth day, blood cell counts have usually reached the normal range.
Hair loss occurs frequently with fluorouracil therapy, and cosmetically significant alopecia has occurred in a substantial number of patients. Regrowth of hair has been reported even in patients receiving repeated courses of the drug. Partial loss of nails has occurred rarely, and diffuse melanosis of the nails has been reported. The most common type of dermatologic toxicity is a pruritic maculopapular rash which usually appears on the extremities and less frequently on the trunk. This rash is generally reversible and usually responsive to symptomatic treatment.
An erythematous, desquamative rash involving the hands and feet has been reported in patients receiving fluorouracil (in some cases, prolonged infusions of high dosages of the drug were administered). The rash may be accompanied by tingling or painful hands and feet, swollen palms and soles, and phalangeal tenderness. These adverse effects, referred to as palmar-plantar erythrodysesthesia or hand-foot syndrome, may gradually disappear over 5-7 days after discontinuance of fluorouracil therapy.
For more Drug Warnings (Complete) data for FLUOROURACIL (31 total), please visit the HSDB record page.

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Pharmacodynamics
Fluorouracil is an antineoplastic anti-metabolite. Anti-metabolites masquerade as purine or pyrimidine - which become the building blocks of DNA. They prevent these substances from becoming incorporated into DNA during the "S" phase (of the cell cycle), stopping normal development and division. Fluorouracil blocks an enzyme which converts the cytosine nucleotide into the deoxy derivative. In addition, DNA synthesis is further inhibited because Fluorouracil blocks the incorporation of the thymidine nucleotide into the DNA strand.

C4H3FN2O2

130.08

130.017

C, 36.93; H, 2.32; F, 14.61; N, 21.54; O, 24.60

51-21-8

3385

White to off-white solid powder

1.7±0.1 g/cm3

401.4±48.0 °C at 760 mmHg

282-286 °C (dec.)(lit.)

196.5±29.6 °C

0.0±1.0 mmHg at 25°C

1.596

-2.1

2

3

0

9

199

0

FC1=C([H])N([H])C(N([H])C1=O)=O

GHASVSINZRGABV-UHFFFAOYSA-N

InChI=1S/C4H3FN2O2/c5-2-1-6-4(9)7-3(2)8/h1H,(H2,6,7,8,9)

5-fluoro-1H-pyrimidine-2,4-dione

NSC 19893; 5-FU; Fluorouracil; NSC-19893; NSC19893; 5-Fluorouracil; 5-Fluorouracil; 5FU; Fluoroplex; Efudex; Adrucil; Carac; Trade name: Adrucil among many others.

2934.99.9001

Powder      -20°C    3 years

                     4°C     2 years

In solvent   -80°C    6 months

                  -20°C    1 month

Note: This product requires protection from light (avoid light exposure) during transportation and storage.

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 (19.22 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 (19.22 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.

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Solubility in Formulation 3: ≥ 2.5 mg/mL (19.22 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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Solubility in Formulation 4: ≥ 2.5 mg/mL (19.22 mM) (saturation unknown) in 5% DMSO + 40% PEG300 + 5% Tween80 + 50% Saline (add these co-solvents sequentially from left to right, and one by one), clear solution.
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 5: ≥ 2.5 mg/mL (19.22 mM) (saturation unknown) in 5% DMSO + 95% (20% SBE-β-CD in Saline) (add these co-solvents sequentially from left to right, and one by one), clear solution.
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.

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Solubility in Formulation 6: 2.5 mg/mL (19.22 mM) in 5% DMSO + 95% Saline (add these co-solvents sequentially from left to right, and one by one), clear solution; with ultrasonication.
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 7: Saline: 10mg/mL

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