DNA Alkylator

Carmustine is a chemotherapy drug used to treat cancer. Neuronal cell proliferation, tumor cytoplasm, and intact N-benzoyltransferase (NAT) activity of 2-aminobenzoic acid (AF) and p-aminobenzoic acid (PABA) are all reduced by carmustine (8, 80, and 800 μM). The DNA-AF addition complex rises with the development of tumor nerve growth cells, while carmustine lowers it [1].

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Carmustine and lomustine are nitrosourea antitumor chemotherapeutic agents which were used to determine whether or not they could affect arylamine N-acetyltransferase (NAT) activity and DNA-2-aminofluorene adducts in rat glial tumor cell line (C6 glioma). The NAT activity was measured by high preformance liquid chromatography (HPLC) assaying for the amounts of N-acetyl-2-aminofluorene (AAF) and N-acetyl-p-aminobenzoic acid (N-Ac-PABA) and remaining 2-aminofluorene (AF) and p-aminobenzoic acid (PABA). The results indicate that NAT activity in glial tumor cell cytosols and intact tumor cells were decreased by carmustine and lomustine in a dose-dependent manner. The apparent values of Km and Vmax of NAT from rat glial tumor cell also decreased after co-treatment of carmustine and lomustine in both examined cytosols and intact cells. Following exposure of glial tumor cells to the various concentrations of AF with or without co-treatment with carmustine and lomustine, DNA-AF adducts were determined by using gamma-[32p]-dATP and HPLC. The DNA-AF adducts in rat glial tumor cells were decreased by co-treatment with carmustine and lomustine. This report is the first demonstration to show carmustine and lomustine did inhibit rat glial tumor cells NAT activity and DNA-AF adduct formation.[1]

In comparison to stents level (GSSG) and reduced glutathione (GSH)/GSSG value, carmustine (BCNU; 25 mg/kg, ip) led to greater levels of death to body weight, the ratio of bound bilirubin, external bile flow, and oxidized glutathione [2].

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This study investigated the effect of trimetazidine (TMZ), known as an anti-oxidant agent, on intrahepatic cholestasis caused by Carmustine (BCNU) in rats. Rats were assigned into four groups. The first group (Saline) consisted of 12 rats, which were injected with 2 ml/kg of saline intraperitoneally (IP) 48 h before the study. The second group (corn oil group, n=15), which were injected with 2 ml/kg of corn oil IP 48 h before the study. The third group (BCNU group, n=16), which were injected with 2 ml/kg of corn oil+25 mg/kg BCNU IP 48 h before the study. The fourth group (TMZ group, n=12), which were injected with 2.5 mg/kg per day of TMZ IP, administered at the same hour of the day as a single-dose. Twelve hour after the first dose of TMZ, corn oil 2 ml/kg+BCNU 25 mg/kg IP were injected, and the rats were included in the study 48 h after the administration of corn oil+BCNU. Following a pentobarbital anaesthesia, abdomen was opened with incision, a cannula was placed into the channel of choledocus, and the amount of bile was measured per hour. Then intracardiac blood sample was taken, and consequently centrifuged to obtain the plasma. Finally, the rats were killed with cervical dislocation, and their livers were removed and weighted. In addition to histopathological examination of liver, the levels of malon dialdehyde (MDA), oxidised glutation (GSSG), and reduced glutation (GSH) were detected. Also the osmolality of bile and plasma was estimated in mOsm/kg. As a result, the biliary flow was seen to decrease in BCNU group (P<0.005), but to be normal in TMZ group. The serum level of conjugated biluribin was higher in BCNU group compared to other groups (P<0.05 for each). Although the level of total glutation was lower (P<0.005) in TMZ group, GSH/GSSG ratio was normal. These findings suggest that TMZ has a protective effect on intrahepatic cholestasis caused by BCNU.[2]

2-Aminofluorene (AF) and p-Aminobenzoic acid (PABA) N-acetylation is determined in an Acetyl-CoAdependent manner. The assay system's incubation mixtures have a total volume of 90 μL and include glial tumor cells cytosols, diluted as needed, in 50 μL of lysis buffer (20 mM Tris/HCl, pH 7.5, 1 mM DTT and 1 mM EDTA), 20 μL of an Acetyl-CoA recycling mixture of 50 mM Tris-HCl (pH7.5), 0.2 mM EDTA, 2 mM DTT, 15 mM acetylcamitine, 2U/mL carnitine acetyltransferase, and AF or PABA at designated concentrations. Addition of 20 μL of acetyl-CoA initiates the reactions. Acetyl-CoA is replaced in the control reactions with 20 μL of distilled water. The final concentrations of AcCoA and PABA for the single point activity measurements are 0.5 mM and 0.1 mM, respectively. 50 μL of 20% trichloroacetic acid is used to stop the PABA reactions and 100 μL of acetonitrile is used to stop the AF reactions after the reaction mixtures, either with or without specific concentrations of carmustine and lomustine, are incubated for 10 minutes at 37°C. Every reaction, including controls and experiments, is carried out in triplicate[1].

Rats: Rats are randomly assigned to four groups after being weighted individually before beginning the study and having their weights recorded. There are twelve rats in Group I (the saline group). The study includes the rats 48 hours after they receive an intraperitoneal (IP) injection of 2 mL/kg of saline 48 hours prior to the study. Fifteen rats make up Group II (corn oil group). The rats receive a 2 mL/kg injection of corn oil (vehicle) IP 48 hours prior to the investigation. Sixteen rats make up Group III (Carmustine group). For three days, the same hour of the day, a single-dose of 1 mL of saline IP is injected into these rats. The rats are added to the study 48 hours after the first dose of saline is administered, and twelve hours later, they receive injections of corn oil (2 mL/kg) and carmustine (25 mg/kg IP). There are twelve rats in Group IV (the trimetazidine group). For three days, these rats receive a single-dose injection of 2.5 mg/kg of trimetazidine (TMZ) IP at the same hour every day. Corn oil (2 mL/kg) and carmustine (25 mg/kg IP) are injected 12 hours after the first dose of TMZ, and the rats are added to the study 48 hours later[2].

Absorption, Distribution and Excretion
5 to 28% bioavailability
Approximately 60% to 70% of a total dose is excreted in the urine in 96 hours and about 10% as respiratory CO2.
Following IV infusion of carmustine, the steady-state volume of distribution averaged 3.25 L/kg. Because of their high lipid solubility, carmustine and/or its metabolites readily cross the blood-brain barrier. Substantial CSF concentrations occur almost immediately after IV administration of carmustine, and CSF concentrations of radioactivity have been variously reported to range from 15-70% of concurrent plasma concentrations. Carmustine metabolites are distributed into milk, but in concentrations less than those in maternal plasma.
The absorption of the copolymer contained in carmustine wafers has not been evaluated in humans. Plasma concentrations of carmustine following intracranial implantation of the wafers have not been determined in humans, but in rabbits undergoing surgical implantation of wafers containing 3.85% carmustine, no detectable levels of carmustine were observed in plasma.
When the carmustine wafer is exposed to the aqueous environment of the resection cavity, hydrolysis of the anhydride bonds in the copolymer occurs, resulting in the release of carmustine and two monomers, carboxyphenoxypropane, and sebacic acid. The carmustine contained in the wafer diffuses into the surrounding brain tissue. The metabolism and excretion of the copolymer contained in carmustine wafers has not been evaluated in humans. Animal studies have shown that more than 70% of the copolymer degrades within 3 weeks following implantation of carmustine wafers into brain tissue; following hydrolysis of the copolymer, carboxyphenoxypropane is eliminated renally, while sebacic acid (an endogenous fatty acid) is metabolized in the liver and expired as carbon dioxide. In humans, wafer remnants have been observed on brain imaging scans or located during subsequent surgical procedures up to 8 months following intracranial implantation. Wafer remnants retrieved from 2 patients approximately 2-3 months after implantation were analyzed and found to consist mostly of water and monomeric components with minimal detectable amounts of carmustine.
The disappearance of 1,3-bis(2-chlorethyl)-1-nitrosourea (BCNU) from plasma, liver, kidney, lung, brain, spleen, tumor tissue and epididymal adipose tissue of Walker 256/B carcinoma-bearing rats and healthy animals was measured by differential pulse polarography after i.v. bolus of the drug. Only BCNU, not its decomposition products, was detected by the polarographic assay. Levels of BCNU in liver of tumor-bearing animals were significantly lower (about 10 times) than those on healthy rats. A bi-exponential fit was used to calculate the kinetics of BCNU in plasma, kidney, lung and brain, but no difference could be found between healthy and Walker tumor-bearing rats. BCNU disappeared faster from adipose tissue of tumor-bearing animals than from normals.
Some 40 minutes after injection, BCNU is no longer an effective antitumour agent, and a few minutes after administration no unchanged BCNU can be detected in plasma. Following its ip or sc injection or oral administration, BCNU was rapidly distributed to most tissues, including brain and cerebrospinal fluid. Excretion was primarily in the urine; it was most rapid in mice (80% of the dose excreted in 24 hours) and less rapid in monkeys and dogs.

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Metabolism / Metabolites
Hepatic and rapid with active metabolites. Metabolites may persist in the plasma for several days.
The in vitro metabolism of the anticancer agent 1,3-bis(2-chloroethyl)-1-nitrosourea (BCNU) has been studied in male Fischer 344 rat liver microsomal preparations. The previously identified product, 1,3-bis(2-chloroethyl)urea (BCU), has been shown to be the major metabolite. Stable isotope labeling and mass spectral analysis of isolated metabolites indicate that BCU is formed exclusively from the metabolic denitrosation of BCNU. The rate of BCNU chemical decomposition in rat liver microsomal preparations deficient in NADPH and the metabolic disappearance rate in preparations containing added NADPH were measured and compared with the measured rate of metabolic formation of BCU under the same conditions. The rate of NADPH-dependent BCNU metabolism and BCU formation are equal within experimental error. BCNU was found to inhibit the rat liver 9000 g supernatant metabolism of 1-(2-chloroethyl)-3-cyclohexyl-1-nitrosourea (CCNU).
Hepatic and rapid with active metabolites. Metabolites may persist in the plasma for several days.
Route of Elimination: Approximately 60% to 70% of a total dose is excreted in the urine in 96 hours and about 10% as respiratory CO2.
Half Life: 15-30 minutes

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Biological Half-Life
15-30 minutes

Toxicity Summary
Carmustine causes cross-links in DNA and RNA, leading to the inhibition of DNA synthesis, RNA production and RNA translation (protein synthesis). Carmustine also binds to and modifies (carbamoylates) glutathione reductase. This leads to cell death.

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Hepatotoxicity
Mild and transient elevations in serum aminotransferase levels are found in up to 25% of patients treated with carmustine. Because carmustine is typically given in combination with other agents, its role in causing these serum enzyme elevations is not always clear. The abnormalities are generally transient, do not cause symptoms and do not require dose modification. Clinically apparent liver injury from carmustine has been limited to a small number of cases of cholestatic hepatitis and more frequent instances of sinusoidal obstruction syndrome, reported mostly with its use in high doses or as a conditioning agent in preparation for hematopoietic cell transplantation. The onset of sinusoidal obstruction syndrome is usually within two to three weeks of the myeloablation and is characterized by a sudden onset of abdominal pain, weight gain, ascites, marked increase in serum aminotransferase levels (and LDH), and subsequent jaundice and hepatic dysfunction. The severity of sinusoidal obstruction syndrome varies from a transient, self-limited injury to acute liver failure. The diagnosis of sinusoidal obstruction syndrome is usually based on clinical features of tenderness and enlargement of the liver, weight gain, ascites and jaundice occurring within 3 weeks of chemotherapy. Liver biopsy is diagnostic but often contraindicated, because of severe thrombocytopenia after hematopoietic cell transplantation.
Likelihood score: E* (unproven but suspected cause of clinically apparent liver injury, particularly when used for myeloablation).

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Effects During Pregnancy and Lactation
◉ Summary of Use during Lactation
No information is available on the use of carmustine during breastfeeding. Most sources consider breastfeeding to be contraindicated during maternal antineoplastic drug therapy, especially alkylating agents such as carmustine. The manufacturer recommends that breastfeeding be discontinued during carmustine therapy and for 1 month after the last dose.
◉ Effects in Breastfed Infants
Relevant published information was not found as of the revision date.
◉ Effects on Lactation and Breastmilk
Some evidence indicates that carmustine can increase serum prolactin. The prolactin level in a mother with established lactation may not affect her ability to breastfeed.

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Protein Binding
80%

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Toxicity Data
The oral LD₅₀s in rat and mouse are 20 mg/kg and 45 mg/kg, respectively.

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Interactions
In patients receiving carmustine and phenytoin, serum concentrations of phenytoin may be decreased. In patients receiving carmustine therapy, serum concentrations of phenytoin should be monitored carefully and dosage adjustments made as necessary.
Qualitative and quantitative changes in tear films leading to damage of the corneal and conjunctival epithelium have been reported in patients receiving high doses of carmustine and mitomycin.
Cimetidine may potentiate the myelosuppressive effects (e.g., neutropenia, agranulocytosis) of myelosuppressive drugs (e.g., alkylating agents, antimetabolites) or therapies (e.g., radiation). Concomitant cimetidine therapy has been reported to potentiate the neutropenic and thrombocytopenic effect of carmustine alone or combined with radiation therapy.

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Non-Human Toxicity Values
LD50 Rat oral 20 mg/kg
LD50 Rat ip 17,420 ug/kg
LD50 Rat sc 83,200 ug/kg
LD50 Rat iv 13,800 ug/kg
For more Non-Human Toxicity Values (Complete) data for Carmustine (9 total), please visit the HSDB record page.

[1]. Hung CF. Effects of carmustine and lomustine on arylamine N-acetyltransferase activity and 2-aminofluorene-DNA adducts in rat glial tumor cells. Neurochem Res. 2000 Jun;25(6):845-51.

[2]. The effect of trimetazidine on intrahepatic cholestasis caused by carmustine in rats. Hepatol Res. 2001 May 1;20(1):133-143.

Therapeutic Uses
BiCNU is indicated as palliative therapy as a single agent or in established combination therapy with other approved chemotherapeutic agents in the following: Brain tumors-glioblastoma, brainstem glioma, medulloblastoma, astrocytoma, ependymoma, and metastatic brain tumors. Multiple myeloma-in combination with prednisone. Hodgkin's Disease-as secondary therapy in combination with other approved drugs in patients who relapse while being treated with primary therapy, or who fail to respond to primary therapy. Non-Hodgkin's lymphomas-as secondary therapy in combination with other approved drugs for patients who relapse while being treated with primary therapy, or who fail to respond to primary therapy.
bis(Chloroethyl) nitrosourea has been used since 1971 as an antineoplastic agent in the treatment of Hodgkin's lymphoma, multiple myeloma, and primary or metastatic brain tumors.
Reported to have antiviral, antibacterial, and antifungal activity, but no evidence was found that it is currently used for these purposes. Former use.
MEDICATION (VET): A chemotherapeutic protocol using carmustine in combination with vincristine and prednisone was tested in dogs with multicentric malignant lymphosarcoma. Of seven dogs treated, six (85.7%) achieved complete remission. A partial response occurred in one dog. Median survival time was 224 days (mean 386 days), and median duration of remission was 183 days (mean 323 days). Marked neutropenia was observed following carmustine administration. There were no significant alterations in platelets and red blood cell counts during treatment, and no abnormalities attributable to the chemotherapy were found in serum biochemical profiles. Results of this study showed that carmustine is an effective alternative option in the treatment of canine lymphosarcoma.

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Drug Warnings
/BOXED WARNING/ WARNING: BiCNU (carmustine for injection) should be administered under the supervision of a qualified physician experienced in the use of cancer chemotherapeutic agents. Bone marrow suppression, notably thrombocytopenia and leukopenia, which may contribute to bleeding and overwhelming infections in an already compromised patient, is the most common and severe of the toxic effects of BiCNU. Since the major toxicity is delayed bone marrow suppression, blood counts should be monitored weekly for at least 6 weeks after a dose. At the recommended dosage, courses of BiCNU should not be given more frequently than every 6 weeks. The bone marrow toxicity of BiCNU is cumulative and therefore dosage adjustment must be considered on the basis of nadir blood counts from prior dose. Pulmonary toxicity from BiCNU appears to be dose related. Patients receiving greater than 1400 mg/sq m cumulative dose are at significantly higher risk than those receiving less. Delayed pulmonary toxicity can occur years after treatment, and can result in death, particularly in patients treated in childhood.
Human systemic effects by parenteral, intravenous, and possibly other routes: nausea or vomiting, reduced white blood cell and blood platelet counts, bone marrow damage, and potentially fatal respiratory system effects, including lung fibrosis, dyspnea, and cyanosis.
In a study of 17 children (aged 1-16 years) receiving carmustine in cumulative doses ranging from 770-1800 mg/sq m combined with cranial radiation therapy for intracranial tumors, 8 children (47%) died of delayed pulmonary fibrosis, including all of those who received initial treatment at less than 5 years of age (5 children). Onset of pulmonary fibrosis has been observed up to 17 years following carmustine therapy. Clinical findings include pulmonary hypoplasia with upper zone contraction on chest radiographs, and an unusual pattern of upper zone fibrosis on thoracic CT scans; no abnormal findings were observed on gallium scans.105 Late onset of reduction in pulmonary function was observed in all long-term survivors in the study. Carmustine-induced pulmonary fibrosis may be slowly progressive and cause death.
Pulmonary toxicity, including acute or delayed onset of pulmonary fibrosis causing death, has occurred in patients receiving systemic carmustine therapy. Pulmonary toxicity characterized by pulmonary infiltrates and/or fibrosis occurring 9 days to 43 months following treatment has been reported in patients receiving carmustine or related nitrosoureas. Most reported cases of pulmonary toxicity have occurred in patients receiving prolonged carmustine therapy with total doses exceeding 1400 mg/sq m; however, pulmonary fibrosis has occurred with lower total doses. Other risk factors include prior history of pulmonary disease and duration of carmustine therapy. Pulmonary toxicity occasionally has been rapidly progressive and/or fatal.
For more Drug Warnings (Complete) data for Carmustine (41 total), please visit the HSDB record page.

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Pharmacodynamics
Carmustine is one of the nitrosoureas indicated as palliative therapy as a single agent or in established combination therapy with other approved chemotherapeutic agents in treatment of brain tumors, multiple myeloma, Hodgkin's disease, and non-Hodgkin's lymphomas. Although it is generally agreed that carmustine alkylates DNA and RNA, it is not cross resistant with other alkylators. As with other nitrosoureas, it may also inhibit several key enzymatic processes by carbamoylation of amino acids in proteins.

C5H9CL2N3O2

214.0499

213.007

C, 28.06; H, 4.24; Cl, 33.12; N, 19.63; O, 14.95

154-93-8

Carmustine-d8

2578

Light yellow solid (low temperature); soild if <30°C; liquid if >30°C

1.5±0.1 g/cm3

404ºC

30 °C(lit.)

1.549

1.3

1

3

4

12

156

0

ClC([H])([H])C([H])([H])N(C(N([H])C([H])([H])C([H])([H])Cl)=O)N=O

DLGOEMSEDOSKAD-UHFFFAOYSA-N

InChI=1S/C5H9Cl2N3O2/c6-1-3-8-5(11)10(9-12)4-2-7/h1-4H2,(H,8,11)

1,3-bis(2-chloroethyl)-1-nitrosourea

NSC409962; NCI-C04773; NCIC04773; NCI C04773; Nitrumon; NSC 409962; NSC-409962; SK 27702; SRI 1720; DTI 015;; FDA 0345; BCNU Becenum; Bi CNU; BiCNU; 154-93-8; 1,3-Bis(2-chloroethyl)-1-nitrosourea; BCNU; Carmustin; Carmubris; Gliadel; Carmustine

29241900

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)

DMSO: ≥ 35 mg/mL (~163.5 mM)
H2O: ~100 mg/mL (~467.2 mM)

Solubility in Formulation 1: ≥ 2.08 mg/mL (9.72 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 20.8 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: ≥ 2.08 mg/mL (9.72 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 20.8 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: ≥ 2.08 mg/mL (9.72 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 20.8 mg/mL clear DMSO stock solution to 900 μL of corn oil and mix evenly.

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Solubility in Formulation 4: 5%DMSO+ 40%PEG300+ 5%Tween 80+ 50%ddH2O: 2.0mg/ml (9.34mM)

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Solubility in Formulation 5: 100 mg/mL (467.18 mM) in PBS (add these co-solvents sequentially from left to right, and one by one), clear solution; with ultrasonication.

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