| Targets |
Procarbazine is a hydrazine derivative with effective antineoplastic activity, used in treating Hodgkin's disease, lung tumor, brain tumor, malignant melanoma and skin tumor. However, it also induces cancer in some experimental animals and humans, and IARC has classified it as probably carcinogenic to humans (Group 2A). The drug undergoes enzymatic oxidation to form carbon-centered free radicals and azoxy derivatives, leading to alkylation. Non-enzymatically, in the presence of Cu(II), procarbazine induces oxidative DNA damage via generation of reactive oxygen species and methyl radicals, contributing to its antitumor effects as well as mutagenesis and carcinogenesis [1].
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| ln Vitro |
Procarbazine plus Cu(II) induces lesions at the 5'-ACG-3' sequence, complementary to a p53 gene hotspot, and the 5'-TG-3' sequence that are formamidopyrimidine-DNA glycosylase-sensitive and piperidine-labile. Procarbazine damages DNA by generating methyl radicals and the Cu(I)-hydroperoxo complex non-enzymatically. [1] Procarbazine is mutagenic in a number of organs following high dose treatment and has a potent clastogenic effect in hematopoietic cells.[2]
In the presence of Cu(II), Procarbazine HCl (NSC-77213) (CAS#: 366-70-1) induced DNA damage in a dose- and time-dependent manner, as shown by piperidine-labile and Fpg-sensitive lesions. The damage occurred preferentially at thymine, cytosine and guanine residues, specifically at the 5'-ACG-3' sequence (complementary to codon 273, a known hotspot of the p53 gene) and the 5'-TG-3' sequence. Catalase partially inhibited DNA damage, while SOD enhanced it. Free hydroxyl radical scavengers (ethanol, mannitol, sodium formate, DMSO) showed no inhibitory effect. Bathocuproine (Cu(I)-specific chelator) inhibited damage. Procarbazine plus Cu(II) significantly increased formation of 8-oxo-7,8-dihydro-2'-deoxyguanosine (8-oxodG) in calf thymus DNA, which was completely inhibited by catalase. ESR spin-trapping experiments detected methyl radicals (carbon-centered radicals) generated from procarbazine and Cu(II) [1].
In Muta Mouse transgenic model, no increase in lacZ mutant frequency was observed in any organ after a single dose of 50 mg/kg, but repeated doses (150 mg/kg/day for 5 days) induced strong mutagenic responses in lung, bone marrow, and spleen (target organs for carcinogenesis), lower responses in kidney, marginal increase in liver, no increase in brain, and a >2-fold increase in testis [2].
In golden hamsters, procarbazine treatment (total 450 mg/kg over 4 weeks) caused a significant decrease in testicular and epididymal weight, a drastic reduction in haploid cells, spermatogenic arrest, and inhibition of spermatid differentiation and chromatin condensation as assessed by flow cytometry (propidium iodide and acridine orange staining). The effect was milder than in rats, with germinal stem cells not completely destroyed. Individual variation in response was observed [3]. |
| ln Vivo |
Procarbazine significantly reduces the weight of the testicles and epididyma, as well as haploid cells and spermatogenic arrest. These effects show variation in the test golden hamster population.[3] In brown adipose tissue, procarbazine causes a dose-dependent potentiation of MAO A. The elevation is more noticeable after monomethylhydrazine, with activity in rat homogenates reaching 350% of that in control homogenates. This amine's metabolism is decreased by procarbazine or monomethylhydrazine to an extent that is comparable to what was found ex vivo in blood vessel homogenates.[4] Procarbazine is a broad-spectrum carcinogen in rodents and monkeys, resulting in tumors of the nervous system, lung, mammary gland, and hematopoietic system. It is also mutagenic, clastogenic, and teratogenic in a variety of test systems of differing complexity. Under complex metabolic conditions, procarbazine in vivo produces a variety of chemically reactive species, such as free radicals and methylating agents.[5]
In vivo: In Muta Mouse, a single intraperitoneal dose of 50 mg/kg Procarbazine HCl (NSC-77213) (CAS#: 366-70-1) did not increase lacZ mutant frequency in bone marrow, liver, testis, lung, kidney, spleen or brain at 28 days post-treatment, but induced micronuclei in peripheral blood reticulocytes (mean 1.51% at 34h and 3.45% at 48h). Five daily doses of 150 mg/kg (total 750 mg/kg) resulted in highly positive lacZ mutant frequencies in lung, bone marrow, and spleen at 14 and 28 days after the last treatment; lower positive responses in kidney; slight increase in liver (92% above control at 14 days but returned to control at 28 days); no increase in brain; testis mutant frequency increased 122% over control at 28 days. The induced mutant frequencies were over 3×10⁻⁴ in bone marrow, spleen, and lung, correlating with known target organs for procarbazine carcinogenesis [2].
In golden hamsters, administration of procarbazine (first dose 150 mg/kg, followed by three weekly doses of 100 mg/kg, total 450 mg/kg) caused significant reduction in testis weight (from 1.824±0.2475 g in controls to 0.245±0.180 g in treated) and epididymis weight (from 476±51.5 mg to 89.8±52.4 mg). Flow cytometric analysis revealed that haploid cells became the minor population, diploid cells predominated, and the haploid/diploid ratio decreased from 4.71±1.27 (control) to 0.26±0.38 (treated). The tetraploid/diploid ratio also decreased from 0.89±0.25 to 0.52±0.17. Acridine orange staining showed spermatid maturation arrest, with most treated animals lacking advanced spermatids beyond the round spermatid stage (regions R1-R2). Histological sections confirmed spermatogenic arrest. The effect was milder than in rats, as seminiferous tubules still contained germinal cells and spermatogonia stem cells appeared unaffected [3].
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| Enzyme Assay |
In vitro enzyme assays: In the Fpg (formamidopyrimidine-DNA glycosylase) sensitivity assay, DNA fragments treated with Procarbazine HCl (NSC-77213) (CAS#: 366-70-1) and Cu(II) were incubated with two units of Fpg protein in reaction buffer (10 mM HEPES-KOH pH 7.4, 100 mM KCl, 10 mM EDTA, 0.1 mg/ml BSA) for 2 hours at 37°C, followed by electrophoresis. Fpg treatment revealed lesions at guanine residues, particularly at the 5'-ACG-3' sequence, indicating oxidative base damage including 8-oxodG [1].
In ESR spin-trapping experiments, reaction mixtures containing 500 µM procarbazine, 100 µM CuCl₂, 5 µM DTPA, and 200 mM spin trapping agent (POBN in 10 mM phosphate buffer or MNP in 50 mM MOPS buffer) were incubated for 60 min at 37°C. ESR spectra were recorded at 25°C using a spectrometer with 100 kHz field modulation, 16 mW microwave power, and 0.1 mT modulation amplitude. POBN trapped carbon-centered radicals showing a six-line signal (a^N = 1.61 mT, a^H = 0.27 mT). MNP trapped methyl radicals showing a 12-line signal consisting of three 1:3:3:1 quartet spectra (a^N = 1.70 mT, a_β^H(3H) = 1.39 mT), assigned to the MNP/•CH₃ radical adduct [1].
Catalase (150 units/ml) and SOD (150 units/ml) were used to study their effects on DNA damage; catalase partially inhibited damage while SOD enhanced it. Bathocuproine (50 µM), a Cu(I)-specific chelator, also inhibited DNA damage [1].
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| Cell Assay |
Cell Line: L1210 cells
Concentration: 5 and 20 nM
Incubation Time: 1 hour
Result: Showed 99.3% and 99.9% survival of cells at 5 mM and 20 mM, respectively.
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| Animal Protocol |
Male muta mouse (7–8 weeks old)
50 and 150 mg/kg
Intraperitoneal injection; 50 and 150 mg/kg; once daily; 5 days
Animal protocols: For the Muta Mouse study, male Muta Mouse (7-8 weeks old) were injected intraperitoneally with Procarbazine HCl (NSC-77213) (CAS#: 366-70-1) dissolved in saline at 10 ml/kg. Single treatment: 50 mg/kg, with blood collected at 34h and 48h for micronucleus assay; mice were killed at 7, 14, and 28 days post-treatment, and liver, bone marrow, testis, lung, kidney, spleen, and brain were collected for lacZ mutant frequency analysis. Repeated treatment: 150 mg/kg once daily for 5 days (total 750 mg/kg), with mice killed at 14 and 28 days after the last treatment. Control mice received saline only [2].
For the golden hamster study, male golden hamsters (Mesocricetus auratus, 53 days old, 85-95 g) were injected intraperitoneally with procarbazine once a week for 4 weeks. The first dose was 150 mg/kg, followed by three weekly doses of 100 mg/kg (total 450 mg/kg). One week after the last treatment, 4 hamsters were sacrificed; the remaining 3 were sacrificed after an additional 4 weeks. Testes and epididymides were excised, weighed, and processed. A portion of each testis was fixed in Bouin's solution, dehydrated, embedded in paraffin, sectioned, and stained with hematoxylin/eosin for histological evaluation. Another portion was minced to prepare single-cell suspensions in TNE buffer (0.01 M Tris, 0.15 M NaCl, 0.001 M EDTA, pH 7.4), filtered through 100-μm nylon mesh, supplemented with 10% glycerol, and frozen at -20°C until flow cytometric analysis [3].
For flow cytometry, thawed cell suspensions were centrifuged at 500g for 10 min, resuspended in TNE buffer, and stained with propidium iodide (12 μg/ml final) for ploidy analysis, or with acridine orange (6 μg/ml in citric acid buffer with Triton X-100 and HCl) for haploid cell analysis. Stained samples were analyzed on a flow cytometer with 488 nm argon laser excitation, recording red fluorescence (BP650 LP) for PI, and both green (BP530/30) and red (BP650 LP) for acridine orange. Data from 10,000 cells per sample were processed using WINMDI software [3].
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| ADME/Pharmacokinetics |
Absorption, Distribution and Excretion
Procarbazine is rapidly and completely absorbed. After oral administration, procarbazine hydrochloride is rapidly and almost completely absorbed from the gastrointestinal tract. Peak plasma drug concentration is reached within 1 hour after a single oral dose of 30 mg of radiolabeled procarbazine hydrochloride. Oral administration typically achieves similar plasma concentrations to intravenous administration. /Procarbazine Hydrochloride/ Approximately 45% to 70% of the dose is excreted in the urine as metabolites within 24 hours after administration. Of the oral or parenteral dose administered to humans, 25% to 70% is recovered in the urine within 24 hours after administration; less than 5% is excreted unchanged, with the remainder mostly excreted as the metabolite N-isopropylterephthalic acid. Distribution studies in animals and humans have shown that radioactive material is present in the liver, kidneys, intestinal wall, and skin after intravenous injection of radiolabeled procarbazine hydrochloride. The drug can cross the blood-brain barrier and distribute into the cerebrospinal fluid. Following oral administration, procarbazine rapidly reaches equilibrium between plasma and cerebrospinal fluid. It is currently unclear whether procarbazine is distributed into breast milk.
For more complete data on absorption, distribution, and excretion of procarbazine (6 items), please visit the HSDB record page.
Metabolism/Metabolites
Procarbazine is primarily metabolized in the liver and kidneys. The drug appears to undergo auto-oxidation, generating an azo derivative and releasing hydrogen peroxide. The azo derivative is isomerized to a hydrazone, which is hydrolyzed and cleaved into a benzaldehyde derivative and methylhydrazine. The methylhydrazine is further degraded to carbon dioxide and methane, possibly also generating hydrazine, while the aldehyde is oxidized to N-isopropylterephthalic acid, which is excreted in the urine.
It is rapidly metabolized in the human body… Oxidation of procarbazine produces the corresponding azo compound and hydrogen peroxide. Further metabolism (possibly in the liver) produces azo oxide derivatives, which circulate in the blood and possess potent cytotoxic activity.
Procarbazine may be converted into demethylprocarbazine in rats; N-isopropyl-α-methylazo-p-toluamide and isopropylamide terephthalate are also generated in rats. /Excerpt from Table/
Intraperitoneal injection of (14)C-monomethyl hydrazine in rats results in its bioconversion to (14)C-methane within 3 minutes. ...Rats produce 25% (14)C-CH4 and 3% (14)C-CO2 within 90 minutes. ...(14)C-procarbazine ... has a low bioconversion efficiency to CH4. After 90 minutes, 4% of the dose ... is excreted as (14)C-CH4. /Hydrochloride/
NADPH-dependent microsomal metabolism of terminally N-methyl-tagged [14C]procarbazine leads to covalent binding of the drug to exogenously added DNA; this reaction can be inhibited by metheprone. The metabolism of procarbazine also revealed that the methyl group of the drug covalently binds to microsomal proteins during metabolism, but this protein binding is at least an order of magnitude less than that of its major oxidative metabolite, N-isopropyl-α-(2-methylazo)-p-toluamide. The covalent binding of the N-methyl group of the azo derivative to microsomal proteins and its methane-producing reaction characteristics share many similarities in apparent kinetic parameters (Km and Vmax), induction and inhibition patterns, suggesting a common metabolic pathway, the generation of active intermediates, and the involvement of cytochrome P450. Reduced glutathione promotes methane formation and inhibits covalent binding to proteins. For more complete metabolic/metabolite data on procarbazine (6 metabolites), please visit the HSDB record page. Procarbazine is primarily metabolized in the liver and kidneys. The drug appears to undergo auto-oxidation, generating azo derivatives and releasing hydrogen peroxide. The azo derivatives are isomerized to hydrazones, which hydrolyze to benzaldehyde derivatives and methylhydrazine. Methylhydrazine further degrades into carbon dioxide and methane, possibly also producing hydrazine, while the aldehyde is oxidized to N-isopropylterephthalic acid, which is excreted in the urine.
Half-life: 10 minutes
Biological half-life
10 minutes
The biological half-life of procarbazine hydrochloride in plasma and cerebrospinal fluid is approximately 1 hour. /Procarbazine hydrochloride/
…The half-life in blood after intravenous injection is approximately 7 minutes.
The plasma half-life of the parent drug is approximately 10 minutes.
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| Toxicity/Toxicokinetics |
Toxicity Summary
The exact mechanism of cytotoxicity of procarbazine is not fully understood. Evidence suggests that the drug may act by inhibiting the synthesis of proteins, RNA, and DNA. Studies have shown that procarbazine may inhibit the process of methionine methyl transfer to tRNA. Loss of functional tRNA may lead to the cessation of protein synthesis, which in turn leads to the cessation of DNA and RNA synthesis. Furthermore, procarbazine may directly damage DNA. Hydrogen peroxide produced during drug autoxidation may attack sulfhydryl groups in residual proteins that are tightly bound to DNA. Hepatotoxicity
Mild and transient elevations in serum transaminase levels are not uncommon during systemic combination chemotherapy, and the role of procarbazine in these abnormalities is generally unclear. More than half of patients experience elevated transaminase levels, with 10% to 20% experiencing levels exceeding five times the upper limit of normal. However, dose adjustments are rarely necessary due to elevated serum enzymes. While clinically significant liver disease with fever and significantly elevated serum transaminase levels has been reported, this is extremely rare. One case report described self-limiting hepatocellular injury without jaundice during a second cycle of combination therapy, which relapsed after re-administration of procarbazine but did not relapse with other antitumor drugs. Probability score: D (likely a rare cause of clinically significant liver injury). Toxicity data: LD50 = 785 mg/kg (oral in rats). Interactions: Concomitant use of procarbazine with alcohol may cause a disulfiram-like reaction and exacerbate central nervous system depression and orthostatic hypotension; furthermore, alcoholic beverages (especially beer, wine, or malt spirits) may contain tyramine, which may lead to a hypertensive response. Concomitant use of local anesthetics (such as epinephrine or levonorepinephrine) or cocaine with procarbazine may cause severe hypertension due to sympathomimetic effects. Cocaine should not be used during or within 14 days after taking monoamine oxidase inhibitors.
The hypotensive effect may be enhanced when using spinal anesthetics and procarbazine concurrently; if spinal anesthesia is planned, it is recommended to discontinue procarbazine at least 10 days before elective surgery.
Concurrent use of anticholinergic drugs or other drugs with anticholinergic activity, antimotor dyskinesia drugs, or antihistamines with procarbazine may enhance the anticholinergic effect due to the secondary anticholinergic activity of monoamine oxidase inhibitors. Furthermore, monoamine oxidase inhibitors (MAO inhibitors) may block the detoxification effect of anticholinergic drugs, thereby enhancing their effect; patients should be advised to report gastrointestinal problems promptly, as concurrent use of MAO inhibitors may lead to paralytic ileus. Concurrent use with MAO inhibitors may also prolong and enhance the central nervous system depressant and anticholinergic effects of antihistamines; concurrent use is not recommended.
For more complete interaction data on procarbazine (22 in total), please visit the HSDB record page.
Non-human toxicity values
Rat intravenous injection LD50: 600 mg/kg body weight (day 59)
Toxicity/Toxicokinetics: In Muta Mouse, Procarbazine HCl (NSC-77213) (CAS#: 366-70-1) induced micronuclei in peripheral blood reticulocytes at a single dose of 50 mg/kg (mean 1.51% at 34h and 3.45% at 48h vs. control ~0.18%). Repeated dosing (150 mg/kg/day for 3 days) further increased micronuclei (mean 6.57% at 24h after third treatment), but depressed erythropoiesis, making it difficult to score 1000 reticulocytes (only 500 were analyzed). The reported LD₅₀ for procarbazine in mice is approximately 770 mg/kg (total dose of 750 mg/kg used in repeated dosing corresponds to near-LD₅₀) [2].
In golden hamsters, procarbazine treatment (total 450 mg/kg) caused significant testicular toxicity: testis weight decreased from 1.824±0.2475 g (control) to 0.245±0.180 g (treated); epididymis weight from 476±51.5 mg to 89.8±52.4 mg; body weight from 149±21 g to 113.6±16 g. Spermatogenesis was severely impaired, with haploid/diploid ratio dropping from 4.71±1.27 to 0.26±0.38. Sperm abnormalities were increased and sperm counts diminished. However, spermatogonia stem cells were not completely destroyed, as seminiferous tubules still contained germinal cells [3].
In vitro, procarbazine plus Cu(II) induced oxidative DNA damage and 8-oxodG formation, which was partially inhibited by catalase, suggesting involvement of H₂O₂. SOD enhanced DNA damage. The drug generates methyl radicals that can form 7-methylguanine and 8-methylguanine, leading to depurination and mutations [1].
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| References |
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| Additional Infomation |
Therapeutic Uses
Antitumor drug; carcinogen
Veterinary drug: Antitumor drug…/For/experimental treatment of various transplanted animal tumors. /Hydrochloride/
Procarbazine, in combination with other drugs, is indicated for the treatment of Hodgkin lymphoma (stages III and IV)…/Included in the US product label/
/Procarbazine, in combination with other drugs, is indicated for the treatment of/certain non-Hodgkin lymphomas. /Not included in the US product label/
For more complete therapeutic use data for procarbazine (6 types in total), please visit the HSDB record page.
Drug Warnings
Procarbazine is a highly toxic drug and should only be used under the continuous supervision of an experienced cancer chemotherapy clinician. If applicable, procarbazine treatment should be initiated during the patient's hospitalization; the patient's clinical and histological diagnosis, as well as hematological, renal, and hepatic conditions, should be carefully evaluated. Although studies on procarbazine in the elderly have not been conducted, given the increased risk of vascular events (especially in cases of sudden hypertension), increased sensitivity to antihypertensive effects, and decreased metabolic capacity in older patients, the use of monoamine oxidase inhibitors (MAOIs) for the first time is not recommended for patients over 60 years of age. When prescribing MAOIs to older patients, their history of depression, adherence to medication, and any potential drug interactions must also be considered. Furthermore, older patients are more prone to age-related renal impairment, which may necessitate dose reduction, or in severe cases, avoidance of procarbazine. Patients taking procarbazine should be advised to abstain from alcohol and avoid foods high in tyramine, such as yogurt, cheese, and bananas. Patients should also be advised to avoid over-the-counter medications containing antihistamines or sympathomimetic drugs and to discuss any prescription medications they are taking with their clinician treating their procarbazine treatment. Pregnancy Risk Grade: D/Positive evidence of risk exists. Human studies, trial data, and post-marketing data have confirmed fetal risk. However, the potential benefits of using this drug may outweigh the potential risks. For example, this drug may be suitable in life-threatening situations or when a patient has a serious illness and other safer medications are unavailable or ineffective. For more complete data on drug warnings for procarbazine (24 in total), please visit the HSDB records page.
Pharmacodynamics
Procarbazine is an alkylating agent used to treat a variety of cancers. Alkylating agents are named for their ability to add alkyl groups to numerous electronegative groups under specific intracellular conditions. They inhibit tumor growth by cross-linking guanine bases in the DNA double helix—directly attacking the DNA. This prevents the DNA strands from unwinding and separating. Since DNA replication requires unwinding and separation, cells cannot divide. Furthermore, these drugs can add methyl or other alkyl groups to molecules that shouldn't be present, inhibiting the proper use of base pairing and leading to DNA mismatches. Procarbazine is a cell-phase specific drug for the S phase of cell division.
Additional Info: Procarbazine HCl (NSC-77213) (CAS#: 366-70-1) is a methylhydrazine derivative used in cancer chemotherapy (Hodgkin's disease, lung cancer, brain tumors, melanoma). Its antineoplastic and carcinogenic effects involve both alkylation (via methyl diazonium ion forming O⁶-methylguanine) and oxidative DNA damage. The non-enzymatic mechanism in the presence of Cu(II) involves reduction of Cu(II) to Cu(I), formation of a Cu(I)-hydroperoxo complex that causes oxidative DNA damage, and generation of methyl radicals that alkylate guanine residues. Procarbazine induces double-base lesions at 5'-ACG-3' (complementary to p53 gene codon 273 hotspot) and 5'-TG-3' sequences. These clustered lesions are poorly repaired and may contribute to mutagenesis. Mutations observed in vivo include G:C→A:T transitions (from O⁶-methylguanine), G:C→T:A and A:T→T:A transversions (from 8-oxodG and depurination). IARC has classified procarbazine as Group 2A (probably carcinogenic to humans). In transgenic mouse assays, procarbazine shows strong clastogenicity (micronucleus induction) at low doses, while gene mutations (lacZ) require higher repeated dosing, and organ-specific mutagenicity correlates with carcinogenic target organs (lung, bone marrow, spleen). The drug also causes germ cell mutations (dominant lethal and specific locus mutations in mice, and increased testis lacZ MF) [1][2][3].
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COA