Mebendazole

Synthetic Routes for Mebendazole Production

Condensation and Cyclization via 3,4-Diaminobenzophenone

The most widely documented method for synthesizing this compound begins with 3,4-diaminobenzophenone as a key intermediate. This compound reacts with methyl cyanocarbamate in a cyclization reaction facilitated by acidic conditions.

Procedure :

• Reaction Setup : 3,4-Diaminobenzophenone is dissolved in acetone and treated with hydrochloric acid to protonate the amine groups.
• Cyclization : Methyl cyanocarbamate is introduced, initiating a nucleophilic attack by the primary amine on the carbonyl group of the carbamate. This forms the benzimidazole core structure.
• Salt Formation : The crude product is treated with nitric acid in methanol to yield this compound nitrate, which enhances crystallinity.
• Crystallization : Conversion to the thermodynamically stable C-polymorph is achieved through controlled cooling and seeding.

Key Parameters :

• Temperature: 35–45°C during cyclization.
• Solvent System: Acetone for initial reaction; methanol for salt formation.
• Yield: 90% for crude product, 95% after crystallization.

Alternative Synthesis from o-Dichlorobenzene and Benzoyl Chloride

An alternative route starts with o-dichlorobenzene and benzoyl chloride, avoiding the need for pre-synthesized 3,4-diaminobenzophenone.

Procedure :

• Condensation : o-Dichlorobenzene reacts with benzoyl chloride in the presence of aluminum chloride to form 3,4-dichlorobenzophenone.
• Ammoniation : The dichloro intermediate undergoes ammonolysis with aqueous ammonia, replacing chlorine atoms with amine groups.
• Cyclization : The resulting 3,4-diaminobenzophenone is treated with methyl cyanocarbamate under reflux in formic acid to complete the benzimidazole ring.

Key Parameters :

• Catalyst: Aluminum chloride for Friedel-Crafts acylation.
• Reaction Time: 6–7 hours at 80°C during cyclization.
• Yield: 85–90% for the final product.

Optimization of Reaction Conditions

Solvent and Catalytic Systems

The choice of solvent significantly impacts reaction efficiency. Acetone and formic acid are preferred for their ability to stabilize intermediates and facilitate proton transfer. Catalytic amounts of hydrochloric acid accelerate cyclization by activating the carbonyl group of methyl cyanocarbamate.

Comparative Data :

Polymorph Control

This compound exists in multiple polymorphic forms, with the C-crystal being the most therapeutically stable. Seeding with C-polymorph crystals during cooling ensures uniform crystallization. Methanol and water mixtures (3:1 v/v) are optimal for inducing nucleation.

Purification and Crystallization Techniques

Salt Formation

Conversion to this compound nitrate improves solubility for subsequent purification. The nitrate salt is precipitated by adding methanol to the reaction mixture, achieving 98% purity after filtration.

Recrystallization

Crude this compound is dissolved in hot formic acid (65°C) and treated with activated carbon to adsorb impurities. Slow addition of water induces crystallization, yielding needle-like C-polymorph crystals.

Conditions :

• Solvent: Formic acid/water (4:1 ratio).
• Cooling Rate: 1°C per minute to prevent amorphous phase formation.

Analytical Methods for Quality Control

Spectrophotometric Analysis

A validated UV-Vis method quantifies this compound using oxidative coupling with N-bromosuccinimide (NBS) and Rhodamine B. The absorbance at 570 nm correlates linearly with concentration (2.5–30 μg/mL, R² = 0.9992).

Procedure :

• Oxidation : this compound is treated with excess NBS in 0.1 M HCl.
• Coupling : The oxidized product reacts with Rhodamine B, forming a chromophore.
• Detection : Absorbance measured at λ_max = 570 nm.

High-Performance Liquid Chromatography (HPLC)

Reverse-phase HPLC with C18 columns and acetonitrile/water mobile phases (70:30 v/v) resolves this compound from degradation products. Retention time: 6.2 minutes.

Comparative Analysis of Preparation Methods

Tipos de reacciones

El mebendazol experimenta diversas reacciones químicas, incluyendo reacciones de oxidación, reducción y sustitución.

Reactivos y condiciones comunes

Oxidación: Se pueden utilizar oxidantes comunes como el peróxido de hidrógeno o el permanganato de potasio.

Reducción: Se pueden emplear agentes reductores como el catalizador de paladio sobre carbón en presencia de hidrógeno.

Sustitución: Las reacciones de sustitución suelen implicar el uso de agentes halogenantes o nucleófilos.

Productos principales

Los productos principales que se forman a partir de estas reacciones dependen de los reactivos y las condiciones específicas que se utilicen. Por ejemplo, la reducción de la 4-amino-3-nitrobenzofenona con catalizador de paladio sobre carbón produce 3,4-diaminobenzofenona .

El mebendazol se ha reutilizado para diversas aplicaciones de investigación científica más allá de su uso original como antihelmíntico. Ha mostrado promesa en el tratamiento de cánceres cerebrales, incluyendo el glioma, al inhibir la progresión maligna y aumentar la sensibilidad de las células gliomatosas a la quimioterapia o radioterapia convencionales . Además, se ha explorado el mebendazol por sus propiedades anticancerígenas en múltiples cánceres, incluyendo la leucemia mieloide aguda, el cáncer de mama y el cáncer gastrointestinal .

El mebendazol funciona inhibiendo la polimerización de la tubulina en los parásitos, interrumpiendo la formación de microtúbulos e interfiriendo con la absorción de glucosa . Esto conduce en última instancia a la muerte del parásito. El compuesto se dirige a vías críticas implicadas en la proliferación celular, la apoptosis y la invasión/migración .

Structural Similarities and Differences

Mebendazole shares a benzimidazole core structure with other compounds in its class, including albendazole , flubendazole , oxibendazole , and parbendazole . Structural variations occur at the substituent groups:

• Flubendazole : A fluoroanalog of this compound, differing by a fluorine substitution at the para position of the benzene ring .
• Albendazole : Features a propylthio group at the carbamate side chain, enhancing lipophilicity and bioavailability .
• Oxibendazole : Contains an oxy group in place of the methyl group in this compound, altering solubility and pharmacokinetics .

Mechanism of Action and Tubulin Binding Affinity

All benzimidazoles target β-tubulin, but binding affinities vary across isoforms and species. Studies comparing this compound, nocodazole, and albendazole reveal:

ND = Not Determined

This compound and nocodazole exhibit comparable binding to both blood cell-specific (BTb) and Caenorhabditis elegans (CeTb) tubulin isoforms, while albendazole shows negligible affinity for BTb .

Anticancer Activity

This compound outperforms several analogs in preclinical cancer models:

Table 3: Anticancer Efficacy in Human Cell Lines

This compound induces mitochondrial cytochrome c release and apoptosis in lung cancer cells, while flubendazole shows comparable efficacy in neuroblastoma models .

Antiviral and Immunomodulatory Effects

• COVID-19 : Reduces C-reactive protein (CRP) by 40% and increases cycle threshold (CT) values in 3 days, indicating viral load reduction .
• Comparative Data : In silico studies identify this compound, oxibendazole, and albendazole as SARS-CoV-2 inhibitors, but only this compound has Phase II clinical trial support .

Pharmacokinetics and Solubility

This compound’s solubility improves with hydrotropes like sodium salicylate (up to 3 mol/L), but its bioavailability remains lower than albendazole’s .

Clinical Trial Data

• Oncology: this compound reduces tumor xenograft growth by 70% in mice and enhances radiation sensitivity in meningioma .
• COVID-19 : Phase II trials show faster inflammation resolution vs. placebo (p < 0.05) .
• Parasitic Diseases: Albendazole demonstrates superior cure rates (90% vs. 75%) in trichinosis compared to this compound .

Mebendazole is a broad-spectrum anthelmintic drug primarily used to treat parasitic infections. However, recent studies have unveiled its potential in oncology, particularly its biological activity against various cancer types. This article delves into the biological mechanisms, pharmacokinetics, and clinical applications of this compound, supported by data tables and case studies.

Antiparasitic Activity
this compound works by inhibiting the polymerization of tubulin, which is essential for microtubule formation in parasites. This disruption leads to impaired glucose uptake and energy depletion in the parasites, ultimately resulting in their death.

Antitumor Activity
Recent research highlights this compound's potential as an anticancer agent. It has been shown to induce apoptosis in cancer cells through several mechanisms:

• Bcl-2 Inactivation : this compound induces apoptosis in melanoma cells by phosphorylating Bcl-2, which prevents its interaction with the pro-apoptotic protein Bax, thus promoting cell death .
• Cell Cycle Arrest : It has been observed to cause cell cycle arrest in various cancer cell lines, including ovarian and colorectal cancers .
• Inhibition of Tumor Angiogenesis : this compound reduces angiogenesis by inhibiting VEGFR2 kinase activity, leading to decreased microvessel density in tumors .

Table 1: Biological Activities of this compound

Pharmacokinetics

This compound exhibits variable pharmacokinetic properties influenced by dosage and formulation. Studies indicate that plasma levels increase with higher doses, and the drug achieves a maximum concentration within hours post-administration.

Table 2: Pharmacokinetic Parameters of this compound

Case Study: this compound in Glioblastoma Treatment

A phase II clinical trial investigated the combination of this compound with temozolomide in patients with newly diagnosed high-grade glioma. The study enrolled 24 patients, revealing promising results regarding safety and overall survival:

• Median Overall Survival (OS) : 21 months.
• Progression-Free Survival (PFS) : 13.1 months for patients receiving more than one month of treatment .

Table 3: Clinical Outcomes from Glioblastoma Study

Case Study: this compound for COVID-19

A recent randomized controlled trial indicated that this compound therapy improved innate immunity and reduced inflammation markers in COVID-19 outpatients compared to a placebo group. Significant reductions in C-reactive protein (CRP) levels were noted within three days of treatment .

Basic Research Question: What experimental design considerations are critical for evaluating mebendazole’s pharmacokinetics in heterogeneous populations?

Methodological Answer:
To assess pharmacokinetics (PK) in diverse populations, researchers should:

• Define subpopulations (e.g., neonates, immunocompromised individuals) based on metabolic or physiological differences .
• Use population PK modeling to account for variability in drug absorption, distribution, and clearance. For example, sparse sampling in pediatric cohorts can reduce ethical and logistical challenges .
• Validate assays (e.g., HPLC, differential pulse polarography) to ensure sensitivity in detecting low plasma concentrations .
• Data sharing protocols must comply with ethical standards, including anonymization and controlled access to sensitive datasets .

Advanced Research Question: How can conflicting efficacy data for this compound in repurposed oncology studies be reconciled?

Methodological Answer:
Conflicting results often arise from:

• Variability in experimental models : Compare outcomes across cell lines (e.g., LNCaP vs. DU145 prostate cancer cells) and in vivo models (e.g., xenografts vs. genetically engineered mice) .
• Dose-response discordance : Use dose-ranging studies to identify therapeutic thresholds. For example, this compound’s anti-cancer effects in PDE4D7-knockdown LNCaPs occur at lower doses than in wild-type cells .
• Mechanistic heterogeneity : Conduct transcriptomic or proteomic analyses to map pathways (e.g., cAMP dynamics, microtubule disruption) influenced by tumor microenvironment factors .
• Meta-analysis frameworks : Apply PRISMA guidelines to aggregate preclinical data and identify bias sources (e.g., publication bias, model selection) .

Basic Research Question: What validated analytical methods are recommended for quantifying this compound in pharmaceutical formulations?

Methodological Answer:

• Electrochemical techniques : Differential pulse polarography (DPP) offers sensitivity at µg/mL levels, validated in pH 7.4 buffers to mimic physiological conditions .
• Chromatography : HPLC with UV detection (λ = 254 nm) provides reproducibility, but requires column optimization to separate this compound from excipients .
• Quality control : Cross-validate results with mass spectrometry (LC-MS) to confirm specificity, especially in complex matrices like serum .

Advanced Research Question: What strategies address this compound’s solubility limitations in preclinical testing?

Methodological Answer:

• Co-solvent systems : Test biocompatible solvents (e.g., PEG-400) to enhance aqueous solubility while monitoring cytotoxicity in vitro .
• Nanoformulation : Develop liposomal or polymeric nanoparticles to improve bioavailability. Characterize particle size (DLS) and encapsulation efficiency (UV-Vis) .
• In silico modeling : Use tools like COSMO-RS to predict solubility in simulated biological fluids and guide formulation design .

Basic Research Question: How should researchers design studies to evaluate this compound resistance in helminthic parasites?

Methodological Answer:

• Longitudinal sampling : Collect parasite isolates pre- and post-treatment to track β-tubulin mutations linked to resistance .
• Phenotypic assays : Measure IC50 shifts in larval motility or egg hatching inhibition assays across multiple generations .
• Genomic sequencing : Identify SNPs in β-tubulin isotype-1 genes and correlate with clinical failure rates .

Advanced Research Question: What methodologies optimize this compound combination therapies for synergistic anti-helminthic effects?

Methodological Answer:

• Checkerboard assays : Determine fractional inhibitory concentration indices (FICI) for this compound paired with albendazole or ivermectin .
• Mechanistic synergy : Use RNAi or CRISPR to validate target pathways (e.g., dual β-tubulin and glutamate-gated chloride channel disruption) .
• In vivo validation : Employ factorial design experiments in rodent models to assess efficacy-toxicity trade-offs .

Basic Research Question: How to ensure reproducibility in this compound’s in vitro cytotoxicity assays?

Methodological Answer:

• Standardize cell lines : Use authenticated stocks (e.g., ATCC-certified DU145) and control for passage number .
• Culture conditions : Maintain consistent O2 levels (5% CO2) and serum concentrations (10% FBS) to minimize batch effects .
• Endpoint validation : Combine MTT assays with live-cell imaging to confirm apoptosis vs. necrosis .

Advanced Research Question: How can computational models predict this compound’s off-target effects in repurposing studies?

Methodological Answer:

• Docking simulations : Use AutoDock Vina to screen this compound against human kinases or GPCRs implicated in side effects .
• Network pharmacology : Construct protein-protein interaction networks to identify secondary targets (e.g., PDE4D7 in prostate cancer) .
• Toxicogenomics : Apply LINCS database queries to predict gene expression changes in non-target tissues .