Kinetics and Mechanism of Hydrolysis of Efavirenz

Purpose. To determine the kinetics and mechanism of hydrolysis of efavirenz [(S)-6-chloro-4-(cyclopropylethynyl)-1,4-dihydro-4-(trifluoromethyl)-2H-3,1-benzoxazin-2-one] in aqueous solutions. Methods. The solution stability was examined at 60°C and an ionic strength of 0.3 M over the pH range of 0.6 to 12.8. The loss of efavirenz and the appearance of degradants were followed with a reverse-phase high-performance liquid chromatography assay. Characterization of the degradants was accomplished with liquid chromatography-mass spectrometry. Results. The degradation of efavirenz followed apparent first-order kinetics over the pH range of 0.6 to 12.8 at 60°C. The catalytic effect of phosphate and borate buffers was negligible while acetate and citrate demonstrated buffer catalysis. The overall rate constant indicated a pH minimum (the pH of maximum stability) of approximately 4. Mass spectra data identified a degradant with a molecular weight consistent with hydrolysis of the cyclic carbamate to the corresponding amino alcohol. The degradation route was confirmed with spiking experiments with an authentic sample of the amino alcohol indicating that the carbamate hydrolysis pathway was the predominant reaction throughout the pH range studied. Subsequent degradation of the amino alcohol proceeded at the extremes of the pH range studied via rearrangement to the quinoline. Conclusions. The pH-rate profile was consistent with a combination of a V-shaped profile in the pH range of 0-9 and a sigmoid-shaped profile in the pH range of 4-13. The plateau that began at pH 10-11 is a result of the ionization of the amine of the carbamate inhibiting the base-catalyzed hydrolysis of efavirenz, given that the ionized form of the carbamate is resonance-stabilized toward hydroxide-catalyzed degradation. Thus, increasing the pH resulted in a parallel decrease in the unionized fraction and increase in hydroxide ion concentration resulting in a constant k_obs value.     

Induction of Multiple Drug Transporters by Efavirenz

Efavirenz, an important component of human immunodeficiency virus 1 (HIV-1) therapy, causes substantial drug interactions as an inducer of cytochromes and the transporter ABCB1. So far its effect on the expression of other transporters is unknown. We therefore investigated the effect of long-term exposure of cells to efavirenz on expression of a large number of important drug transporters and on cell proliferation as a surrogate of intracellular availability. LS180 cells were used as a surrogate for the major site of drug interactions and Jurkat cells were used as a surrogate for the main target cells of HIV therapy. Cells were treated with efavirenz over 4 weeks and mRNA expression of drug transporters was repeatedly quantified. After 4 weeks, efavirenz significantly up-regulated the mRNA of ABCB1, ABCG2, ABCC2, ABCC3, ABCC5, and SLCO3A1 in LS180 cells and ABCG2, ABCC1, ABCC4, ABCC5, and SLCO2B1 in Jurkat cells. However these changes in transporter expression did not influence cell proliferation indicating that intracellular efavirenz concentrations were likely not altered. Efavirenz induces mRNA expression of several drug transporters critically modulating the kinetics of other drugs. While these expressional changes will most likely not influence the efficiency of efavirenz itself, they might change the effect of other co-administered drugs.     

依非韦仑在妊娠早期的安全性评价

目的 评价非核苷逆转录酶抑制剂依非韦仑（efavirenz,EFV）在妊娠早期抗逆转录病毒治疗（antiretroviral therapy,ART）中的安全性。方法 以代表性meta分析为依据,介绍EFV用于妊娠早期的文献资料及各国指南修订变化,对比各指南中妊娠期ART方案中EFV的推荐级别,阐述EFV的药动学和妊娠期用法。结果 EFV对HIV-1病毒有显著抑制作用,早期动物实验和少量人类临床数据显示EFV会导致胎儿神经管畸形,被列为妊娠早期禁用药物。随着样本量增加,数据显示妊娠早期使用EFV不增加整体出生缺陷风险,神经管缺陷风险概率不高于一般人群。但由于人类数据较为有限,仅部分指南推荐妊娠早期使用EFV。结论 针对不同个体的妊娠早期ART方案,仍需针对具体问题进行分析评估。     

The HIV Antiretroviral Drug Efavirenz has LSD-Like Properties

Anecdotal reports have surfaced concerning misuse of the HIV antiretroviral medication efavirenz ((4S)-6-chloro-4-(2-cyclopropylethynyl)-4-(trifluoromethyl)-2,4-dihydro-1H-3,1-benzoxazin-2-one) by HIV patients and non-infected teens who crush the pills and smoke the powder for its psychoactive effects. Molecular profiling of the receptor pharmacology of efavirenz pinpointed interactions with multiple established sites of action for other known drugs of abuse including catecholamine and indolamine transporters, and GABA_A and 5-HT_2A receptors. In rodents, interaction with the 5-HT_2A receptor, a primary site of action of lysergic acid diethylamine (LSD), appears to dominate efavirenz''s behavioral profile. Both LSD and efavirenz reduce ambulation in a novel open-field environment. Efavirenz occasions drug-lever responding in rats discriminating LSD from saline, and this effect is abolished by selective blockade of the 5-HT_2A receptor. Similar to LSD, efavirenz induces head-twitch responses in wild-type, but not in 5-HT_2A-knockout, mice. Despite having GABA_A-potentiating effects (like benzodiazepines and barbiturates), and interactions with dopamine transporter, serotonin transporter, and vesicular monoamine transporter 2 (like cocaine and methamphetamine), efavirenz fails to maintain responding in rats that self-administer cocaine, and it fails to produce a conditioned place preference. Although its molecular pharmacology is multifarious, efavirenz''s prevailing behavioral effect in rodents is consistent with LSD-like activity mediated via the 5-HT_2A receptor. This finding correlates, in part, with the subjective experiences in humans who abuse efavirenz and with specific dose-dependent adverse neuropsychiatric events, such as hallucinations and night terrors, reported by HIV patients taking it as a medication.     

HPLC法测定依非韦伦片中三个特定杂质的含量

目的:建立测定依非韦伦片中三种难分离的特定杂质含量的方法。方法:采用Waters Symmetry C₁₈柱(4.6 mm×250 mm,5μm),以水-乙腈-三氟乙酸为流动相梯度洗脱,检测波长为250 nm,流量为1.5 mL·min^-1。结果:三种特定杂质峰与主峰间的分离度良好,最低检测限为0.5 ng,依非韦伦浓度在0.125~2.5μg·mL^-1范围内与峰面积呈良好的线性关系(r=0.9999,n=6)。结论:本方法简便,专属性强,可用于依非韦伦片中三种特定杂质的控制。     

利福布汀与依非韦伦在大鼠体内药动学的相互作用研究

目的:研究利福布汀(RBT)与依非韦伦(EFV)在大鼠体内药动学的相互影响。方法:将大鼠随机分为EFV单用组(54mg·kg-1)、RBT单用组(40.5mg·kg-1)及其联用组,每组6只,灌胃给药后,液质联用法检测给药后0、0.25、0.5、1、2、3、4、6、9、12、24、48、72h的血药浓度,计算药动学参数。结果:单用时,EFV、RBT的药动学参数分别为:cmax:(4.36±1.23)、(5.62±2.33)mg·L-1,AUC0～72h:(27.37±8.10)、(117.00±50.45)mg·h·L-1,CLZ/F:(2.15±0.77)、(0.36±0.15)L·h-1·kg-1,VZ/F:(8.33±2.49)、(12.06±7.64)L·kg-1;联用组二者的药动学参数分别为cmax:(1.68±0.82)、(3.05±1.66)mg·L-1,AUC0～72h:(13.13±9.95)、(89.60±55.75)mg·h·L-1,CLZ/F:(5.12±2.17)、(0.54±0.24)L·h-1·kg-1,VZ/F:(23.53±11.43)、(13.06±4.75)L·kg-1。与单用时比较,联用组EFV的cmax、AUC0～72h显著减小,CLZ/F、VZ/F显著增加(P<0.05),而RBT药动学各参数未见显著变化。结论:RBT能加快大鼠体内EFV的消除,降低其生物利用度。     

Efavirenz concentrations in HIV-infected patients with and without viral hepatitis

AIMS

Data on efavirenz in HIV/viral hepatitis co-infected patients is non-consensual, probably due to liver function heterogeneity in the patients included.

METHODS

A case control study was performed on 27 HIV-infected patients, with controlled and homogenous markers of hepatic function, either mono-infected or co-infected with HBV/HCV, to ascertain the influence of viral hepatitis on efavirenz concentrations over a 2-year follow-up period.

RESULTS

No differences were found in efavirenz concentrations between groups both during and at the end of the follow-up period: control (2.43 ± 1.91 mg l^–1) vs. co-infected individuals (2.37 ± 0.37 mg l^–1).

CONCLUSION

It was concluded that HBV/HCV infections in themselves do not predispose to an overexposure to efavirenz.

WHAT IS ALREADY KNOWN ABOUT THIS SUBJECT

• HIV-1 co-infection with HBV/HCV is the most important factor determining efavirenz-induced liver toxicity. Higher efavirenz plasma concentrations have been reported in these patients facilitating concentration drug-related adverse effects.
• It is not known whether changes in efavirenz disposition are due to the hepatitis infection/inflammation or to liver failure. As a consequence, the guidelines for the application of therapeutic drug monitoring of efavirenz in HBV/HCV co-infected patients have not been established.

WHAT THIS STUDY ADDS

• The present study has shown that HBV/HCV infection in itself does not predispose to higher efavirenz plasma concentrations. In the absence of hepatic failure, the risk of efavirenz concentration-dependent toxicity is not increased.
• Thus, therapeutic drug monitoring indications in co-infected patients with hepatic function within the normal range should be the same as in HIV-1 mono-infected patients.

     

Instability of Efavirenz Metabolites Identified During Method Development and Validation

Accurate quantification of efavirenz metabolites in patient samples is required to investigate their potential contribution to efavirenz adverse events. This study aimed to validate a LC-MS/MS method to quantify and investigate the stability of efavirenz and metabolites in human plasma. Compounds were extracted from plasma by supported liquid extraction and resolved on a C18 column. Validation was performed following FDA bioanalytical method validation guidelines. Stability under common conditions of sample pre-treatment and storage were assessed. Efavirenz and 8-hydroxyefavirenz were stable for all conditions tested. 7-Hydroxyefavirenz and 8,14-dihydroxyefavirenz were not stable in plasma at room temperature for 24 h (46%-69% loss), -20°C for 90 days (17%-50% loss), or 60°C for 1 h (90%-95% loss). Efavirenz and 8-hydroxyefavirenz concentrations in HIV/AIDS patient (n=5) plasma prepared from pre-treated (60°C for 1 h) whole blood varied from 517-8564 ng/mL and 131-813 ng/mL, respectively. 7-Hydroxyefavirenz and 8,14-dihydroxyefavirenz concentrations were below validated lower limits of quantification (0.25 and 0.5 ng/mL, respectively), most likely due to sample pre-treatment. This is the first report of 7-hydroxyefavirenz and 8,14-dihydroxyefavirenz instability under conditions commonly used in preparation of samples from HIV/AIDS patients. Alternative biosafety measures to heat pre-treatment must therefore be used for accurate quantification of plasma 7-hydroxyefavirenz and 8,14-dihydroxyefavirenz.     

Biowaiver Monographs for Immediate Release Solid Oral Dosage Forms: Efavirenz

Literature data pertaining to the decision to allow a waiver of in vivo bioequivalence testing for the approval of immediate-release (IR) solid oral dosage forms containing efavirenz as the only active pharmaceutical ingredient (API) are reviewed. Because of lack of conclusive data about efavirenz's permeability and its failure to comply with the “high solubility” criteria according to the Biopharmaceutics Classification System (BCS), the API can be classified as BCS Class II/IV. In line with the solubility characteristics, the innovator product does not meet the dissolution criteria for a “rapidly dissolving product.” Furthermore, product variations containing commonly used excipients or in the manufacturing process have been reported to impact the rate and extent of efavirenz absorption. Despite its wide therapeutic index, subtherapeutic levels of efavirenz can lead to treatment failure and also facilitate the emergence of efavirenz-resistant mutants. For all these reasons, a biowaiver for IR solid oral dosage forms containing efavirenz as the sole API is not scientifically justified for reformulated or multisource drug products. © 2012 Wiley Periodicals, Inc. and the American Pharmacists Association J Pharm Sci 102:318–329, 2013     

唐草片对依非韦伦在大鼠体内药动学的影响

目的:研究唐草片对依非韦伦(EFV)在大鼠体内药动学的影响。方法:采用液相色谱-串联质谱(LC-MS/MS)法。色谱柱为Eclipse XDB-C18,流动相为0.1%甲酸水溶液-甲醇(20∶80,V/V),流速为0.5 ml/min,柱温为30℃,进样量为10μl;采用电喷雾电离(ESI),用于定量分析的离子对依次为m/z 515.2→276.2(内标:替米沙坦),m/z 316.0→243.9(EFV)。18只SD大鼠随机均分为3组,分别单独灌胃EFV(54 mg/kg,即EFV组)、同时灌胃EFV和唐草片(54 mg/kg+864 mg/kg,即EFV-0T组)、灌胃EFV 60 min后灌胃唐草片(54 mg/kg+864 mg/kg,即EFV-60T组)。于大鼠灌胃给药前与末次给药后0.25、0.5、1.0、2.0、3.0、4.0、6.0、9.0、12.0、24.0h于颈动脉取血,测定不同时间血药浓度,以DAS 2.0软件计算药动学参数。结果:EFV质量浓度在75.0~9 600.0 ng/ml范围内与其和内标物峰面积之比呈良好线性关系,精密度试验的RSD值均小于10%,准确度为92.1%~102.0%,提取回收率为88.6%~91.0%,稳定性试验的RSD均小于10%。与EFV组比较,EFV-0T组AUC0-∞、t1/2、MRT0-∞降低,CL/F提高,差异有统计学意义(P<0.01或P<0.05);EFV-60T组各指标差异无统计学意义(P>0.05)。各组大鼠tmax、cmax、V/F差异无统计学意义。结论:EFV与唐草片同时服用会降低EFV血药浓度;当两药合用时,须在服用EFV 60 min后服用唐草片。     

Formulation Development and Dissolution Rate Enhancement of Efavirenz by Solid Dispersion Systems

The aim of this study was to enhance the dissolution rate of efavirenz using solid dispersion systems (binary and ternary). A comparison between solvent and fusion method was also investigated. Solid dispersions of efavirenz were prepared using polyethylene glycol 8000, polyvinylpyrrolidone K30 alone and combination of both. Tween 80 was incorporated to obtain a ternary solid dispersion system. Dissolution tests were conducted and evaluated on the basis of cumulative percentage drug release and dissolution efficiency. Physicochemical characterizations of the solid dispersions were carried out using differential scanning calorimetric, powder X-ray diffraction, Fourier transform infrared spectroscopy, and scanning electron microscopy. Dissolution was remarkably improved in both systems compared to pure efavirenz (P<0.05). An optimum ratio was identified at a drug:polymer of 1:10. Incorporation of Tween 80 to 1:10 formulations formed using solvent method showed further improvement in the dissolution rate. Physicochemical characterization results suggested that efavirenz existed in the amorphous form in all the solid dispersion systems providing evidence of improvement in dissolution. No statistically significant difference (P>0.05) in dissolution was observed between the two methods. Binary and ternary solid dispersion systems both have showed a significant improvement in the dissolution rate of efavirenz. Formulations with only polyvinylpyrrolidone K30 showed best dissolution profile and 1:10 was identified as an optimum drug-polymer weight ratio.