Elsevier

Tuberculosis

Volume 94, Issue 3, May 2014, Pages 299-305
Tuberculosis

Host Genetics of Susceptibility
The role of cigarette smoking and liver enzymes polymorphisms in anti-tuberculosis drug-induced hepatotoxicity in Brazilian patients

https://doi.org/10.1016/j.tube.2014.03.006Get rights and content

Summary

Tuberculosis (TB) is still a major health concern and side-effects related to the treatment, especially drug-induced hepatotoxicity (DIH), should be better investigated. In the present study, a possible association between anti-TB DIH and cigarette smoking, N-acetyltransferase 2 (NAT2), Cytochrome P450 2E1 (CYP2E1) and Cytochrome P450 3A4 (CYP3A4) genotypes was studied in 131 TB Brazilian patients. The NAT2 and CYP3A4 genetic polymorphisms were determined using a polymerase chain reaction (PCR) direct sequencing approach and genetic polymorphisms of CYP2E1 gene were determined by restriction fragment length polymorphism (RFLP). The risk of anti-TB DIH was lower in rapid/intermediate acetylators when compared to slow acetylators (OR: 0.34, CI 95: 0.16–0.71; p < 0.01). A decreased risk of developing anti-TB DIH was also observed in active smokers when compared to non-smokers (OR: 0.28, 95 CI: 0.11–0.64; p < 0.01). Significant association between CYP3A4 genotypes and hepatotoxicity was not observed, as well as between CYP2E1 genotype and hepatotoxicity, whose frequency of patients with wild homozygous was more prevalent. The anti-TB drugs interactions with smoking on hepatotoxicity, as well as the NAT2 phenotype, may require to adjust therapeutic regimen dosages or alarm in case of adverse event developments.

Introduction

Tuberculosis (TB) is a major health issue, especially in developing countries, that account for 95% of tuberculosis cases and deaths worldwide according to the World Health Organization (WHO). In Brazil, the notifications recorded more than 70,000 new TB cases in 2012 and more than 10,000 of these new cases occurred in the state of Rio de Janeiro. Although the TB death rate dropped 41% between 1990 and 2011 (data from WHO), side-effects related to the treatment, especially DIH, still increase morbidity and mortality.

Risk factors associated to DIH include age over 60 years [1], female gender [2], [3], poor nutritional status [4], alcohol intake [5], [6], [7], [8] and treatment regimens, among others. Concerning tobacco use, it has been shown that ever smokers are more likely to have cough, dyspnea, chest radiograph appearances of upper zone involvement, cavity and miliary appearance, and positive sputum culture, but are less likely to have isolated extra-pulmonary involvement than non-smokers. Smoking has been found to be associated with both relapse of TB and TB mortality [9].

Keshavjee et al. [10] demonstrated that smoking was associated with worse outcomes in multidrug-resistant tuberculosis patients. Constituents of cigarette smoke, including polycyclic aromatic hydrocarbons, induce several drug-metabolizing enzymes and may interfere with drug clearance [11]. However, most studies do not analyze cigarette smoking as an individual possible risk factor to DIH.

In anti-TB therapy with isoniazid, rifampicin and pyrazinamide, isoniazid has been described as the most common drug associated to hepatotoxicity [8]. Concerning this, genetic polymorphisms in key enzymes associated with isoniazid metabolism also need attention.

In the liver, isoniazid is initially acetylated to acetylisoniazid by the enzyme N-acetyltransferase 2 (NAT2) and then hydrolyzed into acetylhydrazine and isonicotinic acid. Acetylhydrazine is hydrolyzed into hydrazine or acetylated into diacetylhydrazine, a non-toxic residue. Alternatively, acetylhydrazine is oxidate by CYP2E1 generating toxic metabolites. Studies suggest that hydrazine is probably the cause of isoniazid-induced hepatotoxicity [12].

According to NAT2 genotype, individuals can be classified in three acetylator phenotypes: fast, intermediate and slow. The association between acetylator phenotype and isoniazid-induced hepatotoxicity is, however, still controversial. Initial studies have demonstrated that patients with a fast acetylator phenotype had a higher risk of developing hepatotoxicity [13], [14].

More recently, Leiro-Fernandez et al. [15] did not find a significant correlation between acetylator phenotype and hepatotoxicity. However, Santos and et al. [16] and Gupta et al. [17] showed a high correlation between slow acetylators and hepatotoxicity. Corroborating these data, a recent meta-analysis comprising 14 studies showed that TB patients with slow acetylator profile had an increased risk of developing DIH [18].

Concerning liver enzymes other than NAT2, genetic polymorphisms in genes that codify microsomal Cytochrome P450 (CYP) enzymes family could also be related to anti-TB DIH. Different groups have looked at CYP2E1 genetic polymorphisms because this enzyme plays a role in the metabolic pathway of isoniazid [3], [16], [17], [19], however the results have been controversial. Huang et al., for example, showed an association of the CYP2E1 wild type genotype (c1/c1) with an increased risk of DIH [20]. On the other hand, others studies revealed a negative association between hepatotoxicity and CYP2E1 genotype [3], [16], [17]. These differences may be related to different studied populations' ethnicity and divergent criteria used to define hepatotoxicity.

Another CYP enzyme that could be related to anti-TB DIH is CYP3A4. This enzyme is induced by rifampicin and this induction leads to increased release of toxic metabolites by isoniazid metabolism [8]. However, to our knowledge, there are no studies correlating CYP3A4 polymorphisms to anti-TB DIH.

Our group has demonstrated that CYP3A4 5′ region is highly conserved in the analyzed Brazilian population. However, a single nucleotide polymorphism was detected: c.-392A>G [21]. Therefore, the aim of the present study is to investigate a possible correlation between cigarette smoking and anti-TB DIH in Brazilian TB patients, as well as the contribution of NAT2 acetylation status, CYP3A4 and CYP2E1 genetic polymorphisms and clinical/demographical variants to this adverse event occurrence.

Section snippets

Study design and patients

In this retrospective cohort study, we analyzed patients who were treated for tuberculosis at Evandro Chagas Clinical Research Institute (IPEC), Oswaldo Cruz Foundation (FIOCRUZ), Rio de Janeiro, Brazil from 2001 to 2008. The IPEC-FIOCRUZ Ethics Committee approved this research under the SISNEP register: 0013.0.009.000-03/2003.

The eligibility criteria were: signed written consent, sputum smear with acid-fast bacilli or culture positive for Mycobacterium tuberculosis; ongoing TB treatment and

Patient characteristics

A total of 131 TB patients were analyzed in the present study. Among them, 66.4% were male. Most of them were non-white (58%) and the average age was 39.8 years old. The pulmonary form of TB was the most frequent (57.2%), followed by extra-pulmonary (32%) and disseminated-TB (17.5%). Not all patients had serological results for HIV, HCV and HBV. Considering that, 61/129 patients (47.3%) were HIV-positive and 45 patients were on highly active antiretroviral therapy (HAART). HCV prevalence was

Discussion

The understanding of mechanism of anti-TB DIH becomes important to increase TB treatment success rate. In the present study, we analyzed a cohort of Brazilian TB patients from 2001 to 2008.

The 39.7% frequency of DIH found here is above the incidence range of variation that has been reported in several studies (2–28%) [12], although the frequency rate of DIH depends on the investigators' definition of hepatotoxicity as well as the population studied, the high rate of DIH, is probably due the

Acknowledgments

The authors thank the medical, technical and nursing staff of the Evandro Chagas Clinical Research Institute (IPEC) for their support.

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