Evidence for an association of HLA-DRB1*15 and DRB1*09 with leprosy and the impact of DRB1*09 on disease onset in a Chinese Han population

Leprosy is a chronic infectious disease that occurs in genetically predisposed individuals. It is caused by infection with the intracellular macrophage pathogen Mycobacterium leprae. Leprosy is characterized by a spectrum of disease symptoms that result from interactions between the host immune response and invading M. leprae. At the lepromatous extreme, patients exhibit multibacillary infection and an absence of antigen-specific cellular immunity. At the tuberculoid extreme, patients exhibit paucibacillary infection and a strong cellular immune response. Between these two extremes, borderline leprosy patients show intermediate phenotypes [1].

Leprosy was once prevalent worldwide. However, there has been a steady decline in the prevalence of leprosy since the introduction of multidrug therapy by the World Health Organization (WHO-MDT) in 1982 [2]. The detection rate of new cases, however, remains steady in some countries, including several regions of China [3, 4]. Since 2002, nearly 1,600 new cases have been detected annually in China [4].

Previous studies have suggested a strong influence of genetic factors in regulation of the anti-M. leprae immune response [5, 6]. Two recent genome-wide linkage analyses have identified chromosome 6p21 as a major leprosy-susceptibility locus [7, 8]. This region harbors the HLA gene cluster, which has been extensively studied for its role in leprosy pathogenesis [9–12]. In particular, HLA-DR alleles have been consistently found to be associated with leprosy [13]. Several studies reported an association of the HLA-DR2 alleles HLA-DRB1*15, *16 *04, *10 and *12 with susceptibility or resistance to leprosy in Brazilian, Vietnamese, South Indian, Indonesian, Thai and Argentine populations [14–18]. In addition, HLA-DR3 alleles were also found to be associated with leprosy susceptibility in Surinamese and Mexican populations [19, 20]. These studies have not only shown the importance of the HLA-DR locus in leprosy susceptibility, but have also highlighted the fact that different HLA-DR alleles are associated with leprosy in different populations.

Although leprosy remains a major public health concern and one of the infectious diseases most capable of causing disability in China, the genetic susceptibility to leprosy in Chinese populations has received little study. The only such report was an investigation of the association of HLA-DRB1 with leprosy in Southern China; however, due to the small sample used, this study failed to detect this well-established association [21]. In the current study, we investigated the association of HLA-DRB1 with leprosy by genotyping DRB1 alleles in a large Chinese Han sample of 305 leprosy patients and 527 healthy control individuals. We further investigated the association of DRB1 alleles with clinical subtype and age-of-onset of the disease.

All markers with allele frequency above 0.01 have been tested and were in HW equilibrium.

Leprosy is a chronic infectious disease caused by M. leprae. The disease only occurs in a small percentage (1-3%) of infected individuals [23], an observation that supports the important role of host genetic factors in the development of leprosy. The reported male-to-female patient ratio is 2:1 [24], consistent with the known role of gender as an independent risk factor for leprosy susceptibility. In our study, more male cases were collected than female cases; thus, the observed association of the HLA-DRB1 allele with leprosy was based on a mostly male population.

Many studies have reported an association of HLA-DR2 with leprosy, a linkage that is consistent with the finding that HLA-DR antigens are associated with disease and the suggestion that a majority of restriction determinants for M. leprae reside on DR, and not DP or DQ molecules [25]. A functional study of Ag-specific T-cell responses within the context of HLA-DR has shown that HLA-DR-associated immunity may be crucial in the adaptive immune response to infection [26]. Thus, HLA-DR plays a major role in the presentation of M. leprae antigens to T cells in leprosy patients.

By analyzing a large number of Chinese leprosy patients, our study has provided the first evidence for an association of HLA-DRB1 with leprosy in a Chinese population and provided additional support for the important role of HLA in the pathogenesis of leprosy. HLA-DRB1*15 has been demonstrated to be a leprosy-susceptibility allele among Brazilian [14] and Indian [15] populations. In our study, we found that HLA-DRB1*15 was also associated with an increased risk of leprosy in a Chinese population, and exhibited an allele frequency (32%) similar to that observed in Indian populations (31%) but higher than that observed among Brazilians (15%). The HLA-DRB1*09 allele was found to be associated with protection against leprosy in our study; its frequency was significantly lower in leprosy patients than in controls, and the association remained significant after correcting for multiple testing (P = 0.002, Pc = 0.026). This allele also showed a protective effect against the disease in Southern India, but the allele frequency among Indian patients was 0.017 [15], much lower than that observed in the Chinese population studied here (0.08).

It has been reported that the HLA lymphotoxin-alpha (LTA)+80 locus within the 6p21 chromosomal region is a major risk factor for early-onset (i.e., < 16 years old) leprosy [27]. To investigate whether the effect of HLA-DRB1* on leprosy risk was age-onset dependent, we stratified leprosy patients into early-onset (≤ 16 years) and late-onset (> 16 years) groups in association analyses. Interestingly, we found that HLA-DRB1*09 was associated with protection effect against early-onset leprosy (P = 0.003), but not late-onset leprosy (P = 0.285). In contrast, the HLA-DRB1*15 allele showed no age-onset-dependent effects. To our knowledge, this is the first report that HLA-DRB1*09 has an impact on disease onset. Further studies will be needed to investigate the association of the LTA+80 locus with leprosy in Chinese populations and determine whether the association at the HLA-DRB1 locus is independent of the LTA+80 locus.

De Vries et al. have proposed a two-step model for the development of leprosy, proposing that successful infection of M. leprae is first established in genetically vulnerable individuals, and subsequent clinical manifestation of the disease is influenced by additional host and environmental factors [28]. In our study, the frequencies of HLA-DRB1 alleles were similar in the MB and PB patient groups. This is consistent with the two-step model, suggesting that the HLA-DRB1 locus may influence the overall susceptibility to leprosy per se. It has also been reported that certain HLA-DRB1 alleles are associated with MB or PB forms of leprosy [29–31], suggesting that HLA-DRB1 may also be involved in the clinical manifestation of the disease.

Taken together, our association analysis of stratified patient groups indicates that the HLA-DRB1 locus is largely associated with leprosy per se, and only HLA-DRB1*09 shows an age-onset-dependent effect.

In summary, by analyzing a large sample of Chinese Han leprosy patients, our genetic association study of the HLA-DRB1 locus provides strong evidence for an association of HLA-DRB1*15, as a susceptibility allele, and DRB1*09, as a protective allele, with leprosy in a Chinese population. Furthermore, we demonstrate for the first time that HLA-DRB1*09 has an impact on the onset of disease, exerting a protective effect only against early-onset leprosy. These alleles could act alone or in combination with other genes to confer susceptibility or resistance to leprosy in Chinese Han populations.