MDR1 gene C3435T polymorphism and the risk of acquired aplastic anaemia

Field: Medicine (Health Sciences)

Although aplastic anaemia (AA) is thought to be mediated by an autoimmune destruction of haematopoiesis (Young, 2000), the aetiology of autoimmunity in AA is not fully understood. Some cases of acquired AA are an idiosyncratic consequence of exposure to certain drugs and chemicals but, even in this context, the mechanisms by which xenobiotics trigger the immune system have yet to be determined. Altered drug disposition and metabolism may be responsible for increased susceptibility, and pharmacogenetic variations are possible risk factors for idiosyncratic drug-induced AA (Marsh et al, 1999). The MDR1 gene encoded P-glycoprotein (P-gp), linked to multidrug resistance in leukaemia and expressed by normal haematopoietic cells, protects stem cells from toxins by actively extruding a variety of drugs (Chaudhary & Roninson, 1991). We observed previously that P-gp function was significantly reduced in T-cells (Calado et al, 1998) and bone marrow CD34+ cells of patients with AA, most significantly in patients with drug-induced AA (Calado et al, 2001). These findings point to a role of impaired P-gp function in determining predisposition to AA. However, the reason for reduced P-gp activity in this setting is elusive. Strong evidence has emerged recently to demonstrate that P-gp expression is genetically determined. The C3435T transition of the MDR1 gene correlates with reduced P-gp expression and function in the duodenum and CD56+ natural killer (NK) cells (Hoffmeyer et al, 2000; Hitzl et al, 2001). To evaluate whether the MDR1 gene C3435T polymorphism is associated with the occurrence of AA, we studied 35 patients with acquired AA (median, 26 years; range, 2–67 years). Classification was very severe in five, severe in 20 and moderate in 10 patients; 20 AA cases were idiopathic, 13 were related to drugs or chemicals (anti-inflammatory drugs in two, pesticides in nine and solvents in two) and two were hepatitis-associated AA cases; 105 age-, sex- and ethnicity-matched healthy subjects (blood donors) from the same geographical area were studied as controls. Blood and bone marrow samples were collected after informed consent. DNA was extracted from peripheral leucocytes to genotype patients and controls for the MDR1 C3435T polymorphism using polymerase chain reaction (PCR) followed by DpnII enzyme digestion (Hoffmeyer et al, 2000). A technician, who was unaware of the sample status, performed the genotyping. Odds ratios (OR) as a measure of relative risks and 95% confidence intervals (CI 95) were calculated by standard methods. Carrier frequency of 3435t was 62·8% amongst patients and 61% amongst controls, yielding an OR of 1·1 (CI 95, 0·5–2·4). or for heterozygosity was 1·3 (CI 95, 0·6–2·9); it was 0·6 (CI 95, 0·1–2·2) for mutant homozygosity (Table I). When patients with drug-induced AA were analysed separately (n = 13), the overall OR linked to the mutant genotype was 0·7 (CI 95, 0·2–2·3). Heterozygous and mutant homozygous states yielded or of 0·5 (CI 95, 0·1–2·1) and 1·2 (CI 95, 0·2–6·3) respectively. The MDR1 gene C3435T polymorphism is linked to reduced P-gp expression and function, and bone marrow CD34+ cells from 26 out of the 35 AA patients studied presented P-gp function (measured by the rhodamine 123 efflux assay) below the 25% percentile of previously studied normal controls (Calado et al, 2001). However, we failed to demonstrate a relationship between AA and C3435T. This lack of association may be explained by different reasons. First, the proportion of individuals developing AA after drug or chemical exposure is minimal compared with the prevalence of the C3435T polymorphism. It is possible that the decreased P-gp function observed in AA might be associated with other rare unknown MDR1 gene mutation(s) which might determine P-gp function in haematopoietic cells. Second, impaired P-gp function in AA might be the result of acquired rather than genetic factors. Testing whether defective P-gp expression is restricted to the haematopoietic tissue may serve to address this question. Finally, the number of patients analysed here was relatively small making it difficult to definitively rule out an association between MDR1 C3435T polymorphism and AA. In conclusion, although P-gp activity is decreased in AA patients, the MDR1 gene C3435T polymorphism does not seem to be a genetic risk factor for acquired drug-induced aplastic anaemia. The authors thank Sérgio M. Gabellini for technical assistance. This work was supported by Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP) grant 98/14247-6. R.T.C. was a recipient of FAPESP grant 00/13885-0.