Mutations in the E-Domain of RARα Portion of the PML/RARα Chimeric Gene May Confer Clinical Resistance to All-transRetinoic Acid in Acute Promyelocytic Leukemia

Abstract

The binding of all-trans retinoic acid (ATRA) to the ligand-binding region in the E-domain of retinoic acid receptor-alpha modifies the transcriptional activity of RARalpha protein. ATRA probably induces differentiation of acute promyelocytic leukemia (APL) cells by binding to the E-domain of the RARalpha portion (RARalpha /E-domain) of PML/RARalpha chimeric protein. Therefore, molecular alteration in the RARalpha /E-domain of the chimeric gene is one mechanism by which patients with APL may acquire resistance to ATRA therapy. In this study using reverse transcription-polymerase chain reaction and single-strand conformation polymorphism, DNA segments amplified from the RARalpha /E-domain in fresh APL cells of 23 APL patients (8 males and 15 females from 4 to 76 years of age) were screened for mutations. Of those patients, 3 patients (1 with de novo and 2 with relapse) had clinical resistance to ATRA therapy. We found mutations in the RARalpha /E-domain of PML/RARalpha chimeric gene exclusively in the 2 patients who exhibited ATRA-resistance at relapse, whereas the mutations were not detected at their initial onset. Interestingly, these patients received a prolonged or intermittent administration of ATRA before relapse with ATRA-resistance. The mutations lead to the change of amino acid in the ligand-binding region of RARalpha /E-domain, Arg272Gln, or Met297Leu according to the amino acid sequence of RARalpha, respectively. Further study demonstrated that the in vitro ligand-dependent transcriptional activity of the mutant PML/RARalpha protein was significantly decreased as compared with that of wild-type PML/RARalpha. These findings suggest that mutations in the RARalpha /E-domain of the PML/RARalpha chimeric gene may confer clinical resistance to ATRA therapy in patients with APL.