A Preliminary Study On Drug Resistance Mechanism Of Acute Promyelocytic Leukemia With Positive STAT5B-RARα Fusion Gene

Objective:We report a rare case of acute promyelocytic leukemia with STAT5B-RARαtranslocation that is insensitive to retinoic acid and arsenical.A series of in vitro experiments were conducted to explore the possible pathogenesis of STAT5B-RARαtranslocation in APL.So as to provide ideas for summarizing the characteristics of APL and exploring its pathogenesis and treatment strategies.Method:T-A cloning technique and gene sequencing were used to determine the fusion site of STAT5B-RARα fusion gene.PCR splicing method to clone the full length sequence of STAT5B-RARα fusion gene and the STAT5B-RARα expression vector was successfully constructed.The STAT5B-RARα gene was transfected into 293 T cells by transient and stable transfection.The green fluorescence expression was observed through fluorescence microscope,and the protein level expression of the gene was detected by Western blot.The stable transformants of STAT5B-RARαexpression were screened by virus infection in U937 cells.we treated NB4 cell lines,U937-p CDH-vector and U937-p CDH-STAT5B-RARα transfected cell line with ATRA to observe the changes of cell morphology、proliferation、cycle,etc.Rusult:1.The clinical features of this APL patient with STAT5B-RARα translocation are as follows: Bone marrow and peripheral blood smears have similar morphological characteristics to typical PML-RARα;Conventional PML-RARαfusion probe can not detect STAT5B-RARα translocation,which is easy to be missed in clinic.It is insensitive to ATRA and ATO induction therapy.2.The results of gene sequencing show that STAT5B-RARα fusion gene conforms to the common characteristics of RARα fusion gene,that is,the cleavage site of RARα gene is constant,which is in exon 3 and its 3’ sequence.And the western blot results show that the fusion protein mainly exists in the nucleus.After ATRA treatment,the expression of STAT5B-RARα CD11 b increased and the cell morphology changed,showing the cell differentiation and maturation phenomena such as smaller cells,depressed nuclei and divided leaves.The cell growth rate was also significantly inhibited after adding drugs,and the cell cycle was mainly blocked in G2/M phase.Conclusion:1.The APL of STAT5B-RARα translocation has the similar cell morphological characteristics of typical PML-RARα,which can not be detected by conventional PML-RARα fusion probe,so it is easy to miss diagnosis,and is not sensitive to ATRA and ATO induction therapy.2.The in vitro experiments showed that U937-p CDH-STAT5B-RARα transfected cell line had a good drug response to ATRA.3.The inconsistency between in vitro experimental results and clinical treatment response may be related to the heterogeneity of tumor environment in vivo or the regulation of other more complex pathways.