a 68yearold japanese woman was admitted to otemae hospital ( osaka city , japan ) with pancytopenia in may 2012 . initial complete blood counts showed : hemoglobin , 8.2  g / dl ; platelets , 48    10/l ; and white blood cells , 1.65    10/l ( including 1% promyelocytes ) . a bone marrow sample showed marked hypercellularity ; 36.8% were promyelocytes with azurophilic granules and auer rods ( fig . flow cytometry revealed apl cells positive for cluster of differentiation ( cd ) 33 , cd34 , cd64 , cd117 , and human leukocyte antigen ( hla)dr , and negative for cd2 , cd7 , cd13 , and cd19 . fluorescence in  situ hybridization with dual fusion translocation probes detected no typical rara split signals , but rather a small rara signal and two large signals ( 46/100 ) ( fig . accordingly , the patient was diagnosed with apl , and remission induction therapy was initiated with alltrans retinoic acid ( atra ; 45  mg / m / day ) . on day 12 , promyelocytes in the peripheral blood increased to > 10    10/l ; therefore , we added idarubicin ( 9  mg / m / day from days 12 to 14 ) and cytosine arabinoside ( 75  mg / m / day from days 12 to 18 ) . however , reinduction therapy was initiated with gemtuzumab ozogamicin ( go ) ( 9  mg / m ) because of high cd33 expression . one month later , morphological examination and fish of bone marrow aspirate showed hematological remission . in april 2013 , the patient experienced molecular relapse as determined by an increase of wilms tumor1 mrna in the peripheral blood . she was readministered with go ( 9  mg / m ) , and remained in hematological remission until august 2013 when she developed syncope . magnetic resonance images revealed several enhanced regions in the right brain , and her cerebrospinal fluid ( csf ) clearly contained apl cells . because her level of consciousness was depressed despite highdose methotrexate and cytarabine infusion , whole brain irradiation was undertaken . one month later , her neurological symptoms disappeared , and her csf cytology revealed no apl cells . in february 2014 , she developed bilateral leg weakness and walking difficulty . lumbar magnetic resonance imaging showed enhancement in the conus and nerve roots , and apl cells were once again detected in her csf . morphological , cytogenetic , and molecular features of a bone marrow sample from a 68yearold japanese woman with acute promyelocytic leukemia . ( b ) interphase fish using promyelocytic leukemia ( pml ) ( 15q22 ; green ) and retinoic acid receptor  ( rara ) ( 17q21 ; red ) probes did not reveal a pml ( c ) interphase fish assay to detect rara split signal ( using 3rara [ green ] and 5rara [ red ] probes ) showed no split signals but rather a small yellow signal ( 46/100 ; arrow ) . ( d ) mrna expression of interferon regulatory factor 2 binding protein 2 ( irf2bp2 ) and irf2bp2rara . lane 1 shows the result of using two complementary oligonucleotide primers , fp1 and rp1 . the upper detected band was predicted to be 206  bp and the lower band was predicted to be 158  bp ; these bands were supposed to correspond to the expression of normal splice variants of irf2bp2 in the patient 's bone marrow . the upper detected band was predicted to be 398  bp and the lower band was predicted to be 350  bp ; these bands were supposed to correspond to the expression of the fusion variants of irf2bp2rara . the results confirmed that this patient expressed two variants of irf2bp2 ( data not shown ) , as well as the predicted fusion mrnas of irf2bp2rara ; the upper and lower panels show the sequences of the fusion genes from irf2bp2 variants 1 and 2 , respectively . the blue and yellow solid bars represent the coding regions of the irf2bp2 and rara genes , respectively . variant 1 uses an alternate inframe splice junction in the 3 coding region of exon 1 , resulting in an mrna transcript 48  bp longer than variant 2 . the irf2bp2rara fusion is expected to generate two types of fusion mrna depending on the irf2bp2 variants . in the longer type , base 1079 of the irf2bp2 mrna is likely to be fused to base 769 of the rara 
 mrna . in the shorter type , the base 1031 of the irf2bp2 
 mrna is likely to be fused to base 769 of the rara 
 mrna . ( g ) overviews of genomic dna of the irf2bp2,rara , and predicted irf2bp2rara fusion genes . after obtaining written informed consent and institutional review board approval , this translocation gives rise to a putative fusion protein comprising exon 1 of irf2bp2 and exons 39 of rara . irf2bp2 comprises two exons with an intron , and there are two splicing sites within irf2bp2 exon 1 . accordingly , we generated primer pairs to amplify the cdnas of irf2bp2 ( fp and rp1 ) and irf2bp2rara ( fp and rp2 ) . as predicted , two fragments were amplified by rtpcr for both irf2bp2 and irf2bp2rara ( fig . the pcr product sequences corresponded well to the splicing sites ( at the 1031 and 1079  bp positions of irf2bp2 cdna ) ( fig . the rara transcript started at the beginning of exon 3 ( at 769  bp of rara cdna ) . thus , our additional mrna analysis clearly indicated that the irf2bp2rara fusion proteins showed variant forms that were dependent on irf2bp2 splicing . the transcript structures for irf2bp2 and rara , as well as irf2bp2rara , are shown in figure  1f . taken together , the irf2bp2rara gene is likely to be positioned between intron 1 of irf2bp2 and intron 2 of rara ( fig . to our knowledge , this is the second report of a novel apl variant with irf2bp2rara ; this case uniquely showed atypical morphological and clinical features with no dic detected at admission . classic apl cells with pml  rara typically express mature myeloid markers , including cd13 and cd33 , and are negative for cd34 and hladr . moreover , several reports have shown expression of cd34 and/or hladr in a significant proportion of newly diagnosed apl patients.11 , 12 , 13 in our case , the apl cells were positive for cd33 , cd34 , cd64 , cd117 , and hladr , and negative for cd2 , cd7 , cd13 , and cd19 . fluorescence in  situ hybridization with dual fusion translocation probes is useful for detecting unknown rearrangements of the rara gene.14 herein , we failed to detect pml rara fusion ; however , use of a rara dual fusion translocation probe allowed us to detect a novel rearrangement of the rara gene . cases of apl with cryptic pml  rara rearrangement that were negative by fish and conventional karyotyping but positive by rtpcr have been reported;15 these findings were caused by small cryptic insertions . a normal yellow signal indicated normal 17q while another indicated a small deletion in 17q ; furthermore , an abnormal small yellow signal indicated a small insertion of rara ( fig .  1 ) . the irf2bp2cdx1 fusion gene has been reported in mesenchymal chondrosarcoma,16 and there is one previous report of irf2bp2rara in apl.10 yin et  al.10 reported a case of apl with irf2bp2rara that involved exon 2 of irf2bp2 and intron 2 of rara , with breakpoints at positions 1687  bp in irf2bp2 and 41620  bp in rara . the patient achieved cr with atra , arsenic trioxide , and go ; however , relapse occurred 2  months after consolidation therapy . our case differed in its atypical morphology and clinical course , as well as the lack of dic on admission . tissue factors and cancer procoagulant expressed in apl cells are associated with dic ; the lack of dic in our case may be explained by the lower number of apl cells compared to that in the previously reported patient . most patients with apl can achieve cr with atra with or without anthracycline , resulting in favorable clinical outcomes.17 , 18 , 19 in the previous irf2bp2rara case , clinical sensitivity to atra was not properly evaluated because of combination therapy with arsenic trioxide and go . our patient did not respond to atra monotherapy . in the presence of pharmacological concentrations of atra , transcriptional activation can be induced by dissociation of the corepressor complex from pml  rara as well as proteasomedependent pml , atra fails to recognize rara , and corepressor dissociation does not occur , resulting in atra resistance.14 other fusion genes such as plzf the nuclear receptor corepressor ( ncor)/silencing mediator for retinoid and thyroid hormone receptors ( smrt ) corepressor complex interaction with plzf and stat5b translocation is thought to be involved in this resistance . irf2bp2 also interacts with ncor corepressor proteins;21 we speculated that irf2bp2rara is involved in atra resistance because of this . in conclusion the patient had some peculiar clinical characteristics , such as no coagulopathy , a unique cell surface phenotype , and atra resistance . although apl with irf2bp2rara is rarely observed , its unique characteristics have clinical implications for the diagnosis and treatment of apl patients . appendix  s1 primer information for amplification of cdnas of irf2bp2 ( fp and rp1 ) and irf2bp2rara ( fp and rp2 ) .
we present an acute promyelocytic leukemia ( apl ) patient with two subtypes of irf2bp2rara , in which the irf2bp2 gene showed completely new breakpoints . 
 bone marrow examination revealed morphologic features indicative of apl . 
 however , promyelocytic leukemia 
 rara fusion was not detected . a pairedend mrna sequencing followed by rtpcr and 
 direct sequencing revealed two types of fusion transcripts between exon 1b of irf2bp2 and exon 3 of rara . 
 the patient received alltrans retinoic acid and conventional chemotherapy , but showed resistance . 
 this is the second report of irf2bp2 involvement in apl , and we describe various breakpoints for the irf2bp2rara fusion gene .