An 81-year-old female presented with a short history of back pain with a physical examination proving unremarkable. Full blood count revealed pancytopenia (hemoglobin 8.8 g/dl, neutrophils 1.8 x 109/L, and platelets 140 x 109/L) and abnormal circulating promyelocytes that constituted 40% of nucleated cells. A coagulation screen was only mildly deranged with a prothrombin time of 15.1 seconds, an activated partial thromboplastin time of 29.4 seconds, and fibrinogen of 3.2 g/L. Peripheral blood promyelocytes, which accounted for 40% of nucleated cells, had regular nuclei, were hypergranular, and lacked Auer rods (Figure 1(a)) with increased numbers of pseudo-Pelger-Huet neutrophils noted (Figure 1(b)). Immunophenotyping detected autofluorescent blasts that expressed MPO, CD13, CD14, and CD33 but lacked CD34 and HLA-DR expression. The bone marrow was effaced by promyelocytes (Figure 1(c)) which stained strongly positive for Sudan Black. The patient commenced ATRA with a progressive concurrent increase noted in both peripheral blood myeloblasts and promyelocytes (white cell count 40.5 x 109/L), thrombocytopenia (platelets 92 x 109/L), and coagulopathy (fibrinogen 0.7 g/L) despite intensive blood product support. The chest findings were more consistent with aspiration and therefore no response to ATRA with differentiation syndrome excluded. Hydroxyurea and idarubicin were initiated in an attempt to achieve cytoreduction; however, progressive respiratory compromise and worsening coagulopathy led to the patients' death due to pulmonary hemorrhage ten days after presentation.
Cytogenetic G-band analysis identified an aberration involving 17q in 4/13 metaphases analysed. Metaphase fluorescent in situ hybridisation studies revealed a rearrangement involving RARA at 17q12 and confirmed the partner chromosome to be 11q, thus identifying the t(11;17) translocation with a full karyotype of 46,XX,add(17)(q21)[4]/46,XX[9].ish der(11)t(11;17)(q23;q21)(RARA+).nuc ish(PMLx2,RARAx3)[142/200]. A standardised reverse transcription-PCR approach did not detect PML-RARA transcripts, but, using primers previously described, a ZBTB16-RARA fusion transcript, consistent with fusion of ZBTB16 exon 3 fused to RARA exon 2, was detected and confirmed by Sanger sequencing (Figure 2). The reciprocal RARA-ZBTB16 fusion was not detected.
An NGS approach utilising a gene panel to detect additional mutations cooperating with the ZBTB16-RARA fusion in propagating APL was retrospectively employed. Amplicon libraries covering thirty commonly mutated genes implicated in myeloid malignancies were generated using genomic DNA from the diagnostic bone marrow aspirate using an Ion AmpliSeq  approach (Thermo Fisher Scientific, Paisley, UK). A single in-frame CEBPA p.P189delP mutation was detected (c.564-566 delGCC; reference sequence GRCh37: 19:33792755-7) with an allele frequency of 6.2%.