BIRC 5 ( Survivin ) Splice Variant Expression Correlates With Refractory Disease and Poor Outcome in Pediatric Acute Myeloid Leukemia : A Report From the Children ’ s Oncology Group

Acute myeloid leukemia (AML) is characterized by numerous molecular abnormalities impairing differentiation and promoting proliferation [1]. Although recurrent cytogenetic and molecular abnormalities are powerful predictors of relapse and survival in both adult and pediatric AML, identifying reliable prognostic markers of primary refractory disease remains a challenge. Overall, approximately 20–30% of adults (<60 years) and up to 17% of children will fail to achieve complete remission (CR) with cytarabine and anthracycline based induction chemotherapy [1,2]. The prognosis for children with primary refractory disease is extremely poor, with only a third achieving remission with a second therapeutic attempt [3]. Novel prognostic markers that predict induction failure are needed to facilitate the early introduction of intensified and/or novel therapeutic strategies. Survivin, a small inhibitor-of-apoptosis protein (IAP) encoded by the BIRC5 gene, plays critical roles in apoptosis, cell division, and proliferation [4]. Several splice variants of survivin such as full-length survivin (-WT), survivin-2B, and survivin-DEx3 are well characterized [5]. Survivin-WT is encoded by four exons (I–IV), survivin 2B has an additional exon (IIB), and survivin DEx3 has deletion of exon III [5]. Expression of survivin and its splice variants correlates with white blood cell (WBC) count, cytogenetics, clinical response, and survival in AML [6–13]. Studies on survivin expression in childhood AML, however, have been restricted to relatively small cohorts (<50 patients) [6,9,12]. Recent preclinical data show that survivin is a transcriptional target of the FLT3-STAT pathway [14] and regulates proliferation in FLT3-mutated AML [15]. To determine whether survivin expression correlates with disease characteristics, such as FLT3mutational status, and clinical outcome, we performed molecular and expression characterization Background. The inhibitor-of-apoptosis protein survivin, encoded by BIRC5, regulates apoptosis, cell division and proliferation. Several survivin splice variants have been described however, the prognostic significance of their expression has not been well defined in pediatric acute myeloid leukemia (AML). Procedure.Quantitative expression analyses of BIRC5mRNA (n1⁄4306) and survivin transcript splice variants (n1⁄490) were performed on diagnostic bone marrow samples from childrenwith de novoAML treated on the clinical trials CCG-2961 and AAML03P1, then correlated with disease characteristics and clinical outcome. Results. Total BIRC5 expression did not correlate with clinical outcome. Fragment length analysis and sequencing of the entire BIRC5 transcript demonstrated three splice variants. The most prominent product, wild-type survivin, was expressed in all samples tested. Twominor transcriptswere present in 90 patients treated on CCG-2961; survivin-2B and a novel variant, survivin-DEx2, characterized by deletion ofBIRC5 exon II. A high 2B/ DEx2 expression ratio ( 1) correlatedwith increased diagnosticWBC count, monocytic phenotype, þ8 cytogenetics, lower complete remission (45% [n1⁄410] vs. 88% [n1⁄459], P<0.001) and higher induction failure rates (23% [n1⁄45] vs. 3% [n1⁄42], P1⁄40.009). Consistentwith this poor induction response, patients with a 2B/DEx2 ratio 1 had inferior 5-year survival rates (OS 36% vs. 60%, P1⁄40.011; EFS 23%vs. 53%at 5 years, P1⁄40.001) and appear to have increased relapse risk (P1⁄40.056). Subset analyses suggest that relative over-expression of 2B, rather than under-expression of DEx2 determines clinical response. Conclusions. High survivin-2B/DEx2 ratios are associated with refractory disease and inferior survival in childhood AML. Survivin splice variant expression warrants prospective evaluation in clinical trials. Pediatr Blood Cancer 2014;61: 647–652. # 2013 Wiley Periodicals, Inc.