Identification of variant APL translocations PRKAR1A-RARα and ZBTB16-RARα (PLZF-RARα) through the MI-ONCOSEQ platform

Abstract

The cornerstone of management in patients with acute promyelocytic leukemia (APL) is early diagnosis and prompt initiation of treatment with an all-trans retinoic acid (ATRA)-based regimen. Identification of the t(15;17)(PML-RARA) chromosomal translocation through conventional cytogenetics fluorescence in-situ hybridization (FISH) or detection of the promyelocytic leukemia-retinoic acid receptor alpha (PML-RARα) fusion through RT-PCR represent the current standard of care for diagnosing APL. However, about 1–2% of patients with APL have a variant translocation involving other fusion partners with RARα besides PML. These patients present a unique diagnostic and clinical challenge in that conventional cytogenetics in addition to FISH and/or RT-PCR for PML-RARα may fail to identify these clinically relevant genetic lesions leading to an inappropriate diagnosis and treatment. We present two cases of patients who had APL with variant translocations whose bone marrow specimens were sent to the University of Michigan for enrollment in the MI-ONCOSEQ study (HUM00067928) after standard testing failed to identify PML-RARα or t(15;17) despite a phenotypic concern for this diagnosis. In these two patients, whole exome and transcriptome profiling via the MI-ONCOSEQ platform identified a PRKAR1A-RARα fusion in one patient and ZBTB16-RARα fusion in another patient. These cases illustrate the utility of whole exome and transcriptome profiling in diagnosing variant translocations in patients in whom there is a high clinical suspicion for APL based on hematopathology review.

Introduction

The discovery of the characteristic translocation t(15;17)(q22;q12) leading to the PML-RARα fusion transgene and the development of ATRA-based therapy have revolutionized diagnosis and treatment of acute promyelocytic leukemia (APL).[1] Once a disease characterized by fatal DIC and refractoriness to chemotherapy, patients with APL often have a more optimistic prognosis and now receive induction therapy per risk-stratified guidelines with combinations of ATRA, arsenic trioxide (ATO), and/or cytotoxic chemotherapy (anthracyclines and cytarabine).[2]

The favorable prognosis in APL depends, however, on rapid diagnosis and immediate initiation of ATRA to reduce the mortal complications of disseminated intravascular coagulopathy.[3] Morphological characteristics of the disease can raise suspicion for a diagnosis of APL, which may be further supported by immunohistochemistry (IHC) or flow cytometry. However, demonstration of the t(15;17) through conventional karyotype analysis or FISH or detection of the PML-RARα fusion transcript with RT-PCR remains the gold standard for diagnosis. While most APL patients show the typical t(15;17)(q24;q21) on conventional karyotype, up to 8% of cases lack this classical finding.[3] In most cases, the PML-RARα gene fusion can still be detected through FISH (dual fusion or break-apart probes) or via RT-PCR. When this occurs, the lack of the classic t(15;17) on karyotype can be attributed to complex (3 or 4-way) translocations or cryptic (submicroscopic) insertions. The clinical features and prognosis of these patients are not thought to be affected by these unusual cytogenetic findings.[4]

In 1–2% of cases of APL, neither t(15;17) nor PML-RARα can be detected by conventional testing.[5] These patients’ APL may be driven by variant translocations involving other fusion partners of RARα besides PML. To date, 15 additional fusion partners to RARα have been identified: ZBTB16 (also known as PLZF), NPM1, NuMa, STAT5b, PRKAR1A, FIP1L1, BCOR, NABP1 (also known as OBFC2A), IRF2BP2, GTF2I, TBLR1, FNDC3B, ADAMTS17, STAT3, and TFG.[5], [6], [7], [8], [9], [10], [11] ZBTB16-RARα, often involving a balanced chromosomal translocation t(11;17)(q23;q21), is the most common variant translocation, occurring in 0.8% of APL cases. Interestingly, APL with ZBTB16-RARα shows a regular contour to its nucleus and demonstrates resistance to ATRA, leading to a worse prognosis than APL with the conventional t(15;17).[5] Other variant translocations including those with Stat5b, TBLR1, STAT3, and GTF2I are also reportedly insensitive to ATRA.[10] However, the rarity of these translocations, often occurring only as case reports or small series, precludes more generalizable clinical characterization at this time.[5,10]

There is no standard approach to diagnosing variant translocations in APL. However, it is critical that patients with clinical/morphological suspicion for APL and a negative work-up for both t(15;17)(q24;q21) and/or PML-RARα be assessed for a variant translocation to allow for prompt initiation of appropriate treatment. Herein, we characterize two patients with clinical/morphologic suspicion for APL whose bone marrow specimens were sent to the University of Michigan for enrollment in the MI-ONCOSEQ study (HUM00067928) after failure to locally identify t(15;17)(q24;q21) or PML-RARα. In both cases, variant translocations were identified through a next generation sequencing platform, thus demonstrating the utility of whole exome and transcriptome profiling via MI-ONCOSEQ as a potential tool for identifying patients with variant translocations and rendering appropriate treatment in a timely fashion.