Transient Myeloproliferative Disorder and Acute Myeloid Leukemia in Down Syndrome

Abstract Context.—A specific mutation, JAK2V617F, was recently recognized as having diagnostic value for myeloproliferative disorders. No practical assay is currently available for routine use in a clinical laboratory. Objective.—We report the development of a real-time polymerase chain reaction melting curve analysis assay that is appropriate for molecular diagnostics testing. Design.—Specific primers and fluorescence resonance energy transfer probes were designed, and patients with a previously diagnosed myeloproliferative disorder, de novo acute myeloid leukemia, or reactive condition were selected. The DNA was extracted from fresh and archived peripheral blood and bone marrow specimens, and real-time polymerase chain reaction melting curve analysis was performed on the LightCycler platform (Roche Applied Science, Indianapolis, Ind). Results.—The JAK2 region was successfully amplified, and wild-type amplicons were reproducibly discriminated from JAK2V617F amplicons. Titration studies using homozygous wild-type and mutant cell lines showed the relative areas under a melting curve were proportional to allele proportion, and the assay reliably detected one mutant in 20 total cells. JAK2V617F was identified in patients previously diagnosed with a myeloproliferative disorder or acute myeloid leukemia transformed from myeloproliferative disorder, whereas a wild-type genotype was identified in patients with reactive conditions or de novo acute myeloid leukemia. Conclusions.—These findings demonstrate the suitability of this assay for identifying JAK2V617F in a clinical laboratory setting. Furthermore, the semiquantitative detection of JAK2V617F in archived specimens provides a new tool for studying the prognostic significance of this mutation.

Download Full-text

Differences of Somatic Mutations and Gene Expression in Blasts of Transient Leukemia and Acute Myeloid Leukemia of Down Syndrome

Blood ◽

10.1182/blood.v124.21.2364.2364 ◽

2014 ◽

Vol 124 (21) ◽

pp. 2364-2364

Author(s):

Jian Chen ◽

Fouad Yousif ◽

Timothy Beck ◽

John D McPherson ◽

Johann K. Hitzler

Keyword(s):

Gene Expression ◽

Acute Myeloid Leukemia ◽

Down Syndrome ◽

Myeloid Leukemia ◽

Somatic Mutations ◽

Digital Pcr ◽

Droplet Digital Pcr ◽

Transient Leukemia ◽

Genetic Events ◽

Acute Myeloid

Abstract Background: Transient leukemia (TL) occurs in 30% of newborns with Down syndrome (DS) and typically resolves spontaneously. Approximately 20% of infants with TL go on to develop acute myeloid leukemia of DS (DS-AML) within the first four years of life. The blasts of both TL and DS-AML harbor somatic mutations of GATA1 . The objective of this study was to identify additional genetic events, which associated with the progression of TL to DS-AML. Methods: Leukemic blasts of TL, DS-AML and normal T lymphocytes were sorted from blood and bone marrow samples of five patients who successively developed both disorders. In addition, blasts of one patient with subsequent relapse of DS-AML were analyzed. Mutational spectrum and gene expression and were determined by exome sequencing and RNASeq (Illumina HiSeq2000). The presence of mutations, which were identified with this approach in DS-AML blasts, was examined by droplet digital PCR in TL blasts (BioRad QX200). Results: Blasts of TL overall harbored fewer mutations than those of DS-AML. Mutations of cohesin and RAS pathway genes were identified in a subset of DS-AML but not TL. In the patient who developed a relapse, different cohesin gene mutations were detected at initial diagnosis of AML and relapse; a minor clone present at initial diagnosis of AML emerged as the predominant clone at relapse. Concordant somatic GATA1 mutations were present in both TL and DS-AML blasts derived from the same patient. In contrast, other genetic events, which were detected in DS-AML blasts by exome sequencing, were confirmed to be absent in TL (by droplet digital PCR). The majority of differentially expressed genes showed higher expression levels in blasts of TL compared to DS-AML. They included genes encoding chemokines and related to IL1 and TGFb signaling. Conclusions: The pathogenic sequence starting with TL and culminating in AML is uniquely initiated in children with DS by somatic mutation of GATA1. In contrast, the events associated with the transformation of TL to DS-AML resemble progression factors also found in non-DS AML. These progression events were not detectable even in minor subclones of TL suggesting they are acquired after the onset of TL. This research was supported by funding from the Canadian Cancer Society Research Institute and Ontario Institute for Cancer Research. Disclosures No relevant conflicts of interest to declare.

Download Full-text