High throughput digital quantification of mRNA abundance in primary human acute myeloid leukemia samples
Jacqueline E. Payton, … , Mark A. Watson, Timothy J. Ley
Jacqueline E. Payton, … , Mark A. Watson, Timothy J. Ley
Published May 18, 2009
Citation Information: J Clin Invest. 2009;119(6):1714-1726. https://doi.org/10.1172/JCI38248.
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Technical Advance Hematology

High throughput digital quantification of mRNA abundance in primary human acute myeloid leukemia samples

Abstract

Acute promyelocytic leukemia (APL) is characterized by the t(15;17) chromosomal translocation, which results in fusion of the retinoic acid receptor α (RARA) gene to another gene, most commonly promyelocytic leukemia (PML). The resulting fusion protein, PML-RARA, initiates APL, which is a subtype (M3) of acute myeloid leukemia (AML). In this report, we identify a gene expression signature that is specific to M3 samples; it was not found in other AML subtypes and did not simply represent the normal gene expression pattern of primary promyelocytes. To validate this signature for a large number of genes, we tested a recently developed high throughput digital technology (NanoString nCounter). Nearly all of the genes tested demonstrated highly significant concordance with our microarray data (P < 0.05). The validated gene signature reliably identified M3 samples in 2 other AML datasets, and the validated genes were substantially enriched in our mouse model of APL, but not in a cell line that inducibly expressed PML-RARA. These results demonstrate that nCounter is a highly reproducible, customizable system for mRNA quantification using limited amounts of clinical material, which provides a valuable tool for biomarker measurement in low-abundance patient samples.

Authors

Jacqueline E. Payton, Nicole R. Grieselhuber, Li-Wei Chang, Mark Murakami, Gary K. Geiss, Daniel C. Link, Rakesh Nagarajan, Mark A. Watson, Timothy J. Ley

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Figure 6

The validated 33-gene M3-specific signature is consistently dysregulated in other AML datasets and a mouse model of APL, but not in a PML-RARA+ cell line.

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The validated 33-gene M3-specific signature is consistently dysregulated...
The top portion of each GSEA plot shows the running enrichment score for the validated M3-specific genes as the analysis moves down the ranked list. The peak score for each plot is the enrichment score for the gene set. The bottom portion of each plot shows the value of the ranking metric as it moves down the list of ranked genes. The FDR is an expression of the significance level of the enrichment, after multiple test correction. (A) GSEA plot of 325 M0–M4 AML samples (GSE6891), comparing M3 with other FAB subtypes, demonstrates significant enrichment. (B) GSEA plot of mCG-PML-RARA murine APL cells (20, 24) compared with day 2 wild-type murine myeloid cells (mostly promyelocytes) demonstrates significant enrichment. (C) GSEA plot of uninduced versus PML-RARA–induced PR-9 cells demonstrates no enrichment of the M3-specific genes at any time point.