Nature Genetics: doi: /ng Supplementary Figure 1. HOX fusions enhance self-renewal capacity.
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1 Supplementary Figure 1 HOX fusions enhance self-renewal capacity. Mouse bone marrow was transduced with a retrovirus carrying one of three HOX fusion genes or the empty mcherry reporter construct as described in Online Methods. (a) At 1-week intervals, colonies were counted and cells were replated in duplicate for a total of four platings. Each bar donates colony counts for a replating round. Error bars indicate standard error of the mean. Data are compiled from two separate experiments with similar results. (b) Flow cytometry of colonies. Data shown are from one of two experiments with similar results.
2 Supplementary Figure 2 HOX rearranged subgroup gene expression signatures. (a) HOX fusion partner genes are upregulated in HOX rearranged cases. Relative HOX gene expression values (row percentiles) are indicated by heat color (color key). Genes are ordered by chromosomal position. Arrows indicate direction of transcription of proteincoding HOX genes. Fusion genes are divided by coding (top) and non-coding HOX fusions and ordered by HOX fusion partner. Right column summarizes the expression value of the HOX gene involved in the rearrangement. (b) HOXA9 target genes are upregulated in the HOX rearranged subgroup. Distribution of expression of genes within the HOXA9 target upregulated gene set showing the highest association with AMKL subgroup 10. Expression is highest in the HOX rearranged cases for both protein coding and non-protein coding HOX fusions (red circles).
3 Supplementary Figure 3 Genomic driver mutations in adult AMKL. Recurrently mutated genes in the adult cohort as determined by whole exome sequencing, copy number alterations, and cytogenetics (see Supplementary Tables 7 and 8 for detailed information on each mutation).
4 Supplementary Figure 4 Genomic driver mutations in pediatric AMKL. Recurrently mutated genes in the pediatric cohort as determined by whole exome sequencing, copy number alterations, and cytogenetics (see Supplementary Tables 6 and 8 for detailed information on each mutation).
5 Supplementary Figure 5 Down syndrome AML and chromosome 21 gene expression patterns. (a) GATA1 mutant non-ds-amkl and DS-AML gene expression patterns correlate. Scatterplots of relative gene expression values (log 2 ratios) of differentially expressed genes in DS-AML compared to non-ds AML (y-axes) and those of the different non-ds AMKL subgroups relative to the other groups combined (x-axes). Spearman correlation coefficients of comparisons are shown to the right. (b) Chromosome 21 expression levels. Relative expression of genes on chromosome 21 in the AMKL subgroups is shown. Genomic amplification status for each sample is indicated by the circles. Filled circles, amplified chromosome 21; open circles, chromosome 21 status not analyzed/unknown.
6 Supplementary Figure 6 RB1 expression levels. RB1 expression levels as determined by RNA-seq are shown for NUP98-KDM5A specimens vs. others.
7 Supplementary Figure 7 Cooperation of MPL mutations and HOX rearrangements. Murine bone marrow was co-transduced with a retrovirus carrying a HOX fusion gene and an MPL gene in the following combinations: mcherry (MIC)+GFP(MIG); MIC+MPL wild type (MPL wt ); MIC+MPL S505N mutant (MPL S505N ); MIC+MPL W515L mutant (MPL W515L ); GATA2-HOXA9+MIG; GATA2-HOXA9+MPL wt ; GATA2-HOXA9+MPL S505N ; GATA2-HOXA9+MPL W515L ; NIPBL-HOXB9+MIG; NIPBL- HOXB9+MPL wt ; NIPBL-HOXB9+MPL S505N ; NIPBL-HOXB9+MPL W515L. Double-positive transduced cells were flow sorted using mcherry and GFP markers. (a) MPL mutations do not enhance self-renewal. Sorted cells were grown on methylcellulose supplemented with
8 cytokines. At 1-week intervals, colonies were counted and cells were replated for a total of three platings. Each bar donates colony counts for a replating round. Error bars indicate standard error of the mean. Data are compiled from two separate experiments with similar results. (b) MPL mutations enhance cytokine-independent growth. 5 x 10 5 cells from the third CFU plating were removed and placed in liquid culture in the absence of cytokines in triplicate. Viable cells were counted daily for one week. Error bars indicate standard error of the mean. Data shown are from one of two experiments with similar results.
9 Supplementary Figure 8 MPL mutations enhance JAK2 signaling. Colonies from the third plating as described in Supplementary Figure 7 were removed and placed in liquid culture in the absence of cytokines. After 48 hours of growth, cells were harvested and subjected to western blot analysis. The ratio of phosphorylated JAK2 to total JAK2 protein as determined by densitometry and phosphorylated STAT5 to total STAT5 protein are shown below. Data shown are from one of two experiments with similar results.
10 Supplementary Figure 9 Non-DS-AMKL subgroups correlate with outcome. (a-c) Comparison of event free survival (a), overall survival (b), and cumulative incidence of relapse or non-response (c) across AMKL subgroups. Each group was tested against the other groups combined. P values were determined by log-rank tests from either Cox proportional hazards regression models (a,b) or Gray's K-sample tests (c). All P values for competing risk differences were > 0.25.
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Supplementary Figure S1 Expression of mir-181b in EOC (A) Kaplan-Meier curves for progression-free survival (PFS) and overall survival (OS) in a cohort of patients (N=52) with stage III primary ovarian
Nature Getetics: doi: /ng.3471
Supplementary Figure 1 Summary of exome sequencing data. ( a ) Exome tumor normal sample sizes for bladder cancer (BLCA), breast cancer (BRCA), carcinoid (CARC), chronic lymphocytic leukemia (CLLX), colorectal