SUPPLEMENTAL DATA

Supplemental Figure 1. Highly variable expression of MMRN1 in pediatric AML patient specimens. Initial discovery studies using microarray data from a set of 211 diagnostic specimens from participants of COG trials (including 58 from AAML03P1 [52 of which were also available for qRT-PCR analyses] and 130 from AAML0531 [122 of which were also available for qRT-PCR analyses]) and 4 normal bone marrow (NBM) specimens. The MLL subset of AMLs was used to arbitrarily define a cut-off of >9.0 (indicated with dashed line) for “high” (n=35, top 16.6%) and ≤9.0 for “low” (n=176, bottom 83.4%) MMRN1 expression, respectively.

Supplemental Figure 2. Clinical outcome in patients with high and low MMRN1 expression. Estimates of the probability of (A) OS and (B) EFS in patients with high (n=35) vs. low (n=176) MMRN1 expression.

Supplemental Figure 3. Relationship between bone marrow blast percentage and MMRN1 expression in the AAML0531 patient cohort. Spearman correlation: p=0.731.

SUPPLEMENTAL TABLE 1. Comparison of Baseline Characteristics of Patients with Low (<0.5) vs. High (≥0.5) MMRN1 Expression in AAML03P1

Patient Characteristics

Relative MMRN1 Expression

P-value

Low (<0.5)

n = 138

High (≥0.5)

n = 45

Median Age, years (range)

10.9 (0.1-20.8)

6.1 (0.2-18.0)

0.020

Male Sex, n (%)

86 (62%)

24 (53%)

0.285

WBC (x103/µL) - median (range)

37.8 (1.8 - 495)

23.7 (2.1 - 409)

0.827

Median bone marrow blasts, %

71 (5-100)

72 (2-100)

0.572

Platelet count (x103/µL) - median (range)

45.5 (4 - 578)

64 (10 - 520)

0.028

Hemoglobin (g/dL) - median (range)

8.6 (3.3 - 15.6)

8.3 (4.3 - 14.0)

0.717

Cytogenetics, n (%)

Normal

30 (24%)

6 (14%)

0.151

t(8;21)(q22;q22)

22 (18%)

0 (0%)

0.003

inv(16)/t(16;16)(p13.1;q22)

24 (20%)

4 (9%)

0.124

t(9;11)(p22;q23) or other abn 11q23

19 (15%)

12 (28%)

0.071

t(6;9)(p23;q34)

5 (4%)

1 (2%)

1.000

Monosomy 7

0 (0%)

5 (12%)

<0.001

Del7q

2 (2%)

2 (5%)

0.276

-5/5q-

0 (0%)

2 (5%)

0.066

Trisomy 8

9 (7%)

4 (9%)

0.743

Other

12 (10%)

7 (16%)

0.270

Unknown

15

2

Risk Group, n (%)

Standard

46 (36%)

30 (70%)

<0.001

Low

66 (52%)

4 (9%)

<0.001

High

15 (12%)

9 (21%)

0.138

Unknown

11

2

Molecular alterations, %

FLT3/ITD

15%

5%

0.075

NPM1 mutation

10%

0%

0.040

CEBPA mutation

9%

0%

0.040

WT1 mutation

14%

7%

0.249

Hepatomegaly, n (%)

26%

38%

0.149

Splenomegaly, n (%)

32%

32%

0.971

Extramedullary disease, n (%)

13%

7%

0.244

CNS disease, n (%)

7%

0%

0.063

Chloroma, n (%)

7%

7%

1.000

SUPPLEMENTAL TABLE 2. Univariate and Multivariate Regression Models of OS, EFS, and RR for the AAML0531 Study, with Outcome Censored at Time of HCT

OS

EFS

RR

n

HR

95% CI

P-value

n

HR

95% CI

P-value

n

HR

95% CI

P-value

Univariate Model

MMRN1 Expression

<0.5

590

1 (Ref)

590

1 (Ref)

449

1 (Ref)

≥0.5

160

2.197

1.65-2.93

<0.001

160

1.864

1.46-2.38

<0.001

106

2.088

1.52-2.87

<0.001

Disease Risk*

Standard-risk

349

1 (Ref)

349

1 (Ref)

251

1 (Ref)

Low-risk

283

0.326

0.23-0.46

<0.001

283

0.428

0.33-0.55

<0.001

233

0.389

0.28-0.53

<0.001

High-Risk

98

1.334

0.91-1.95

0.137

98

1.694

1.23-2.33

0.001

56

1.262

0.75-2.11

0.379

BM Blast % (per 20%)

697

1.108

0.98-1.25

0.097

697

1.244

1.13-1.38

<0.001

519

1.262

1.10-1.44

0.001

Platelet Count (per 50x103/µL)

748

1.052

0.99-1.11

0.083

748

1.035

0.99-1.09

0.164

553

1.038

0.97-1.11

0.262

Multivariate Model**

MMRN1 Expression

<0.5

538

1 (Ref)

538

1 (Ref)

415

1 (Ref)

≥0.5

144

1.554

1.12-2.15

0.008

144

1.353

1.03-1.78

0.031

93

1.442

1.01-2.06

0.044

Disease Risk*

 

Standard-risk

323

1 (Ref)

323

1 (Ref)

232

1 (Ref)

Low-risk

267

0.282

0.19-0.42

<0.001

267

0.411

0.30-0.56

<0.001

223

0.428

0.29-0.63

<0.001

High-Risk

92

1.033

0.68-1.56

0.875

92

1.412

1.00-1.99

0.048

53

1.073

0.62-1.87

0.802

BM Blast % (per 20%)

682

0.984

0.87-1.11

0.793

682

1.152

1.04-1.28

0.007

508

1.208

1.05-1.38

0.006

*See methods section for definition of cytogenetic/molecular disease risk **Models were also adjusted for treatment arm and age.

SUPPLEMENTAL TABLE 3. Relative MMRN1 expression in paired samples of CD34+/CD33- and CD34+/CD33+ cells

 

Cell Population

AML Specimen

 

#1

#2

#3

#4

#5

#6

#7

#8

#9

#10

MMRN1/GUSB Expression

CD34+/CD33-

1.79

0.03

0.09

4.82

0.01

1.64

0.33

1.35

0.06

0.32

CD34+/CD33+

1.14

0.07

0.06

1.44

0.02

0.48

0.13

0.73

0.11

0.15

FACS was used to separate less mature CD34+/CD33- and more mature CD34+/CD33+ cells. Equal amounts of total RNA were then used to quantify MMRN1 and GUSB transcript levels by qRT-PCR.

MM

RN

1 ex

pres

sion

(log

bas

e 2)

11.2 10.4

9.7 8.9 8.1 7.4 6.6 5.8 5.0 4.3 3.5

M

M

R

N

1

e

x

p

r

e

s

s

i

o

n

(

l

o

g

b

a

s

e

2

)

11.2

10.4

9.7

8.9

8.1

7.4

6.6

5.8

5.0

4.3

3.5

0 2 4 6 8 100

25

50

75

100

Years from Study Entry

Ove

rall

Surv

ival

P=0.0040

0 2 4 6 8 100

25

50

75

100

Years from Study Entry

Even

t-fre

e Su

rviv

al

MMRN1 LowMMRN1 High

P=0.0049

A B

0246810

0

25

50

75

100

Years from Study Entry

O

v

e

r

a

l

l

S

u

r

v

i

v

a

l

P=0.0040

0246810

0

25

50

75

100

Years from Study Entry

E

v

e

n

t

-

f

r

e

e

S

u

r

v

i

v

a

l

MMRN1 Low

MMRN1 High

P=0.0049

A

B