Key Points

  • NPM1 mutations in AML arise from replication errors primed by illegitimate TdT activity.

  • The involvement of TdT in both NPM1 and FLT3-ITD mutagenesis suggests a significant proportion of AML is a by-product of adaptive immunity.

Abstract

Nucleophosmin (NPM1) is the most commonly mutated gene in acute myeloid leukemia (AML). AML with mutated NPM1 is recognized as a separate entity in the World Health Organization 2016 classification and carries a relatively favorable prognosis. NPM1 mutations are predominantly 4-bp duplications or insertions in the terminal exon that arise through an unknown mechanism. Here we analyze 2430 NPM1 mutations from 2329 adult and 101 pediatric patients to address their origin. We show that NPM1 mutations display the hallmarks of replication slippage, but lack suitable germline microhomology available for priming. Insertion mutations display G/C-rich N-nucleotide tracts, with a significant bias toward polypurine and polypyrimidine stacking (P < .001). These features suggest terminal deoxynucleotidyl transferase (TdT) primes replication slippage through N-nucleotide addition, with longer syntheses manifesting as N-regions. The recurrent type A, type D, and type B mutations require 1, 2, and 3 N-nucleotide extensions of T, CC, and CAT, respectively, with the last nucleotide used as occult microhomology. This TdT-mutator model successfully predicts the relative incidence of the 256 potential 4-bp insertion/duplication mutations at position c.863_864 over 4 orders of magnitude (ρ = 0.484, P < .0001). Children have a different NPM1 mutation spectrum to adults, including a shift away from type A mutations and toward longer N-regions, consistent with higher TdT activity in pediatric myeloid stem cells. These findings complement our FLT3-ITD data, suggesting illegitimate TdT activity contributes to around one-half of AMLs. AML may therefore reflect the price for adaptive immunity.

REFERENCES

REFERENCES
1.
Grimwade
D
,
Ivey
A
,
Huntly
BJP
.
Molecular landscape of acute myeloid leukemia in younger adults and its clinical relevance
.
Blood
.
2016
;
127
(
1
):
29
-
41
.
2.
Falini
B
,
Mecucci
C
,
Tiacci
E
, et al;
GIMEMA Acute Leukemia Working Party
.
Cytoplasmic nucleophosmin in acute myelogenous leukemia with a normal karyotype
.
N Engl J Med
.
2005
;
352
(
3
):
254
-
266
.
3.
Arber
DA
,
Orazi
A
,
Hasserjian
R
, et al
.
The 2016 revision to the World Health Organization classification of myeloid neoplasms and acute leukemia
.
Blood
.
2016
;
127
(
20
):
2391
-
2405
.
4.
Haferlach
C
,
Mecucci
C
,
Schnittger
S
, et al
.
AML with mutated NPM1 carrying a normal or aberrant karyotype show overlapping biologic, pathologic, immunophenotypic, and prognostic features
.
Blood
.
2009
;
114
(
14
):
3024
-
3032
.
5.
Rau
R
,
Brown
P
.
Nucleophosmin (NPM1) mutations in adult and childhood acute myeloid leukaemia: towards definition of a new leukaemia entity
.
Hematol Oncol
.
2009
;
27
(
4
):
171
-
181
.
6.
Falini
B
,
Nicoletti
I
,
Martelli
MF
,
Mecucci
C
.
Acute myeloid leukemia carrying cytoplasmic/mutated nucleophosmin (NPMc+ AML): biologic and clinical features
.
Blood
.
2007
;
109
(
3
):
874
-
885
.
7.
Alcalay
M
,
Tiacci
E
,
Bergomas
R
, et al
.
Acute myeloid leukemia bearing cytoplasmic nucleophosmin (NPMc+ AML) shows a distinct gene expression profile characterized by up-regulation of genes involved in stem-cell maintenance
.
Blood
.
2005
;
106
(
3
):
899
-
902
.
8.
Mullighan
CG
,
Kennedy
A
,
Zhou
X
, et al
.
Pediatric acute myeloid leukemia with NPM1 mutations is characterized by a gene expression profile with dysregulated HOX gene expression distinct from MLL-rearranged leukemias
.
Leukemia
.
2007
;
21
(
9
):
2000
-
2009
.
9.
Schnittger
S
,
Schoch
C
,
Kern
W
, et al
.
Nucleophosmin gene mutations are predictors of favorable prognosis in acute myelogenous leukemia with a normal karyotype
.
Blood
.
2005
;
106
(
12
):
3733
-
3739
.
10.
Verhaak
RGW
,
Goudswaard
CS
,
van Putten
W
, et al
.
Mutations in nucleophosmin (NPM1) in acute myeloid leukemia (AML): association with other gene abnormalities and previously established gene expression signatures and their favorable prognostic significance
.
Blood
.
2005
;
106
(
12
):
3747
-
3754
.
11.
Nakagawa
M
,
Kameoka
Y
,
Suzuki
R
.
Nucleophosmin in acute myelogenous leukemia
.
N Engl J Med
.
2005
;
352
(
17
):
1819
-
1820
,
author reply 1819-1820
.
12.
Nishimura
Y
,
Ohkubo
T
,
Furuichi
Y
,
Umekawa
H
.
Tryptophans 286 and 288 in the C-terminal region of protein B23.1 are important for its nucleolar localization
.
Biosci Biotechnol Biochem
.
2002
;
66
(
10
):
2239
-
2242
.
13.
Falini
B
,
Bolli
N
,
Shan
J
, et al
.
Both carboxy-terminus NES motif and mutated tryptophan(s) are crucial for aberrant nuclear export of nucleophosmin leukemic mutants in NPMc+ AML
.
Blood
.
2006
;
107
(
11
):
4514
-
4523
.
14.
Brunetti
L
,
Gundry
MC
,
Sorcini
D
, et al
.
Mutant NPM1 maintains the leukemic state through HOX expression
.
Cancer Cell
.
2018
;
34
(
3
):
499
-
512
.
15.
Brown
P
,
McIntyre
E
,
Rau
R
, et al
.
The incidence and clinical significance of nucleophosmin mutations in childhood AML
.
Blood
.
2007
;
110
(
3
):
979
-
985
.
16.
Brown
P
,
Meshinchi
S
,
Levis
M
, et al
.
Pediatric AML primary samples with FLT3/ITD mutations are preferentially killed by FLT3 inhibition
.
Blood
.
2004
;
104
(
6
):
1841
-
1849
.
17.
Thiede
C
,
Creutzig
E
,
Reinhardt
D
,
Ehninger
G
,
Creutzig
U
.
Different types of NPM1 mutations in children and adults: evidence for an effect of patient age on the prevalence of the TCTG-tandem duplication in NPM1-exon 12
.
Leukemia
.
2007
;
21
(
2
):
366
-
367
.
18.
Borrow
J
,
Dyer
SA
,
Akiki
S
,
Griffiths
MJ
.
Terminal deoxynucleotidyl transferase promotes acute myeloid leukemia by priming FLT3-ITD replication slippage
.
Blood
.
2019
;
134
(
25
):
2281
-
2290
.
19.
Dovey
OM
,
Cooper
JL
,
Mupo
A
, et al
.
Molecular synergy underlies the co-occurrence patterns and phenotype of NPM1-mutant acute myeloid leukemia
.
Blood
.
2017
;
130
(
17
):
1911
-
1922
.
20.
Lieber
MR
.
The mechanism of double-strand DNA break repair by the nonhomologous DNA end-joining pathway
.
Annu Rev Biochem
.
2010
;
79
(
1
):
181
-
211
.
21.
Messer
PW
,
Arndt
PF
.
The majority of recent short DNA insertions in the human genome are tandem duplications
.
Mol Biol Evol
.
2007
;
24
(
5
):
1190
-
1197
.
22.
Roth
DB
,
Chang
XB
,
Wilson
JH
.
Comparison of filler DNA at immune, nonimmune, and oncogenic rearrangements suggests multiple mechanisms of formation
.
Mol Cell Biol
.
1989
;
9
(
7
):
3049
-
3057
.
23.
Bangs
LA
,
Sanz
IE
,
Teale
JM
.
Comparison of D, JH, and junctional diversity in the fetal, adult, and aged B cell repertoires
.
J Immunol
.
1991
;
146
(
6
):
1996
-
2004
.
24.
Waanders
E
,
Scheijen
B
,
van der Meer
LT
, et al
.
The origin and nature of tightly clustered BTG1 deletions in precursor B-cell acute lymphoblastic leukemia support a model of multiclonal evolution
.
PLoS Genet
.
2012
;
8
(
2
):
e1002533
.
25.
Gauss
GH
,
Lieber
MR
.
Mechanistic constraints on diversity in human V(D)J recombination
.
Mol Cell Biol
.
1996
;
16
(
1
):
258
-
269
.
26.
Kosugi
S
,
Yanagawa
H
,
Terauchi
R
,
Tabata
S
.
NESmapper: accurate prediction of leucine-rich nuclear export signals using activity-based profiles
.
PLOS Comput Biol
.
2014
;
10
(
9
):
e1003841
.
27.
Marculescu
R
,
Vanura
K
,
Montpellier
B
, et al
.
Recombinase, chromosomal translocations and lymphoid neoplasia: targeting mistakes and repair failures
.
DNA Repair (Amst)
.
2006
;
5
(
9-10
):
1246
-
1258
.
28.
Rebhandl
S
,
Huemer
M
,
Greil
R
,
Geisberger
R
.
AID/APOBEC deaminases and cancer
.
Oncoscience
.
2015
;
2
(
4
):
320
-
333
.
29.
Boeckx
N
,
Willemse
MJ
,
Szczepanski
T
, et al
.
Fusion gene transcripts and Ig/TCR gene rearrangements are complementary but infrequent targets for PCR-based detection of minimal residual disease in acute myeloid leukemia
.
Leukemia
.
2002
;
16
(
3
):
368
-
375
.
30.
Murray
JM
,
O’Neill
JP
,
Messier
T
, et al
.
V(D)J recombinase-mediated processing of coding junctions at cryptic recombination signal sequences in peripheral T cells during human development
.
J Immunol
.
2006
;
177
(
8
):
5393
-
5404
.
31.
Schneider
M
,
Panzer
S
,
Stolz
F
,
Fischer
S
,
Gadner
H
,
Panzer-Grümayer
ER
.
Crosslineage TCR delta rearrangements occur shortly after the DJ joinings of the IgH genes in childhood precursor B ALL and display age-specific characteristics
.
Br J Haematol
.
1997
;
99
(
1
):
115
-
121
.
32.
Champagne
DP
,
Shockett
PE
.
Illegitimate V(D)J recombination-mediated deletions in Notch1 and Bcl11b are not sufficient for extensive clonal expansion and show minimal age or sex bias in frequency or junctional processing
.
Mutat Res
.
2014
;
761
:
34
-
48
.
33.
Creutzig
U
,
van den Heuvel-Eibrink
MM
,
Gibson
B
, et al;
AML Committee of the International BFM Study Group
.
Diagnosis and management of acute myeloid leukemia in children and adolescents: recommendations from an international expert panel
.
Blood
.
2012
;
120
(
16
):
3187
-
3205
.
You do not currently have access to this content.