Key Points

  • Differential spatial expression of the chemokine CXCL4/platelet factor-4 marks the progression of fibrosis.

  • The absence of hematopoietic CXCL4 ameliorates the MPN phenotype and reduces stromal cell activation and BM fibrosis.

Abstract

Primary myelofibrosis (PMF) is a myeloproliferative neoplasm (MPN) that leads to progressive bone marrow (BM) fibrosis. Although the cellular mutations involved in the pathogenesis of PMF have been extensively investigated, the sequential events that drive stromal activation and fibrosis by hematopoietic–stromal cross-talk remain elusive. Using an unbiased approach and validation in patients with MPN, we determined that the differential spatial expression of the chemokine CXCL4/platelet factor-4 marks the progression of fibrosis. We show that the absence of hematopoietic CXCL4 ameliorates the MPN phenotype, reduces stromal cell activation and BM fibrosis, and decreases the activation of profibrotic pathways in megakaryocytes, inflammation in fibrosis-driving cells, and JAK/STAT activation in both megakaryocytes and stromal cells in 3 murine PMF models. Our data indicate that higher CXCL4 expression in MPN has profibrotic effects and is a mediator of the characteristic inflammation. Therefore, targeting CXCL4 might be a promising strategy to reduce inflammation in PMF.

REFERENCES

REFERENCES
1.
Grinfeld
J
,
Nangalia
J
,
Baxter
EJ
, et al
.
Classification and personalized prognosis in myeloproliferative neoplasms
.
N Engl J Med
.
2018
;
379
(
15
):
1416
-
1430
.
2.
Baxter
EJ
,
Scott
LM
,
Campbell
PJ
, et al;
Cancer Genome Project
.
Acquired mutation of the tyrosine kinase JAK2 in human myeloproliferative disorders
.
Lancet
.
2005
;
365
(
9464
):
1054
-
1061
.
3.
James
C
,
Ugo
V
,
Le Couédic
JP
, et al
.
A unique clonal JAK2 mutation leading to constitutive signalling causes polycythaemia vera
.
Nature
.
2005
;
434
(
7037
):
1144
-
1148
.
4.
Kralovics
R
,
Passamonti
F
,
Buser
AS
, et al
.
A gain-of-function mutation of JAK2 in myeloproliferative disorders
.
N Engl J Med
.
2005
;
352
(
17
):
1779
-
1790
.
5.
Levine
RL
,
Pardanani
A
,
Tefferi
A
,
Gilliland
DG
.
Role of JAK2 in the pathogenesis and therapy of myeloproliferative disorders
.
Nat Rev Cancer
.
2007
;
7
(
9
):
673
-
683
.
6.
Thorsten
K
,
Gisslinger
H
,
Harutyunyan
AS
, et al
.
Frequent mutations in the calreticulin gene CALR in myeloproliferative neoplasms
.
Blood
.
2013
;
122
(
21
):
LBA
-
1
.
7.
Schneider
RK
,
Mullally
A
,
Dugourd
A
, et al
.
Gli1+ mesenchymal stromal cells are a key driver of bone marrow fibrosis and an important cellular therapeutic target
.
Cell Stem Cell
.
2017
;
20
(
6
):
785
-
800.e788
.
8.
Decker
M
,
Martinez-Morentin
L
,
Wang
G
, et al
.
Leptin-receptor-expressing bone marrow stromal cells are myofibroblasts in primary myelofibrosis
.
Nat Cell Biol
.
2017
;
19
(
6
):
677
-
688
.
9.
Levine
SP
,
Wohl
H
.
Human platelet factor 4: purification and characterization by affinity chromatography. Purification of human platelet factor 4
.
J Biol Chem
.
1976
;
251
(
2
):
324
-
328
.
10.
Schaffner
A
,
Rhyn
P
,
Schoedon
G
,
Schaer
DJ
.
Regulated expression of platelet factor 4 in human monocytes—role of PARs as a quantitatively important monocyte activation pathway
.
J Leukoc Biol
.
2005
;
78
(
1
):
202
-
209
.
11.
Vandercappellen
J
,
Van Damme
J
,
Struyf
S
.
The role of the CXC chemokines platelet factor-4 (CXCL4/PF-4) and its variant (CXCL4L1/PF-4var) in inflammation, angiogenesis and cancer
.
Cytokine Growth Factor Rev
.
2011
;
22
(
1
):
1
-
18
.
12.
Affandi
AJ
,
Silva-Cardoso
SC
,
Garcia
S
, et al
.
CXCL4 is a novel inducer of human Th17 cells and correlates with IL-17 and IL-22 in psoriatic arthritis
.
Eur J Immunol
.
2018
;
48
(
3
):
522
-
531
.
13.
Vrij
AA
,
Rijken
J
,
Van Wersch
JW
,
Stockbrügger
RW
.
Platelet factor 4 and beta-thromboglobulin in inflammatory bowel disease and giant cell arteritis
.
Eur J Clin Invest
.
2000
;
30
(
3
):
188
-
194
.
14.
Pitsilos
S
,
Hunt
J
,
Mohler
ER
, et al
.
Platelet factor 4 localization in carotid atherosclerotic plaques: correlation with clinical parameters
.
Thromb Haemost
.
2003
;
90
(
6
):
1112
-
1120
.
15.
Aivado
M
,
Spentzos
D
,
Germing
U
, et al
.
Serum proteome profiling detects myelodysplastic syndromes and identifies CXC chemokine ligands 4 and 7 as markers for advanced disease
.
Proc Natl Acad Sci U S A
.
2007
;
104
(
4
):
1307
-
1312
.
16.
Kasper
B
,
Petersen
F
.
Molecular pathways of platelet factor 4/CXCL4 signaling
.
Eur J Cell Biol
.
2011
;
90
(
6-7
):
521
-
526
.
17.
Schubert
M
,
Klinger
B
,
Klünemann
M
, et al
.
Perturbation-response genes reveal signaling footprints in cancer gene expression
.
Nat Commun
.
2018
;
9
(
1
):
20
.
18.
Holland
CH
,
Szalai
B
,
Saez-Rodriguez
J
.
Transfer of regulatory knowledge from human to mouse for functional genomics analysis
.
Biochim Biophys Acta Gene Regul Mech
.
2020
;
1863
(
6
):
194431
.
19.
Darby
IA
,
Hewitson
TD
.
Hypoxia in tissue repair and fibrosis
.
Cell Tissue Res
.
2016
;
365
(
3
):
553
-
562
.
20.
Naba
A
,
Clauser
KR
,
Hoersch
S
,
Liu
H
,
Carr
SA
,
Hynes
RO
.
The matrisome: in silico definition and in vivo characterization by proteomics of normal and tumor extracellular matrices
.
Mol Cell Proteomics
.
2012
;
11
(
4
):
014647
.
21.
Väremo
L
,
Nielsen
J
,
Nookaew
I
.
Enriching the gene set analysis of genome-wide data by incorporating directionality of gene expression and combining statistical hypotheses and methods
.
Nucleic Acids Res
.
2013
;
41
(
8
):
4378
-
4391
.
22.
Sahin
H
,
Wasmuth
HE
.
Chemokines in tissue fibrosis
.
Biochim Biophys Acta
.
2013
;
1832
(
7
):
1041
-
1048
.
23.
Bruns
I
,
Lucas
D
,
Pinho
S
, et al
.
Megakaryocytes regulate hematopoietic stem cell quiescence through CXCL4 secretion
.
Nat Med
.
2014
;
20
(
11
):
1315
-
1320
.
24.
Tefferi
Ayalew
,
Nicolosi
Maura
,
Mudireddy
Mythri
, et al
.
Driver mutations and prognosis in primary myelofibrosis: Mayo-Careggi MPN alliance study of 1,095 patients
.
Am. J. Hematol
.
2018
;
93
(
3
):
348
-
355
.
25.
Garcia-Alonso
L
,
Holland
CH
,
Ibrahim
MM
,
Turei
D
,
Saez-Rodriguez
J
.
Benchmark and integration of resources for the estimation of human transcription factor activities
.
Genome Res
.
2019
;
29
(
8
):
1363
-
1375
.
26.
Rampal
R
,
Al-Shahrour
F
,
Abdel-Wahab
O
, et al
.
Integrated genomic analysis illustrates the central role of JAK-STAT pathway activation in myeloproliferative neoplasm pathogenesis
.
Blood
.
2014
;
123
(
22
):
e123
-
e133
.
27.
Chen
E
,
Beer
PA
,
Godfrey
AL
, et al
.
Distinct clinical phenotypes associated with JAK2V617F reflect differential STAT1 signaling
.
Cancer Cell
.
2010
;
18
(
5
):
524
-
535
.
28.
Čokić
VP
,
Mitrović-Ajtić
O
,
Beleslin-Čokić
BB
, et al
.
Proinflammatory cytokine IL-6 and JAK-STAT signaling pathway in myeloproliferative neoplasms
.
Mediators Inflamm
.
2015
;
2015
:
453020
.
29.
Sollazzo
D
,
Forte
D
,
Polverelli
N
, et al
.
Circulating calreticulin is increased in myelofibrosis: correlation with interleukin-6 plasma levels, bone marrow fibrosis, and splenomegaly
.
Mediators Inflamm
.
2016
;
2016
:
5860657
.
30.
Alavi
MV
,
Mao
M
,
Pawlikowski
BT
, et al
.
Col4a1 mutations cause progressive retinal neovascular defects and retinopathy
.
Sci Rep
.
2016
;
6
(
1
):
18602
.
31.
Mylotte
LA
,
Duffy
AM
,
Murphy
M
, et al
.
Metabolic flexibility permits mesenchymal stem cell survival in an ischemic environment
.
Stem Cells
.
2008
;
26
(
5
):
1325
-
1336
.
32.
Hu
X
,
Yu
SP
,
Fraser
JL
, et al
.
Transplantation of hypoxia-preconditioned mesenchymal stem cells improves infarcted heart function via enhanced survival of implanted cells and angiogenesis
.
J Thorac Cardiovasc Surg
.
2008
;
135
(
4
):
799
-
808
.
33.
Grayson
WL
,
Zhao
F
,
Bunnell
B
,
Ma
T
.
Hypoxia enhances proliferation and tissue formation of human mesenchymal stem cells
.
Biochem Biophys Res Commun
.
2007
;
358
(
3
):
948
-
953
.
34.
van Bon
L
,
Affandi
AJ
,
Broen
J
, et al
.
Proteome-wide analysis and CXCL4 as a biomarker in systemic sclerosis
.
N Engl J Med
.
2014
;
370
(
5
):
433
-
443
.
35.
Zaldivar
MM
,
Pauels
K
,
von Hundelshausen
P
, et al
.
CXC chemokine ligand 4 (Cxcl4) is a platelet-derived mediator of experimental liver fibrosis
.
Hepatology
.
2010
;
51
(
4
):
1345
-
1353
.
36.
Kiu
H
,
Nicholson
SE
.
Biology and significance of the JAK/STAT signalling pathways
.
Growth Factors
.
2012
;
30
(
2
):
88
-
106
.
37.
Kleppe
M
,
Kwak
M
,
Koppikar
P
, et al
.
JAK-STAT pathway activation in malignant and nonmalignant cells contributes to MPN pathogenesis and therapeutic response
.
Cancer Discov
.
2015
;
5
(
3
):
316
-
331
.
38.
Tefferi
A
,
Vaidya
R
,
Caramazza
D
,
Finke
C
,
Lasho
T
,
Pardanani
A
.
Circulating interleukin (IL)-8, IL-2R, IL-12, and IL-15 levels are independently prognostic in primary myelofibrosis: a comprehensive cytokine profiling study
.
J Clin Oncol
.
2011
;
29
(
10
):
1356
-
1363
.
39.
Heinrich
PC
,
Behrmann
I
,
Haan
S
,
Hermanns
HM
,
Müller-Newen
G
,
Schaper
F
.
Principles of interleukin (IL)-6-type cytokine signalling and its regulation
.
Biochem J
.
2003
;
374
(
Pt 1
):
1
-
20
.
40.
Manshouri
T
,
Estrov
Z
,
Quintás-Cardama
A
, et al
.
Bone marrow stroma-secreted cytokines protect JAK2(V617F)-mutated cells from the effects of a JAK2 inhibitor
.
Cancer Res
.
2011
;
71
(
11
):
3831
-
3840
.
41.
Yue
L
,
Bartenstein
M
,
Zhao
W
, et al
.
Efficacy of ALK5 inhibition in myelofibrosis
.
JCI Insight
.
2017
;
2
(
7
):
e90932
.
42.
Ballen
KK
,
Shrestha
S
,
Sobocinski
KA
, et al
.
Outcome of transplantation for myelofibrosis
.
Biol Blood Marrow Transplant
.
2010
;
16
(
3
):
358
-
367
.
43.
Cervantes
F
,
Vannucchi
AM
,
Kiladjian
JJ
, et al;
COMFORT-II investigators
.
Three-year efficacy, safety, and survival findings from COMFORT-II, a phase 3 study comparing ruxolitinib with best available therapy for myelofibrosis [published correction appears in Blood. 2016;128(25):3013]
.
Blood
.
2013
;
122
(
25
):
4047
-
4053
.
44.
Tefferi
A
.
Challenges facing JAK inhibitor therapy for myeloproliferative neoplasms
.
N Engl J Med
.
2012
;
366
(
9
):
844
-
846
.
45.
Verstovsek
S
,
Kantarjian
H
,
Mesa
RA
, et al
.
Safety and efficacy of INCB018424, a JAK1 and JAK2 inhibitor, in myelofibrosis
.
N Engl J Med
.
2010
;
363
(
12
):
1117
-
1127
.
46.
Ceglia
I
,
Dueck
AC
,
Masiello
F
, et al
.
Preclinical rationale for TGF-β inhibition as a therapeutic target for the treatment of myelofibrosis
.
Exp Hematol
.
2016
;
44
(
12
):
1138
-
1155.e4
.
47.
Kampan
NC
,
Xiang
SD
,
McNally
OM
,
Stephens
AN
,
Quinn
MA
,
Plebanski
M
.
Immunotherapeutic interleukin-6 or interleukin-6 receptor blockade in cancer: challenges and opportunities
.
Curr Med Chem
.
2018
;
25
(
36
):
4785
-
4806
.
48.
Huselton
E
,
Cashen
AF
,
Jacoby
M
,
Pusic
I
,
Romee
R
,
Uy
GL
.
CX-01, an inhibitor of CXCL12/CXCR4 axis and of platelet factor 4 (PF4), with azacitidine (AZA) in patients with hypomethylating agent (HMA) refractory AML and MDS
.
J Clin Oncol
.
2018
;
36
(
15
):
7027
.
49.
Kovacsovics
TJ
,
Mims
A
,
Salama
ME
, et al
.
Combination of the low anticoagulant heparin CX-01 with chemotherapy for the treatment of acute myeloid leukemia
.
Blood Adv
.
2018
;
2
(
4
):
381
-
389
.
50.
Garbers
C
,
Heink
S
,
Korn
T
,
Rose-John
S
.
Interleukin-6: designing specific therapeutics for a complex cytokine
.
Nat Rev Drug Discov
.
2018
;
17
(
6
):
395
-
412
.
You do not currently have access to this content.