We developed a real-time copy number polymerase chain reaction assay for deletions on chromosome 20q (del20q), screened peripheral blood granulocytes from 664 patients with myeloproliferative disorders, and identified 19 patients with del20q (2.9%), of which 14 (74%) were also positive for JAK2-V617F. To examine the temporal relationship between the occurrence of del20q and JAK2-V617F, we performed colony assays in methylcellulose, picked individual burst-forming units–erythroid (BFU-E) and colony-forming units–granulocyte (CFU-G) colonies, and genotyped each colony individually for del20q and JAK2-V617F. In 2 of 9 patients, we found that some colonies with del20q carried only wild-type JAK2, whereas other del20q colonies were JAK2-V617F positive, indicating that del20q occurred before the acquisition of JAK2-V617F. However, in colonies from 3 of 9 patients, we observed the opposite order of events. The lack of a strict temporal order of occurrence makes it doubtful that del20q represents a predisposing event for JAK2-V617F. In 2 patients with JAK2-V617F and 1 patient with MPL-W515L, microsatellite analysis revealed that del20q affected chromosomes of different parental origin and/or 9pLOH occurred at least twice. The fact that rare somatic events, such as del20q or 9pLOH, occurred more than once in subclones from the same patients suggests that the myeloproliferative disorder clone carries a predisposition to acquiring such genetic alterations.

Deletions of the long arm of chromosome 20 (del20q) have been observed frequently in patients with myeloproliferative disorders (MPDs), myelodysplastic syndromes (MDSs), and acute myeloid leukemia (AML).1,,4  A common deleted region (CDR) of 2.7 megabases (Mb) was mapped for patients with MPD, and a 2.6-Mb CDR was defined for MDS/AML with an overlap of 1.7 Mb between the 2 CDRs.5,,,,10  However, to date, it was not possible to identify a gene mutation within the CDR that is functionally linked to the expansion of the del20q clone.11  In MPD patients, del20q was found in the bone marrow from approximately 1% of patients with essential thrombocythemia (ET), up to 9% of polycythemia vera (PV) and up to 12% of primary myelofibrosis (PMF).10,12  The del20q occurred preferentially together with the JAK2-V617F mutation in 28 of 29 MPD patients studied.13  We previously analyzed 2 such patients with del20q and JAK2-V617F and found that the size of the del20q clone by far exceeded the size of the clone carrying JAK2-V617F,14  suggesting that del20q preceded the acquisition of the JAK2-V617F mutation. Here we expanded these studies by screening for del20q in 664 patients with MPD and examined the temporal order of the acquisition of JAK2-V617F and del20q in patients carrying both mutations by analyzing single colonies grown in methylcellulose

Patients

The collection of blood samples was performed at the study centers in Basel, Switzerland, Vienna, Austria, and Dijon and Nantes, France and was approved by the local Ethics Committees (Ethik Kommission Beider Basel, the Ethik Kommission der Universität Wien und des Allgemeinen Krankenhauses der Stadt Wien-AK, and the institutional boards of Dijon and Nantes). Written informed consent was obtained from all patients in accordance with the Declaration of Helsinki. The diagnosis of MPD was established according to the criteria of the World Health Organization.4 

Cells and DNA analysis

Purification of granulocytes and peripheral mononuclear cells (PBMCs) and extraction of DNA were performed as described.15,16  The granulocyte DNA was assayed using a real-time polymerase chain reaction (PCR)–based copy number assay as previously published.16  The following primers were used to detect the copy number of 2 different exon of L3MBTL: 1736-GATCCCAATCAGGACCCCC, 1737-CGCCTGGCACTGACAGGT for exon 2 and 1995-CATGAAGCTGGAGGCTGTTG, 1996-GCCACGCAGACAAGGGAC for exon 9. Primers for LOC221154 (1283-CCATGGACGACGGGTTTCT, 1284-TGTACAGGACGTAGGAGGGTGA) were used as a diploid reference. Samples that showed a decrease in copy number in both exons were validated using primers in the neighboring genes PTPRT(1740-TGCCCCGGAACCATGATA, 1741-GGTCCAGAGGCAGCACGT) and TOX2 (1949-CCTGCCTACTCCTATCAGGCC, 1950-GTTGGACACCATGATGGCTG). The colony assays were performed using PBMCs from patients as published.17  Methylcellulose-based media (no. 4431) containing erythropoietin from StemCell Technologies (Vancouver, BC) was used to grow burst-forming units–erythroid (BFU-E) and colony-forming units–granulocyte (CFU-G). To ensure that plates with an optimal density were obtained so that colonies can be picked without contamination by cells from neighboring colonies, PBMCs were plated at 3 different concentrations (50 000, 100 000, and 200 000 per mL).

Comparative genomic hybridization

The custom oligo comparative genomic hybridization (CGH) array was designed using the eArray platform from Agilent Technologies (Palo Alto, CA). The chips were processed following the instructions of the supplier. Either a pool of male whole blood DNAs (Promega, Madison, WI) or T-cell DNAs from the respective patients were used as reference DNA. The data were analyzed using Capweb software (http://bioinfo-out.curie.fr/CAPweb/).18 

Purified peripheral blood granulocyte DNAs of 664 MPD patients (320 ET, 262 PV, 82 PMF) were screened for del20q by monitoring the copy number of L3MBTL, a gene located in the common deleted region on chromosome 20q. Two real-time PCR assays with primers placed in exons 2 and 9 of L3MBTL were used in parallel (Figure 1A). Patients with decreased gene copy numbers in both assays were further examined with real-time PCR assays for PTPRT and TOX2, 2 genes flanking L3MBTL (Figure 1B). The del20q was confirmed in 19 of 664 (2.9%) MPD patients, or specifically in 4 of 320 (1.3%) ET, 7 of 262 (2.7%) PV, and 8 of 82 (9.8%) PMF. The JAK2-V617F mutation was present in 14 of 19 (74%) patients with del20q, whereas the MPL-W515L was present in 1 patient without JAK2-V617F. No mutations in JAK2 exon 12 were found among the patients with del20q (Table 1). Cytogenetic analysis was available for 8 of the 19 patients with del20q and confirmed the results obtained by real-time PCR in all 8 cases. To map the breakpoints of the individual deletions, we performed custom high-density oligonucleotide CGH arrays on 8 del20q patients from whom DNA of sufficient quality was available. A CDR of 8.98 Mb (35.95-44.93 Mb) containing 93 genes was defined (Figure 2).

Figure 1

Screening for del20q by real-time PCR. (A) The copy number of the L3MBTL gene was determined in DNA from purified peripheral blood granulocytes using 2 SYBR real-time PCR assays with primer pairs located in L3MBTL exon 2 (quadrangles) and exon 9 (circles). Samples with gene copy values less than a threshold of 1.8 in both assays are marked in red and identified by unique patient numbers. (B) Patients with decreased L3MBTL gene copy number were further examined with real-time PCR assays for the neighboring genes PTPRT and TOX2. The decreased copy number was confirmed in all cases. nd indicates not determined.

Figure 1

Screening for del20q by real-time PCR. (A) The copy number of the L3MBTL gene was determined in DNA from purified peripheral blood granulocytes using 2 SYBR real-time PCR assays with primer pairs located in L3MBTL exon 2 (quadrangles) and exon 9 (circles). Samples with gene copy values less than a threshold of 1.8 in both assays are marked in red and identified by unique patient numbers. (B) Patients with decreased L3MBTL gene copy number were further examined with real-time PCR assays for the neighboring genes PTPRT and TOX2. The decreased copy number was confirmed in all cases. nd indicates not determined.

Close modal
Figure 2

CGH. Granulocyte DNA from patients and a reference DNA were used to perform custom high-density oligonucleotide CGH arrays. The graphs show the log ratios for all probes located on chromosome 20q. Green represents regions deleted in the patient's DNA. The boundaries are marked by a clear drop in the ratio between the patient's DNA and the reference. Patient Vi384 delineates the centromeric and patient Na293 the telomeric border of the CDR, which spans 9 Mb and includes 93 genes.

Figure 2

CGH. Granulocyte DNA from patients and a reference DNA were used to perform custom high-density oligonucleotide CGH arrays. The graphs show the log ratios for all probes located on chromosome 20q. Green represents regions deleted in the patient's DNA. The boundaries are marked by a clear drop in the ratio between the patient's DNA and the reference. Patient Vi384 delineates the centromeric and patient Na293 the telomeric border of the CDR, which spans 9 Mb and includes 93 genes.

Close modal

To determine the temporal relationship between the occurrence of del20q and JAK2-V617F, we performed colony assays in methylcellulose, picked single BFU-E and CFU-G colonies grown in the presence of erythropoietin, and genotyped each colony individually for del20q, JAK2-V617F, or in patient Vi346 for MPL-W515L (Figure 3). We observed 3 different patterns: First, in patients p187 and Na293, we found that some del20q-positive colonies carried the wild-type JAK2, whereas other del20q-positive colonies were positive for JAK2-V617F, indicating that the del20q clone is larger than the JAK2-V617F clone (Figure 3A). Because all colonies positive for JAK2-V617F also displayed del20q, we infer that del20q occurred before the JAK2 mutation. Second, in 3 patients, we observed the reverse order, with JAK2-V617F preceding del20q, as illustrated by the presence of JAK2-V617F–positive colonies with and without del20q (Figure 3B). Patient Vi233 appears to have acquired del20q in a cell heterozygous for JAK2-V617F, whereas in patients p048 and p103 the transition to del20q occurred in a cell homozygous for JAK2-V617F. Third, in patients p007, Vi102, and Vi346, we found a more complex pattern (Figure 3C). In the first sample of p007 from February 2005, only a few CFU-G heterozygous for JAK2-V617F carried del20q, whereas colonies homozygous for JAK2-V617F already existed. Two years later, del20q was also present in a subset of colonies homozygous for JAK2-V617F. This pattern could be explained either by postulating 2 independent del20q events or 2 independent 9pLOH events. The del20q-positive BFU-E colonies in patient Vi102 were wild-type for JAK2, and the JAK2-V617F–positive colonies were all negative for del20q, which is compatible with originating from 2 independent clones. However, the finding of a CFU-G colony positive for both JAK2-V617F and del20q suggests that either del20q or JAK2-V617F occurred twice independently. Patient Vi346 was negative for JAK2-V617F but carried the MPL-W515L mutation. The del20q was only detectable in CFU-G colonies, and all of these colonies were also heterozygous for MPL-W515L. Some colonies were homozygous for MPL-W515L but negative for del20q.

Figure 3

Single colony assays for del20q and JAK2-V617F. PBMCs were grown in methylcellulose in the presence of erythropoietin. Single erythroid colonies (BFU-E) and granulocytic colonies (CFU-G) were picked and analyzed individually for the presence of del20q and JAK2-V617F by microsatellite PCR and allele-specific PCR, respectively. Each colony is represented by a dot that is placed into one of 6 quadrangles representing the 6 possible genotypes: wild-type (wt), heterozygous (het), and homozygous (hom) for JAK2-V617F or MPL-W515L on the vertical axis, and absence (open quadrangles) or presence of del20q (gray quadrangles) on the horizontal axis. Results for BFU-Es are shown in the upper part and CFU-Gs in the lower part of the panels. The unique patient numbers, the diagnoses (PMF, ET, PV, sMF indicates secondary myelofibrosis), the allelic ratio of JAK2-V617F or MPL-W515L in purified granulocytes (%T), and the date of the sample drawing are shown above the corresponding boxes. (A) This group of patients has acquired the del20q before JAK2-V617F, as demonstrated by the presence of JAK2 wt colonies carrying del20q. For both patients shown, 2 sequential samples were analyzed. (B) In this group of patients, del20q was acquired in a cell already carrying JAK2-V617F, as indicated by the presence of JAK2-V617F-positive colonies with or without del20q. (C) Patients with a complex pattern of del20q acquisition that does not comply with a linear temporal order of events. In patient Vi346, the del20q coexists with MPL-W515L. (D) A patient with del20q that is negative for the JAK2-V617F mutation.

Figure 3

Single colony assays for del20q and JAK2-V617F. PBMCs were grown in methylcellulose in the presence of erythropoietin. Single erythroid colonies (BFU-E) and granulocytic colonies (CFU-G) were picked and analyzed individually for the presence of del20q and JAK2-V617F by microsatellite PCR and allele-specific PCR, respectively. Each colony is represented by a dot that is placed into one of 6 quadrangles representing the 6 possible genotypes: wild-type (wt), heterozygous (het), and homozygous (hom) for JAK2-V617F or MPL-W515L on the vertical axis, and absence (open quadrangles) or presence of del20q (gray quadrangles) on the horizontal axis. Results for BFU-Es are shown in the upper part and CFU-Gs in the lower part of the panels. The unique patient numbers, the diagnoses (PMF, ET, PV, sMF indicates secondary myelofibrosis), the allelic ratio of JAK2-V617F or MPL-W515L in purified granulocytes (%T), and the date of the sample drawing are shown above the corresponding boxes. (A) This group of patients has acquired the del20q before JAK2-V617F, as demonstrated by the presence of JAK2 wt colonies carrying del20q. For both patients shown, 2 sequential samples were analyzed. (B) In this group of patients, del20q was acquired in a cell already carrying JAK2-V617F, as indicated by the presence of JAK2-V617F-positive colonies with or without del20q. (C) Patients with a complex pattern of del20q acquisition that does not comply with a linear temporal order of events. In patient Vi346, the del20q coexists with MPL-W515L. (D) A patient with del20q that is negative for the JAK2-V617F mutation.

Close modal

In one patient (Na391) with 50% of erythroblasts in the peripheral blood, all colonies were positive for both del20q and JAK2-V617F, which precluded us from determining the order of events (not shown), and one patient (p020) had del20q without a mutation in the JAK2 or MPL gene (Figure 3D). We analyzed p020 at 3 different time points, and we were able to detect an increase in del20q-positive colonies in CFU-Gs in 2007. However, one year later, the percentage of del20q-positive colonies was again comparable with the first time point. An increase in the percentage of del20q-positive colonies was observed in serial samples from Na293 and p007, whereas in p187 the colonies were all del20q-positive and remained so during follow-up, except for one BFU-E and one CFU-G (Figure 3A,C).

To examine the complex pattern in patients p007, Vi102, and Vi346, we performed a more detailed microsatellite analysis for regions on chromosomes 9 and 20 (Figure 4). In p007, the deletion in colony no. 185 affected the chromosome 20q of a different parental origin than in colony no. 6, indicating that 2 independent deletion events must have occurred (Figure 4A left panel). Furthermore, 4 different sizes of the latter del20q haplotype were detected when additional colonies were analyzed. This pattern could arise when a small del20q event was followed by sequential events that increased the size of the deleted region in the same cell, or alternatively, each of the del20q regions could represent a separate de novo deletion event. At present, we cannot distinguish between these possibilities. In addition, colony no. 185 also differed in the size of the 9pLOH region, suggesting the presence of 2 different 9pLOH events (Figure 4A right panel). In contrast, analysis of the deleted regions in colonies from other patients showed a unique size for each of the del20q regions (not shown), making it doubtful that the pattern observed in p007 is the result of artifacts that occurred during the methylcellulose culture. Evidence for 2 independent del20q events was also found in patients Vi102 (Figure 4B) and Vi346 (Figure 4C). In both cases, colonies were found that showed loss of heterozygosity affecting 2 different parental chromosomes 20.

Figure 4

Multiple del20q and 9pLOH events in colonies from 3 MPD patients. (A) The deleted region on chromosome 20q was mapped in individual colonies from patient p007 using 6 microsatellites distributed along chromosome 20q. T-cell DNA was used to define the 2 alleles for each informative microsatellite. The gray boxes mark the deleted region for each individual colony. del20q in colony no. 185 created a different haplotype than the deletion in colony no. 6 (marked by arrows), indicating that 2 del20q events occurred independently and affected the chromosome 20q of different parental origin. Furthermore, 4 different sizes of the del20q haplotype were detected when additional colonies were analyzed. The results from the mapping of the 9pLOH region are shown on the right side. The 9pLOH region is smaller in colony no. 185, as shown by the heterozygosity of D9S2148 in this colony. (B) Two separate del20q events affecting the chromosome 20q of different parental origin occurred in patient Vi102. (C) Two separate del20q events affecting the chromosome 20q of different parental origin occurred in patient Vi346. The chromatograms for the MPL-W515L mutation are shown for the 2 colonies analyzed.

Figure 4

Multiple del20q and 9pLOH events in colonies from 3 MPD patients. (A) The deleted region on chromosome 20q was mapped in individual colonies from patient p007 using 6 microsatellites distributed along chromosome 20q. T-cell DNA was used to define the 2 alleles for each informative microsatellite. The gray boxes mark the deleted region for each individual colony. del20q in colony no. 185 created a different haplotype than the deletion in colony no. 6 (marked by arrows), indicating that 2 del20q events occurred independently and affected the chromosome 20q of different parental origin. Furthermore, 4 different sizes of the del20q haplotype were detected when additional colonies were analyzed. The results from the mapping of the 9pLOH region are shown on the right side. The 9pLOH region is smaller in colony no. 185, as shown by the heterozygosity of D9S2148 in this colony. (B) Two separate del20q events affecting the chromosome 20q of different parental origin occurred in patient Vi102. (C) Two separate del20q events affecting the chromosome 20q of different parental origin occurred in patient Vi346. The chromatograms for the MPL-W515L mutation are shown for the 2 colonies analyzed.

Close modal

In the present study, we assessed the potential connection between JAK2-V617F and del20q by analyzing the temporal order of acquisition of the 2 events. Our copy number assay allowed us to screen for del20q by real-time PCR in peripheral blood from a large number of MPD patients. The L3MBTL gene, which we have chosen for the copy number assay, is located within the CDR shared between patients with MPD and MDS/AML.9  This assay requires that the majority of granulocytes carry del20q and therefore selects for del20q events that dominate in granulocytes in the peripheral blood. Nevertheless, our observed frequencies in patients with ET, PV, and PMF of del20q are comparable with results obtained with cytogenetic studies of bone marrow.10,12  Furthermore, real-time PCR and cytogenetic analysis were in agreement in all 8 patients in whom both analyses have been performed. We found no difference in the frequencies of JAK2-V617F in PV and PMF patients with and without del20q. Our results do not confirm a previously reported preferential association of del20q with JAK2-V617F-positive cases of PMF (Table 2).13  The reason for this discrepancy is currently unclear. In ET, 4 of 4 patients with del20q were JAK2-V617F–positive, similar to 5 of 5 del20q-positive ET cases reported previously (Table 2).13  Thus, by combining the studies, 9 of 9 ET patients with del20q were also JAK2-V617F–positive. However, this association should be examined in a larger series of patients with del20q. Mapping of the del20q region by CGH revealed that in most patients large deletions have occurred (Figure 2) and that the CDR derived from analyzing granulocytes overlaps with the published region obtained by mapping bone marrow samples.9 

Disparity between the size of the JAK2-V617F–positive clone and the clone determined by analysis of the X-chromosome inactivation pattern,14,19  presence of endogenous erythroid colonies that are negative for JAK2-V617F,17,20,21  and coexistence of JAK2-V617F and MPL-W515L/K in the same patients,22,23  suggested that clonal events preceding the acquisition of JAK2-V617F exist in patients with MPD. We previously found 2 patients in whom the clone carrying del20q was larger than the clone positive for JAK2-V617F, and we hypothesized that JAK2-V617F preferentially occurs on the background of clonal hematopoiesis, which in some cases may be caused by del20q.14  Our single clone analysis confirmed the previously observed temporal order of events in 2 patients (Figure 3A), but in 3 additional patients we detected the inverse order, ie, JAK2-V617F preceding del20q (Figure 3B). Thus, there appears to be no strict temporal order of acquisition, making it doubtful that del20q represents a predisposing event for JAK2-V617F. At present, we cannot formally exclude the possibility that del20q may represent a “passenger mutation,” ie, a genetic alteration without functional consequence.24,25  The fact that in serial samples of patients Na293 and p007 we observed an expansion of the del20q subclone (Figure 3) could be interpreted in favor of a growth advantage provided by del20q and thus for a role as a “driver mutation.” Similarly, in patients p048 and p103, nearly all colonies were del20q-positive, suggesting a growth advantage of the cells harboring del20q.

Unexpectedly, in 3 patients (p007, Vi102, and Vi346), we found evidence for several independent del20q events and in p007 for 2 independent 9pLOH events (Figure 4). Patient p007 was diagnosed with PV 22 years ago and for 11 years showed signs of a progression to secondary myelofibrosis. The long disease duration and/or progression to spent phase may favor accumulation of de novo mutation events. However, the disease duration in ET patient Vi102 (4 years) and PMF patient Vi346 (6 years) was shorter. Genomic instability as a consequence of JAK2-V617F was recently reported.26  This could explain the findings in p007. However, in patient Vi102, one of the del20q events occurred in a JAK2-V617F–negative cell (Figure 3C) and patient Vi346 was negative for JAK2-V617F, suggesting that genomic instability in these patients is independent of JAK2.

The publication costs of this article were defrayed in part by page charge payment. Therefore, and solely to indicate this fact, this article is hereby marked “advertisement” in accordance with 18 USC section 1734.

We thank Michael Medinger for contributing clinical data, Jean-Luc Harousseau (Nantes) and Bruno Villemagne (La Roche sur Yon) for providing patient samples, and Jürg Schwaller and Alex Theocharides for comments on the manuscript.

This work was supported by the Swiss National Science Foundation (grants 310000-108006/1) and the Swiss Cancer League (OCS-01 742-08-2005) to R.C.S. and from the Initiative for Cancer Research of the Medical University of Vienna and the Austrian Science Fund FWF (P20033-B11) to R.K.

Contribution: F.X.S. performed research, analyzed data, and wrote the paper; R.J., R.L., and H.H.-S. performed research; S.H., F.G., A.T., H.G., and R.K. provided essential reagents and analyzed data; and R.C.S. designed research, analyzed data, and wrote the paper.

Conflict-of-interest disclosure: The authors declare no competing financial interests.

Correspondence: Radek C. Skoda, Department of Biomedicine, Experimental Hematology, University Hospital Basel, Hebelstrasse 20, 4031 Basel, Switzerland; e-mail: radek.skoda@unibas.ch.

1
Rege-Cambrin
G
Mecucci
C
Tricot
G
et al
A chromosomal profile of polycythemia vera.
Cancer Genet Cytogenet
1987
25
233
245
2
Demory
JL
Dupriez
B
Fenaux
P
et al
Cytogenetic studies and their prognostic significance in agnogenic myeloid metaplasia: a report on 47 cases.
Blood
1988
72
855
859
3
Diez-Martin
JL
Graham
DL
Petitt
RM
Dewald
GW
Chromosome studies in 104 patients with polycythemia vera.
Mayo Clin Proc
1991
66
287
299
4
Mertens
F
Johansson
B
Heim
S
Kristoffersson
U
Mitelman
F
Karyotypic patterns in chronic myeloproliferative disorders: report on 74 cases and review of the literature.
Leukemia
1991
5
214
220
5
Asimakopoulos
FA
White
NJ
Nacheva
E
Green
AR
Molecular analysis of chromosome 20q deletions associated with myeloproliferative disorders and myelodysplastic syndromes.
Blood
1994
84
3086
3094
6
Asimakopoulos
FA
Gilbert
JG
Aldred
MA
Pearson
TC
Green
AR
Interstitial deletion constitutes the major mechanism for loss of heterozygosity on chromosome 20q in polycythemia vera.
Blood
1996
88
2690
2698
7
Bench
AJ
Aldred
MA
Humphray
SJ
et al
A detailed physical and transcriptional map of the region of chromosome 20 that is deleted in myeloproliferative disorders and refinement of the common deleted region.
Genomics
1998
49
351
362
8
Bench
AJ
Nacheva
EP
Champion
KM
Green
AR
Molecular genetics and cytogenetics of myeloproliferative disorders.
Baillieres Clin Haematol
1998
11
819
848
9
UK Cancer Cytogenetics Group (UKCCG)
Bench
AJ
Nacheva
EP
Hood
TL
et al
Chromosome 20 deletions in myeloid malignancies: reduction of the common deleted region, generation of a PAC/BAC contig and identification of candidate genes.
Oncogene
2000
19
3902
3913
10
Douet-Guilbert
N
Basinko
A
Morel
F
et al
Chromosome 20 deletions in myelodysplastic syndromes and Philadelphia-chromosome-negative myeloproliferative disorders: characterization by molecular cytogenetics of commonly deleted and retained regions.
Ann Hematol
2008
87
537
544
11
Li
J
Bench
AJ
Vassiliou
GS
Fourouclas
N
Ferguson-Smith
AC
Green
AR
Imprinting of the human L3MBTL gene, a polycomb family member located in a region of chromosome 20 deleted in human myeloid malignancies.
Proc Natl Acad Sci U S A
2004
101
7341
7346
12
Gangat
N
Strand
J
Lasho
TL
et al
Cytogenetic studies at diagnosis in polycythemia vera: clinical and JAK2V617F allele burden correlates.
Eur J Haematol
2008
80
197
200
13
Campbell
PJ
Baxter
EJ
Beer
PA
et al
Mutation of JAK2 in the myeloproliferative disorders: timing, clonality studies, cytogenetic associations, and role in leukemic transformation.
Blood
2006
108
3548
3555
14
Kralovics
R
Teo
SS
Li
S
et al
Acquisition of the V617F mutation of JAK2 is a late genetic event in a subset of patients with myeloproliferative disorders.
Blood
2006
108
1377
1380
15
Kralovics
R
Buser
AS
Teo
SS
et al
Comparison of molecular markers in a cohort of patients with chronic myeloproliferative disorders.
Blood
2003
102
1869
1871
16
Kralovics
R
Passamonti
F
Buser
AS
et al
A gain-of-function mutation of JAK2 in myeloproliferative disorders.
N Engl J Med
2005
352
1779
1790
17
Li
S
Kralovics
R
De Libero
G
Theocharides
A
Gisslinger
H
Skoda
RC
Clonal heterogeneity in polycythemia vera patients with JAK2 exon12 and JAK2-V617F mutations.
Blood
2008
111
3863
3866
18
Liva
S
Hupe
P
Neuvial
P
et al
CAPweb: a bioinformatics CGH array Analysis Platform.
Nucleic Acids Res
2006
34
W477
W481
19
Levine
RL
Belisle
C
Wadleigh
M
et al
X-inactivation-based clonality analysis and quantitative JAK2V617F assessment reveal a strong association between clonality and JAK2V617F in PV but not ET/MMM, and identifies a subset of JAK2V617F-negative ET and MMM patients with clonal hematopoiesis.
Blood
2006
107
4139
4141
20
Nussenzveig
RH
Swierczek
SI
Jelinek
J
et al
Polycythemia vera is not initiated by JAK2V617F mutation.
Exp Hematol
2007
35
32
38
21
Pardanani
A
Lasho
TL
Finke
C
et al
Extending Jak2V617F and MplW515 mutation analysis to single hematopoietic colonies and B and T lymphocytes.
Stem Cells
2007
25
2358
2362
22
Pardanani
AD
Levine
RL
Lasho
T
et al
MPL515 mutations in myeloproliferative and other myeloid disorders: a study of 1182 patients.
Blood
2006
108
3472
3476
23
Lasho
TL
Pardanani
A
McClure
RF
et al
Concurrent MPL515 and JAK2V617F mutations in myelofibrosis: chronology of clonal emergence and changes in mutant allele burden over time.
Br J Haematol
2006
135
683
687
24
Greenman
C
Stephens
P
Smith
R
et al
Patterns of somatic mutation in human cancer genomes.
Nature
2007
446
153
158
25
Sjoblom
T
Jones
S
Wood
LD
et al
The consensus coding sequences of human breast and colorectal cancers.
Science
2006
314
268
274
26
Plo
I
Nakatake
M
Malivert
L
et al
JAK2 stimulates homologous recombination and genetic instability: potential implication in the heterogeneity of myeloproliferative disorders.
Blood
2008
112
1402
1412
Sign in via your Institution