Effectiveness and safety of high-dose cyclophosphamide as
salvage therapy for high-risk multiple myeloma and plasma cell
leukemia refractory to new biological agents
Guillermo L. Rivell,1Chris Y. Brunson,2Laura Milligan,1Robert K. Stuart,1and Luciano J. Costa1
Multiple Myeloma (MM) patients refractory to bortezomib and one or
more immunomodulatory drugs have a poor clinical outcome. Alkylat-
ing agents are often avoided in the initial management of MM in part
due to concerns for impairment of HSC mobilization. High doses of
cyclophosphamide (HiCy) administered without HSC support may be
an effective treatment to rescue MM patients refractory to novel biolog-
ical agents. We performed a retrospective single institution analysis of
17 consecutive MM patients receiving high dose cyclophosphamide
(HiCy, 3000 mg/m2) after failure of bortezomib and, in most cases, at
least one immunomodulatory agent (IMiD). Despite the prevalence of
high-risk features in this cohort we found HiCy to be an effective sal-
vage therapy for high-risk MM patients refractory to new biological
Multiple Myeloma (MM) is a malignant plasma cell disorder with no stand-
ard curative therapy  affecting 4.3 per 100,000 individuals yearly in the
united states, accounting for about 1% of all cancers and 10% of all hemato-
logical malignancies . Patients who receive conventional chemotherapy or
autologous stem cell transplantation have a mean overall survival of 3.7
years , although survival is likely increasing with the assimilation of new
biological agents into the management of MM [3–7].
For decades, melphalan and prednisone were the cornerstones of MM
management. Complete responses under this regimen are rare, and the
median time for progression is not higher than 15 months [5,8]. The immu-
nomodulatory (IMiD) drugs thalidomide and lenalidomide [6,7] along with the
first proteasome inhibitor bortezomib have shown efficacy in managing
relapsed and refractory MM [4,9,10]. These biological agents have been
quickly incorporated in the upfront management of MM so that regimens
containing one or more biological agents have produced the highest
response rates ever reported for previously untreated MM patients [3,11–
Despite these advances, patients refractory to, or progressing after, treat-
ment with one or more biological agents have a very poor prognosis and no
satisfactory therapeutic option [12,20]. A recent study from the International
Myeloma Working Group reported the outcomes of 270 patients relapsing
on or refractory to bortezomib and one IMiD. Only 30% of the patients had
an objective response to the next line of therapy. The median overall survival
and event free survival time were dismal eight months and five month,
With the enthusiasm generated by autologous hematopoietic stem cell
transplantation (HSCT) and the new biological agents, alkylating agents are
often excluded from the initial therapy of MM and patients failing new biologi-
cal agents are often alkylating-naı ¨ve. High-dose, single agent cyclophospha-
mide can be an attractive alternative in this setting since it does not require
stem cell support and is compatible with poor renal function. In this report
we present a retrospective analysis of consecutive patients with bortezomib-
refractory (and most IMiD-refractory) MM treated with high-dose cyclophos-
phamide and growth factor support at our institution.
A total of 17 patients treated between March 2009 and January 2011
were included in the analysis. Characteristics of the patients are displayed in
Table I. The median age of patients was 53 years (range, 26–73) and the
median time from initial diagnosis until cyclophosphamide treatment was
17.3 months (range, 1.6–106.4). Patients were heavily pretreated with the
median number of prior treatments being 3 (range, 1–6). All patients were
refractory to the last therapeutic regimen and had failed to respond or were
refractory to a regimen containing bortezomib. Twelve patients (70.6%) had
previously received at least one IMiD and three patients (17.6%) had previ-
ously received autologous HSC transplantation. Six patients (35.3%) met cri-
teria for plasma cell leukemia at the time of cyclophosphamide treatment
and eight (47.1%) had renal dysfunction at the time of treatment including
two patients on chronic hemodialysis. The patients included in the series
were not eligible for any trials available at the institution or in the area due
to one or more of the following reasons: advanced renal dysfunction (n 5
8), plasma cell leukemia (n 5 6), ECOG performance status >2 (n 5 5) or
bortezomib refractoriness (n 5 17).
Median follow up was 4.2 months (range, 1.7–21.4) from cyclophospha-
mide administration. Response was assessed four to six weeks after admin-
istration of cyclophosphamide. Overall, nine patients had a partial response
or better (52.9%, 95%C.I. 29.2–76.7%), including 4 patients with a very
good partial response (23.5%, 95%C.I. 3.4–43.7%). All the remaining
patients had stable disease. Five of the 12 patients previously treated with
at least one IMiD obtained a response (41.6%) along with 4/7 patients previ-
ously treated with an alkylating agent (57.1%). Median overall survival was
14.1 (±7.1) months (see Fig. 1). Interestingly, all six patients with plasma
cell leukemia had complete clearance of plasma cells from the peripheral
blood. Among the 12 patients with intent to proceed with HSCT (cyclophos-
phamide used as ‘‘bridge’’ therapy), eight patients underwent either autolo-
gous (n 5 7) or allogeneic (n 5 1) transplantation. The median survival for
these patients is estimated at 21 months. The remaining four patients did
not receive HSCT because of refusal (n 5 1), fast disease progression (n 5
2), or worsening performance status and comorbidities (n 5 1). Overall,
eight patients have died, one due to toxicity of treatment and seven from dis-
Alkylating agents are an important component of both conventional and
high-dose chemotherapy-based treatment of MM. In recent years, due to the
enthusiasm about new biological agents and the concerns regarding impair-
ment of future HSC mobilization, alkylating agents have often been avoided
in the initial management of MM. We believe that many of the patients
becoming refractory to proteasome inhibitors and IMiDs were still sensitive
to alkylating agents and could be rescued with high-dose cyclophosphamide
without HSC support.
There are few series on the management of MM refractory to proteasome
inhibitors and IMiD . These patients should be urged to participate in
clinical trials with new drugs, including new proteasome inhibitors, histone
deacetylase inhibitors, anti-CS1 antibodies and third generation IMiDs. A
recent report including 270 ‘‘double refractory’’ patients from the Multicenter
International Myeloma Working Group indicates a median survival of eight
months from the time the patients failed last therapy. Remarkably, only 30%
of patients had an objective response to the subsequent line of therapy .
Even though the present series include higher proportions of patients with
Stage 3 disease at the diagnosis, unfavorable chromosomal abnormalities,
and 3 or more prior lines of therapy, objective response (partial response or
better) was documented in 52.9% of the patients. Another interesting obser-
vation is that all six patients with plasma cell leukemia had clearance of cir-
culating plasma cells within four weeks of administration of cyclophospha-
mide. Comparisons between our series and the much larger series by
Kumar et al.  requires caution due to the small sample size, lower pro-
portion of patients refractory to an IMiD (70.6 vs. 100%) and lower propor-
tion of patients with prior autologous hematopietic stem cell transplantation
(17.6 vs. 74%).
There are significant limitations of this therapy that need to be empha-
sized. Despite antibiotic prophylaxis and growth factor support, the majority
of patients developed fever and neutropenia requiring hospitalization, and
one patient died from complications of sepsis. This is therefore a toxic regi-
men requiring careful patient selection. The most important limitation how-
ever is the short duration of the responses obtained. In fact, all responding
patients either died as consequence of disease progression or required
C 2011 Wiley-Liss, Inc.
American Journal of Hematology
subsequent therapy within three months of receiving cyclophosphamide.
This aspect is responsible, at least in part, for the short median overall sur-
vival despite the considerable rate of objective responses. We suggest
therefore that cyclophosphamide can be used as a ‘‘bridge’’ strategy for
more definitive therapy, particularly in patients needing immediate disease
The most important point of the current report is that in the era of biologi-
cal agents and deferred use of alkylating agents, many patients who
become ‘‘double refractory’’ are still sensitive to alkylating agents. Although
high doses of cyclophosphamide is one valid option to explore such sensitiv-
ity, combination of cyclophosphamide and biological agents may be a safer
and equally successful approach [19,22]. In fact, there is justifiable enthusi-
asm for the potential of bortezomib to inhibit NF-kB mediated activation of
the Fanconi/BRCA pathway and enhance DNA damage induced by alkylat-
ing agents .
In summary, high doses of cyclophosphamide can transiently control high-
risk MM refractory to novel biological agents. Clinical trials are necessary to
redefine the best use of alkylating agents in MM in a therapeutic landscape
dominated by safe and highly active biological agents.
This is a retrospective single institution review of all consecutive patients
with bortezomib-refractory MM treated with high-dose cyclophosphamide at
the Medical University of South Carolina. Inclusion criteria were firm diagno-
sis of symptomatic MM, prior disease progression while receiving therapy or
within 60 days of completing therapy with a bortezomib containing regimen,
and intent to treat with high doses of cyclophosphamide without hemato-
poietic stem cell (HSC) support.
Treatment consisted of cyclophosphamide 3,000 mg/m2administered over
1 hr. The dose of cyclophosphamide was fractioned in two days (1,500 mg/
m2/day) for patients receiving outpatient therapy. All patients, with the excep-
tion of two anuric patients in chronic dialysis, received mesna for prophylaxis
of hemorrhagic cystitis (20% of the cyclophosphamide dose at 0, 4, and 8 hr
of cyclophosphamide infusion). All patients received antibiotic prophylaxis
with ciprofloxacin, acyclovir, and fluconazole and growth factor support with
We obtained Institutional Review Board approval to perform this analysis.
Data extracted included de-identified patient demographics, disease charac-
teristics, treatment history, toxicity, response to therapy, subsequent therapy,
and survival data. Disease was characterized according to the International
Staging System  and chromosomal abnormalities determined by stand-
ard metaphase karyotype and/or fluorescence in situ hybridization (FISH).
Treatment-induced adverse events were graded according to the National
Cancer Institute common terminology criteria for adverse events (CTCAE)
version 4.0. For response assessment we utilized the International Uniform
Response Criteria for Mutiple Myeloma .
The main endpoint of the study was to determine the overall response rate
for high-dose cyclophosphamide in bortezomib-refractory MM patients. Secon-
dary endpoints were to determine median overall survival, frequency and
severity of adverse events. Progression free survival and duration of response
was not estimated since this endpoint was distorted by the high proportion of
patients undergoing transplantation immediately after a response was
obtained. We reported proportions with their respective 95% confidence inter-
vals. Continuous numerical variables were described using their median and
range. Overall survival was described utilizing the method of Kaplan-Meier.
Statistical analysis was performed the software SPSS (version 16).
1Division of Hematology and Oncology, Medical University of South Carolina,
Charleston, South Carolina
2Mabry Cancer Center, Orangeburg, South Carolina
*Correspondence to: Luciano J. Costa, MD, PhD, 96 Jonathan Lucas Street, CSB
903, Charleston, SC, 29414
Received for publication 22 February 2011; Revision 5 April 2011;
Accepted 10 April 2011
Conflict of interest: Nothing to report.
Published online 18 April 2011 in Wiley Online Library
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TABLE I. Characteristics of Patients Refractory to Novel Biological Agents
and Treated with High-Dose Cyclophosphamide
Median age (range)
3 or more abnormalities
ECOG Performance Status
Plasma cell leukemia
Median time diagnosis-treatment in months (range)
Number of prior lines of therapy
3 or more
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700 American Journal of Hematology
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Infrequent occurrence of MPL exon 10 mutations in
polycythemia vera and post-polycythemia vera myelofibrosis
Animesh Pardanani,Terra L. Lasho,Christy M. Finke, and Ayalew Tefferi*
Polycythemia vera (PV) is molecularly well characterized with 96% and
3% of patients exhibiting JAK2V617F and JAK2 exon 12 mutations,
respectively [1,2]. MPL exon 10 mutations have not been identified in
PV; among the Philadelphia-negative classic myeloproliferative neo-
plasms (MPN), MPL mutations appear to be restricted to essential
thrombocythemia (ET) or myelofibrosis (primary or post-ET) with over-
all mutation frequency of 4 and 8%, respectively [3–6]. Although
JAK2V617F and MPL mutations have been identified concurrently in
ET and primary myelofibrosis (PMF), such occurrences are relatively
infrequent, and this combination has not been described in PV [3–6].
We screened 168 patients, 149 with PV and 19 with post-PV MF, for
presence of JAK2 and MPL mutations; 162 patients (96.4%) and two
patients (1.2%) harbored JAK2 and MPL mutations, respectively. Both
MPL mutated patients, one each with PV and post-PV MF, were also
JAK2V617F-positive. Thus, PV patients may infrequently harbor MPL
mutations suggesting the disease is molecularly more complex than
previously appreciated. It reinforces the view that currently identified
MPN-relevant mutations are neither mutually exclusive nor are they
We screened a total of 168 patients for presence of JAK2V617F and MPL
exon 10 mutations; of these, 149 patients had PV and 19 patients post-PV
MF. Overall 162 patients (96.4%) harbored JAK2 mutations; of these, 160
patients exhibited JAK2V617F and two patients JAK2 exon 12 mutations
(both with PV). JAK2 mutation frequency in PV and post-PV MF was 95.9
and 100%, respectively. Two patients (1.2%), one each with PV and post-PV
MF, harbored a MPL exon 10 mutation; thus, MPL mutation frequency was
< 1% in PV and 5% in post-PV MF in this cohort. The PV patient harbored
the MPLW515R allele and the post-PV MF patient MPLW515K (Fig. 1). Both
patients with MPL mutations harbored JAK2V617F concurrently.
Clinical and laboratory information for the two MPL mutated patients is as
follows: Patient #1 was a 45-year-old male who diagnosed with PV approxi-
mately a year prior to his initial presentation at our institution. A review of
his laboratory records showed peak hemoglobin (Hgb) value of 19.4 g/dL.
The serum erythropoietin level was undetectable or demonstrated to be
abnormally low on several occasions. Phlebotomies were instituted after the
diagnosis was established; however, no cytoreductive therapy was initiated.
At the time of Mayo referral, the laboratory values were as follows: Hgb 15.2
g/dL, hematocrit 47.1%, red blood cell mean corpuscular volume 75.9 fl,
serum ferritin level 6 mg/L, and serum erythropoietin level 1.9 mU/mL
and MPL exon 10 mutations in a patient with (a) PV and (b) post-PV myelofibrosis
(post-PV MF). The former patient (PV) harbored MPLW515R and the latter (post-
PV MF) MPLW515K. The arrows indicate the nucleotide substitution for missense
mutations. [Color figure can be viewed in the online issue, which is available at
DNA sequence traces showing concurrent presence of JAK2V617F
American Journal of Hematology701
(normal range 4-24). Bone marrow examination showed morphologic find-
ings consistent with treated PV (absent iron stores); reticulin stain showed
normal reticulin fibers and cytogenetic studies revealed a normal karyotype.
Analysis of DNA collected at the time of referral showed presence of
JAK2V617F and the MPLW515R mutation in the heterozygous state (Fig. 1,
Patient #2 was a 54-year-old female with a diagnosis of blastic transfor-
mation of post-PV MF; at the time of referral to our institution she had re-
achieved chronic-phase disease after having received an allogeneic stem
cell graft from her HLA-identical sibling brother. PV had been diagnosed 22
years prior; subsequent to the interim fibrotic transformation of PV, she had
received multiple therapies including hydroxyurea, erythropoiesis stimulating
agents, androgens, and had also undergone splenectomy. At referral, the
patient was pancytopenic and bone marrow examination showed myelofibro-
sis with approximately half the metaphases exhibiting recipient-derived com-
plex cytogenetic abnormalities. Analysis of DNA collected at the time of
referral showed presence of JAK2V617F and the MPLW515K mutation in
the heterozygous state (Fig. 1, bottom panel).
The identification of MPL mutations in PV and post-PV MF was somewhat
unexpected given previous reports of the putative anti-erythropoietic effect
of such mutations. In contrast to JAK2V617F-mutated patients, we and
others have failed to obtain erythropoietin-independent erythroid colonies in
vitro in studies of PMF or ET patients [3,7,8]. Furthermore, in ET, patients
harboring MPL mutations exhibit significantly lower hemoglobin values,
higher serum erythropoietin levels, and reduced total and erythroid bone
marrow cellularity, as compared to MPL wild-type/JAK2V617F positive
patients [3,6]. Also, some  but not other studies (Pardanani et al., manu-
script submitted) in PMF patients have identified an association between
MPL mutations and more severe anemia as well as increased dependency
for red cell concentrate transfusions.
Although this study clearly demonstrates that JAK2V617F and MPL muta-
tions can coexist in PV/post-PV MF, the cumulative published data to date
suggest that such occurrences are likely to be rare. Likewise, in ET the
mutation concurrence rate is likely ? 1% [3,6]. In contrast, in PMF, the pub-
lished mutation concurrence rate has varied widely from 0% (Pardanani
et al., submitted) to up to 38% [4,5], likely reflecting the smaller patient num-
bers relative to ET and a more heterogeneous patient population.
The number of MPL-mutated PV/post-PV MF patients is too small to draw
any conclusions regarding possible clinical or laboratory associations or
impact on clinical outcome; the current observations do however further
demonstrate the lack of disease specificity of MPN-relevant mutations and
call into question the presumed phenotype-modifying properties of such
mutations. Given that the MPL mutations in the aforementioned PV/post-PV
MF cases were easily identified by direct DNA sequencing and given the
observed mutant: wild-type allele peak heights on DNA chromatograms (Fig.
1), the current observations cannot be easily explained by presence of a
minor MPL mutant clone on a background of a dominant JAK2V617F har-
boring clone. Further, at least in the case of the aforementioned PV case,
the MPLW515R mutation was demonstrable relatively early during the
course of the disease and in the absence of any cytoreductive or potentially
mutagenic therapy being administered. A similar early acquisition of MPL
mutations has been previously demonstrated in PMF and ET [3,9,10].
Although these observations alone do not provide conclusive evidence for
the pathogenetic relevance of MPL mutations in PV, it does not exclude this
The current data do not clarify whether each mutation targets an inde-
pendent stem cell clone (i.e., biclonal disease) or whether both mutations
arise in the same stem cell clone with sequential mutation acquisition (i.e.,
clonal evolution) . Unfortunately, unavailability of viably preserved cells
from the two PV/post-PV MF patients precluded genotyping of individual
hematopoietic colonies grown in vitro that may have clarified this issue.
This study was approved by the Mayo Clinic institutional review board.
Patients provided informed written consent for study sample collection as
well as permission for its use in research. Inclusion to this study required
availability of archived peripheral blood and/or bone marrow sample col-
lected at the time of diagnosis or first referral. The diagnoses of PV and
post-PV MF were according to the World Health Organization criteria .
Screening of patients for MPL exon 10 genetic variations was performed
using the High Resolution Melting (HRM) Dye assay using the LightCycler1
480 Instrument (Roche-Applied-Science, Mannheim, Germany). For each
20-mL reaction, 15–30 ng of DNA template was added to a reaction mix of:
1.5-mM MgCl2, 500 nM of both forward and reverse primer (MPL-forward 50-
TAGTGTGCAG-30), ddH20, and 1X of enzyme mix (HRM Master Kit, Roche-
Applied-Science, Mannheim, Germany). Each reaction was first amplified by
real-time PCR using the following cycling parameters: 958C for 10 min fol-
lowed by 45 cycles of 958C for 10 s, 608C for 15 s, and 728C for 30 s. After
amplification, each target sequence was subjected to an increasing tempera-
ture and data from the intercalated HRM Melting Dye signal resulting from
dissociation of double-stranded DNA to single-stranded DNA was plotted.
Analysis of melting curves was executed using Gene Scanning Software
(Roche-Applied-Science, Mannheim, Germany) and was normalized to wild-
type sequence. Any sample sequence that deviated from the normal plot
was Sanger sequenced for confirmation of presence of MPL genetic varia-
tion. Sequencing was performed bidirectionally using two ABI PRISM2
3730xl DNA analyzers (96-capillary). JAK2V617F mutation screening was
performed as previously described .
A Pardanani, TL Lasho, and A Tefferi designed the study, contributed
patient samples, analyzed the data, and wrote the paper. TL Lasho and C
Finke performed the mutational analysis.
Division of Hematology, Department of Medicine, Mayo Clinic
Contract grant sponsor: The Henry J Predolin Foundation
*Correspondence to: Ayalew Tefferi, MD, Division of Hematology, Mayo Clinic
200 First Street SW, Rochester, MN
Conflict of interest: Nothing to report.
Published online 19 April 2011 in Wiley Online Library
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702 American Journal of Hematology
Novel CUX1 missense mutation in association with 7q2 at
leukemic transformation of MPN
Nils H. Thoennissen,1,2* Terra Lasho,3Gabriela B. Thoennissen,1,2Seishi Ogawa,4
Ayalew Tefferi,3and H. Phillip Koeffler1,5
Patients with monosomy 7 (27) or del(7q) comprise a heterogeneous
subgroup in acute myeloid leukemia (AML) but no specific target
genes have been identified [1–4]. Recently, we detected a commonly
deleted region on the long arm of chromosome 7 in a large cohort of
(MPN) at the time of leukemic transformation using single-nucleotide
polymorphism array (SNP-A) [5,6]. This region was located on 7q22.1
encompassing only two genes, CUX1 and SH2B2. Currently, we
screened for acquired mutations in these two candidate genes using
15 secondary AML cases with preceding MPN and loss-of-heterozygos-
ity (LOH) on chromosome 7q. We detected a novel hemizygous mis-
sense mutation (V1288L) in the HOX domain of the transcriptional reg-
ulator CUX1 in one case with primary myelofibrosis (PMF) at time of
leukemic evolution. Although only detected in 1/15 (6.5%) of the cases
with secondary AML, an acquired mutation of CUX1 may play a critical
role in myeloid malignancies with 7q aberrations.
Abnormalities involving chromosome 7q are frequently detectable in mye-
loid malignancies [1–4]. The association of losses in 7q with AML suggests
that this region contains a tumor suppressor gene or genes whose loss of
function contributes to leukemic transformation or tumor progression. Based
on several preceding chromosome banding studies, two minimal deleted
regions have been identified; one locus at centromeric band 7q22 and the
other at telomeric breakpoint varying from q32 to q36 [7–9]. The complexity
of 7q rearrangements suggests that a synergy of different genetic factors,
rather than the alteration of a single tumor suppressor gene, could be
involved in the pathogenesis of 7q2 in myeloid disorders. Two recent studies
identified monoallelic or biallelic loss-of-function mutations in the histone H3
methyltransferase EZH2 on 7q36.1 in patients with myelodysplastic/s (12%)
and in those with myelofibrosis (13%); the authors suggested EZH2 func-
tions as a tumor-suppressor gene in these malignancies rather than an
oncogene as in some other malignancies [10,11]. Notably, no EZH2 muta-
tions were found so far in de novo or secondary patients with AML having
complete or partial monosomy for chromosome 7.
Using high-density SNP-A provides a robust and detailed approach to
detect large and small copy-number changes, as well as copy-number neu-
tral (CNN-) LOH. We recently applied this interrogational method and per-
formed a systemic analysis of 159 samples obtained from patients with
either MPN or secondary AML with preceding MPN to obtain a comprehen-
sive profile of genomic alterations associated with leukemic transformation in
MPN disease . Complete or partial deletion (27/7q2) and CNN-LOH of
the long arm of chromosome 7 were one of the most common abnormalities
detected by SNP-A analysis in 25% of samples with secondary AML, and
associated with inferior outcome. The minimal deleted region spanned a
small region at 7q22.1.
In this study, we used 15 secondary AML cases with chromosome seven
aberrations and prior MPN (Table I) to screen for novel tumor suppressor
genes. Using both cytogenetics and SNP-A, 11 samples (73%) showed het-
erozygous deletions on 7q; three samples (20%) had CNN-LOH on 7q; and
the analysis of one case (7%; #138) revealed both CNN-LOH on 7q21.13-
qter and a small homozygous deletion on 7q22.1 (0.88 Mbp) encompassing
only two candidate genes, CUX1 and SH2B2 (Fig. 1). One of the major pro-
tein function of SH2B2 (alias APS) is the recruitment of c-Cbl into the recep-
tor/JAK complex, and thereby inhibiting JAK/STAT signaling activity [12,13].
CUX1 (alias Cut homeobox 1 or CUTL1) belongs to a family of transcription
factors with homeodomain (HOX) involved in the control of cell proliferation
and differentiation .
First, we investigated all coding exons of SH2B2 in the 15 patients with
secondary AML by direct sequencing, but no genetic variation, either SNP
or mutation, could be detected. Instead, after screening the coding regions
of the three Cut domains and the HOX domain of the CUX1 gene, we
detected not only a SNP in case #34, #79, and #80 (Table I) but also a
genetic variation in a highly conserved nucleotide position in case #90
(V1288L; Table I), which has not been previously described in the ‘‘Database
of Genomic Variants’’ (http://projects.tcag.ca/variation/), the ‘‘UCSC Genome
Browser’’ (http://genome.ucsc.edu/), or the ‘‘Cosmic Catalogue for Somatic
Mutations in Cancer’’ (http://www.sanger.ac.uk/genetics/CGP/cosmic/). We
were able to characterize the novel nucleotide variant (GTC ? CTC) as a
somatic mutation by showing a normal genetic code in a serial sample of
case #90 originating from the chronic phase of MPN before leukemic evolu-
tion (Fig. 2A). Interestingly, not only the V1288L missense mutation in CUX1
was acquired at the time of transformation to secondary AML but also the
loss of the normal allele with del(7)(q11.21qter) as indicated by SNP-A .
The V1288L mutation is located in the coding region of the DNA-binding
HOX domain of CUX1 (Fig. 2B) and has statistically a detrimental effect on
TABLE I. Genetic Aberrations on Chromosome 7 in 15 Patient Samples from Time of Leukemic Transformation after Prior MPN
Case #DiagnoseJAK2V617F Prior MPNCytogenetics SNP-A (breakpoints)Aberration size [Mbp] Genetic Variation in CUX1
del(7)(q11 .22 qter)
del(7)(q11 21 qter)
rs76202142 G/T (SNP; Intron 16-17)
rs73712454 G/T (SNP; Intron 22-23)
rs73712454 G/T (SNP; Intron 22-23)
V1288L (rnissense mutation; Exon 24)
Indicated are detected aberrations on chromosome 7 and in CUX1 on 7q22.1. ET, essential thrombocythemia; PMF, primary myelofibrosis; PV, polycythemia vera; sAML,
secondary acute myelogenous leukemia; Mbp, megabase pairs.
American Journal of Hematology703
transformed MPN. Each horizontal line represents abnormality detected in each patient; green: CNN-LOH; blue: deletion. (B) One patient, #138, with transformed MPN
showed CNN-LOH on 7q, as well as a homozygous deletion on 7q22.1 encompassing only CUX1 and SH2B2. AsCN, allele-specific copy number; CN, copy number;
CNN-LOH, copy-number neutral loss-of-heterozygosity; SNP, single nucleotide polymorphism. [Color figure can be viewed in the online issue, which is available at
Aberrations on chromosome 7 in patients with transformed MPN. (A) Indicated are SNP-Chip findings on Chromosome 7 in 15 patients (see Table I) with
(V1288L) in the CUX1 gene after leukemic transformation. (B) Indicated is the schematic overview of the CUX1 protein including its three CUT domains, the HOX
domain, as well as the three known isoforms p75, p110, and p200 (according to Sansregret et al, 2008) . AA, amino acid. [Color figure can be viewed in the online
issue, which is available at wileyonlinelibrary.com.]
Novel somatic mutation of CUX1 in a paient with transformed MPN and 7q2. (A) Nucleotide sequencing of MPN case #90 revealed one missense mutation
704 American Journal of Hematology
the CUX1 protein (non-neutral, reliability index 2, accuracy 70%) according
to the SNAP software, a neural-network–based method to make prediction
regarding the functionality of a mutated protein .
The human CUX1 spans at least 340 kb, contains 33 exons, and at least
three protein isoforms can be expressed as the result of proteolytic proc-
essing or transcription initiation at an alternative start site (Fig. 2B) .
The full-length protein, p200, is a complex protein with four evolutionarily
conserved DNA-binding domains: three Cut repeats and a Cut homeodo-
main (HOX; Fig. 2B). CUX1 was originally shown to function in precursor
cells of various lineages as a transcriptional repressor that down-modulates
lineage specific genes that later become expressed in terminally differenti-
ated cells [14,16]. Noteworthy, homozygous mutant mice expressing a
hypomorphic and non-functional HOX domain of the CUX1 protein demon-
strated myeloid hyperplasia, suggesting that CUX1 functions as a tumor
suppressor and that its loss is a significant event in the generation or pro-
gression of myeloid disorders . However, one study performing a muta-
tional analysis of childhood samples with AML and monosomy 7 revealed
no somatic mutations in CUX1 . In contrast, transgenic mice expressing
the short isoform of CUX1, p75, displayed heightened susceptibility to
mammary tumors and a myeloproliferative disease-like myeloid leukemia,
pointing to an oncogenic role of CUX1 [19,20]. In consequence, regarding
to present literature, CUX1 has a critical regulatory influence on the myeloid
development and may act as either transcriptional repressor or activator
depending on its isoform.
In summary, this is the first report of an acquired missense mutation of
CUX1 in a patient with secondary AML and 7q2. Although detected in only
one case (6.5%), it may contribute to the pathogenesis of AML in a subset
of patients including the transformation or progression of chronic myeloid
diseases in association with 7q2. Larger cohorts of myeloid diseases and
further functional analysis are required to explore the impact of the novel
mutation in the CUX1 gene.
H.P.K. is the holder of the Mark Goodson endowed Chair in Oncology
Research at Cedars Sinai Medical Center and is a member of the Jonsson
Cancer Center and the Molecular Biology Institute, UCLA.
1Division of Hematology and Oncology, Cedars-Sinai Medical Center, UCLA
School of Medicine, Los Angeles, California;2Department of Hematology and
Oncology, University Hospital of Mu ¨nster, Mu ¨nster, Germany;3Department of
Hematology, Mayo Clinic, Rochester, Minnesota;4Department of Regeneration
Medicine for Hematopoiesis, Graduate School of Medicine, University of Tokyo,
University of Tokyo Hospital, Tokyo, Japan;5National Cancer Institute of
Singapore, National University of Singapore, Singapore
Grant sponsor: Deutsche Forschungsgemeinschaft (DFG, Bonn, Germany); Grant
numbers: TH 1438/1-1, N.H.T. Grant sponsor: National Institutes of Health (NIH,
Bethesda, MD); Grant number: 5R01CA026038-32. Grant sponsor: National
University of Singapore (A*STAR grant of Singapore)
*Correspondence to: Nils H. Thoennissen, MD, Department of Medicine
A—Hematology and Oncology, University Hospital Mu ´nster,
Albert-Schweitzer-Strasse 33, 48149 Mu ´nster, Germany
A.T. and H.P.K contributed equally to this work.
Conflict of interest: Nothing to report.
Published online 27 April 2011 in Wiley Online Library
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Laboratory and echocardiography markers in sickle cell patients
with leg ulcers
Caterina P. Minniti,1* James G. Taylor VI,1Mariana Hildesheim,1Patricia O’Neal,2Jonathan Wilson,1
Oswaldo Castro,2Victor R. Gordeuk,2and Gregory J. Kato1
Chronic leg ulcers are a debilitating complication of sickle cell dis-
ease, associated with increased morbidity and perhaps mortality that
affect 8–50% of patients. We evaluated the characteristics of sickle cell
disease (SCD) patients with a history of leg ulceration, including
hemolytic rate, estimated pulmonary artery systolic pressure, and
other parameters in a cohort of 505 adults with SCD. Ninety-four sub-
jects (18%) had either active ulcers at enrollment or history of leg
ulceration. Patients affected were older and predominantly had homo-
zygous hemoglobin SS, lower body mass index, and pulse oximetry,
higher tricuspid regurgitation velocities, markers of hemolysis, serum
American Journal of Hematology705
uric acid and serum amino-terminal probrain type natriuretic peptide,
when compared to subjects without such history. In this prospective
cohort of adults with SCD, we confirm that leg ulcers are still frequent
and are associated with elevated tricuspid jet velocity (TRV) and
markers of hemolysis. We describe a novel association of leg ulcer
with hyperuricemia and oxygen desaturation and suggest potential
implications for uric acid as a marker of vascular dysfunction.
The prevalence of leg ulceration in sickle cell disease patients varies,
being low before 10 years of age, and in genotypes other than SS, and it is
influenced by geographical location, with an occurrence as high as 75% in
Jamaica and 8–10% in North America [1–3]. Advances in understanding the
pathophysiology and management of leg ulcers have been slow. Abnormal
vascular tone and activated, adhesive endothelium have been proposed as
a pathway of end organ damage in sickle cell disease. Vasculopathy has
been implicated in the development of sickle leg ulcers and other complica-
tions and is associated with hemolytic severity . Patients with leg ulcera-
tion are at risk to develop serious complications of sickle cell disease, such
as pulmonary hypertension, priapism, and possibly, renal disease [5–8].
Our investigation was carried out in a large cohort of adult patients with
SCD screened prospectively for pulmonary hypertension by echocardio-
gram. We aimed to corroborate that leg ulcers are a manifestation of the
hemolysis/vasculopathy subphenotype and identify additional clinical and
laboratory characteristics associated with leg ulcers.
The characteristics of the subjects are reported in Table I.
Ninety-four (18%) of the 505 subjects reported having an active ulcer (24
subjects) or a history of leg ulceration (70 subjects). Most of the affected
subjects had homozygous HbS (85 or 90.4%), whereas nine had double het-
erozygosity for HbS and HbC (9.6%). Affected patients were older (median
age of 39 years compared to 31 years in the nonleg ulcer group, P <
0.0006). The body mass index (BMI) of subjects with leg ulcers was signifi-
cantly lower than in patients without such history, with over a third of them
having a BMI < 20.5 (P 5 0.01), implying that patients affected with ulcers
are generally underweight. The overall prevalence of hydroxyurea use was
39%, specifically 40.5% of the subjects without a history of leg ulcers were
taking hydroxyurea compared to 34% of the subjects with a history of leg
ulcers (P 5 0.2). Similar findings were obtained among SS only: 46% versus
36% (P 5 0.14). Hemoglobin F percentage was not different in the two
groups (P 5 0.6). The protective effect of fetal hemoglobin for the occur-
rence of leg ulcers has been inconsistent among several studies [3,6,8,9]. A
potentially confounding factor in modern cohorts is the widespread use of
hydroxyurea in contrast to earlier analyses [3,9]. It is possible that hydrox-
yurea statistically unlinks leg ulceration and fetal hemoglobin levels. Our
study design could not answer whether hydroxyurea use in SCD is useful or
detrimental for leg ulcers, because we did not establish the temporal rela-
tionship between hydroxyurea and development of leg ulceration.
Some authors have recommended transfusion therapy for the management
of leg ulcers . We did not show a correlation between levels of hemoglobin
A, a marker of recent transfusion, and prevalence of leg ulcers. On the other
hand, lower hemoglobin was associated with a history of leg ulcers in patients
with and without a-thalassemia. Patients with a history of leg ulcers exhibited
more abnormal levels of markers of hemolysis, such as significantly higher
serum lactate dehydrogenase (LDH) levels (P < 0.0001), absolute reticulocyte
counts (P < 0.005), aspartate transaminase AST (P 5 0.0004), and lower
hemoglobin concentration (P < 0.0001). They also had 20% higher serum uric
acid concentrations (P < 0.0001) alkaline phosphatase, and slightly lower
serum albumin (P < 0.0001). Median ferritin level was higher in the leg ulcer
group (528 vs. 388 mg/dL), which might be indicative of chronic inflammation
and/or a population that is more frequently transfused because of more severe
disease. The proportion of subjects that reported history of chronic transfusion
was similar in the two groups, 16% in the ‘‘no ulcer’’ and 20% in the ‘‘yes
ulcer,’’ leading to speculation that high ferritin is at least in part because of
chronic inflammation. Higher iron burden may also contribute to vascular dys-
function. The role of a-thalassemia trait as a modulator for leg ulcer risk has
been proposed by some investigators [3,9]. Alpha thalassemia status was
known in a subset of subjects, 294/505 (58%) and did not affect the preva-
lence of leg ulcers. The relationship of a-thalassemia trait to elevated esti-
TABLE I. Laboratory Characteristics and Demographics of Study Participants by History of Leg Ulcers
History of leg ulcers
N Median (IQR)a
N Median (IQR)a
N Median (IQR)a
Men, N (%)
HbSC disease, N (%)
Deaths, N (%)
BMI < 20.5, N (%)
HbA ? 50%, N (%)
Hydroxyurea, N (%)
Alpha thalassemia (a/2), N (%)
White blood cell count (31023/mL)
Platelet count (31023/mL)
Reticulocyte count (31023/mL)
Fetal hemoglobin (%)
Blood urea nitrogen (mg/dL)
Lactate dehydrogenase (U/L)
Alanine aminotransferase (U/L)
Aspartate aminotransferase (U/L)
Alkaline phosphatase (U/L)
Total bilirubin (mg/dL)
Direct bilirubin (mg/dL)
Uric acid (mg/dL)
O2Sat < 93%, N (%)
O2Sat < 96%, N (%)
NT–Pro BNP (pg/mL)
TRV ? 2.5 (m/s) (%)
TRV ? 3.0 (m/s) (%)
aExcept where otherwise noted.
bFrom Wilcoxon rank-sum test or Pearson’s chi square statistic between history of leg ulcer groups.
706American Journal of Hematology
mated pulmonary arterial pressure has been similarly inconsistent [11–13].
These apparently conflicting results could be due to the small magnitude of
the effect of a-thalassemia, below the detection of some studies. Moreover, a-
thalassemia determination was available on only in ?60% of the total popula-
tion studied in our study, limiting our statistical power.
After adjusting for age, hemoglobin, LDH, AST, albumin, uric acid, reticulo-
cyte count, direct bilirubin, and amino-terminal probrain type natriuretic pep-
tide (NT-proBNP) were still significantly associated with leg ulcer history.
Multivariate logistic regression analysis, presented in Table II, identified older
age (P < 0.001), lower hemoglobin concentration (P < 0.001), and higher
AST (P 5 0.06) as independent correlates of leg ulcer history in our cohort.
We also evaluated leg ulcer history as a potential predictor of elevated TRV.
After adjustment for other significant predictors of elevated TRV in our
cohort, leg ulcer history was marginally associated with TRV ? 2.5 m/sec (P
5 0.06), but not with TRV ? 3.0 m/sec (P 5 0.4).
Pulse oximetry readings were lower in subjects with leg ulcers. An oxygen
saturation level <93% was three times more prevalent among subjects with
a history of leg ulcers (22.4%) compared to subjects without a history of leg
ulcers (7.6%, P 5 0.0003). Recent publications report that, in SCD patients,
small decreases of steady-state oxygen saturation correlate to the degree of
anemia and indices of hemolysis, TRV, 6-min walk distance and risk for
stroke or other central nervous system events [14–19]. The mechanisms by
which a slight desaturation contributes to the risk of leg ulceration are not
clear. However, steady-state desaturation is associated with more prominent
desaturation during sleep which in turn causes endothelial, leukocyte, and
erythrocyte activation and adhesion [16,18]. These events could contribute
to vasculopathy and decreased blood flow analogous to other conditions
known to be associated with vasculopathy and leg ulcers, such as diabetes
mellitus. Hemoglobin desaturation decreases arterial oxygen content and
could compromise oxygen delivery to the skin.
A measurable TRV, a noninvasive estimate of pulmonary arterial pressure,
was obtained in 478 subjects. SCD subjects with leg ulcer history had a sig-
nificantly higher prevalence of elevated TRV: TRV ? 2.5 m/sec was present
in 59% of the subjects in the leg ulcer group and in 43% of the group with-
out leg ulcer history (P 5 0.006), and a TRV ? 3.0 m/sec, in 20% versus
13% (P 5 0.004), for subjects with and without leg ulcers history, respec-
tively. NT-proBNP is a widely used clinical laboratory marker of left ventricu-
lar stress that is elevated in adults with right ventricular stress because of
pulmonary hypertension associated with SCD and other disorders [20,21]
and is associated with early mortality in adults with SCD. In this study, NT
proBNP was significantly higher in subjects with leg ulcers (P < 0.0001). An
increase in NT-proBNP has been associated with markers of hemolysis in
SCD and other chronic anemias, with improvement after therapy , there-
fore it is possible that the higher NT-proBNP in the leg ulcer group reflects
higher hemolysis. We also plan to investigate the possibility that diastolic-
associated venous congestion contributes to leg ulceration.
We evaluated the frequency of self-reported lifelong clinical complications
in SCD subjects with leg ulcers. Clinical parameters included history of
stroke or transient ischemic attack, headaches, avascular necrosis of bone,
asthma, acute chest syndrome, cardiovascular dysfunction, priapism, kidney
disease, and frequency of hospitalization for pain. None of them were signifi-
cantly associated with leg ulceration history. Interestingly, the results of a
prior cooperative study for sickle cell disease leg ulcer study show concord-
ance with our results obtained using self-reported history . The qualita-
tively very similar results support the self-reported approach.
Of the 505 subjects in our cohort, 448 who enrolled on or before 12/31/2008
were followed for mortality (89%). Sixty-four of these subjects (14 %) had died
by the time of analysis. Mean follow up was 54.3 months (25th and 75th per-
centiles: 24 and 85 months, respectively). Kaplan–Meier survival curves did
not show a difference in survival between groups. Age, TRV, and serum ferritin
were the independent predictors of death in our population, Table III.
In conclusion, we demonstrate that leg ulcers are still a relatively common
and serious complication of sickle cell disease, with 21% of HbSS patients
reporting an ulcer in the past or having an active ulcer, similar to the 25%
incidence reported by Koshy over two decades ago . Leg ulceration in
patients with SCD is associated with markers of hemolytic severity, cardio-
vascular risk, and surrogate markers of pulmonary hypertension. These epi-
demiologic data support the model of a ‘‘leg ulcer phenotype’’ originally pro-
posed by Ballas, revised by Alexander et al. and subsequently by our group
suggesting an overlap of the multifactorial pathobiological mechanisms of
leg ulcer and vasculopathy [4,23,24].
A new observation that emerged in our analysis is the association of
increased levels of uric acid with history of chronic leg ulcers. This associa-
tion was considerably stronger than the one observed with creatinine or
blood urea nitrogen (BUN), thus suggesting that it was not merely a nonspe-
cific reflection of renal dysfunction. It is known that in patients without SCD,
uric acid is a possible cause of hypertension and a marker for cardiovascu-
lar disease, pulmonary hypertension, and early mortality [25–28]. High uric
acid may be a consequence of higher hemolysis because of the high hema-
topoietic turnover. The evolving evidence for uric acid as a risk factor for
vasculopathy is consistent with our association between uric acid and leg
ulceration as part of a vasculopathic complex in SCD.
Therapies of limited efficacy abound for chronic sickle cell leg ulcers, but
more uniformly effective therapy is needed.
Five hundred and twenty-five SCD adult subjects were prospectively
enrolled in a National Heart, Lung and Blood Institute (NHLBI)-approved pro-
tocol (Clinical Trial.gov no. NCT00011648) from February 2001 to March
2010. All patients provided written informed consent before enrollment. Our
analysis included 505 (96%) of these subjects based on the availability of
either a history of leg ulcers or an assessment of active leg ulcers at enroll-
ment. A preliminary analysis of the first 325 subjects was presented in part
at the Fourth Annual Sickle Cell Disease Research and Educational Sympo-
sium & Annual National Sickle Cell Disease Scientific Meeting (Hollywood,
FL) and published in its meeting report . This is an updated and
extended analysis of an expanded cohort. Data prospectively collected
included, a detailed past medical history and physical examinations, echo-
cardiographic imaging, laboratory analyses, and pulse oximetry measure-
ments. Echocardiography was performed as previously described . Labo-
ratory analysis included complete blood count, chemistries, serum LDH, and
NT-proBNP. Subjects were evaluated while at steady state, at least 2 weeks
after any type of acute exacerbation of SCD. Patients were followed for a
median of 52.3 months (25th and 75th percentiles: 24 and 84 months).
The diagnosis of sickle cell disease genotype was made by high pressure
liquid chromatography (HPLC) and confirmed by DNA sequencing in the first
270 subjects, where it was questionable . The presence of coincident a-tha-
lassemia (a3.7deletion) was determined in 294 patients as previously
Statistical Analysis. Clinical and laboratory characteristics were com-
pared between subjects with active ulcers or a history of leg ulceration and
subjects without a history of ulcers using the nonparametric Wilcoxon rank
sum test for continuous variables and Pearson’s Chi-square statistic for cate-
gorical variables. P values less than 0.05 were considered significant. TRV
was examined by categorizing subjects into groups based on a cut-off of
two standard deviations above the mean (?2.5 m/sec) and a cut-off of
approximately three standard deviations above the mean (?3.0 m/sec).
TABLE II. Independent Associations of Clinical and Laboratory Characteris-
tics with Leg Ulcer History in Logistic Regression Analysis
Independent variableOR (95% CI)a
aFor continuous variables, odds ratio is given for the 75th relative to 25th percen-
tile and is adjusted for all other covariates in the model.
TABLE III. Cox Proportional Hazards Regression Analysis of Mortality
among Sickle Cell Patients, Adjusted for Leg Ulcer History
Independent variableHazard ratio (95% CI)a
TRV ? 3.0
History of leg ulcers
aFor continuous variables, hazard ratio is given for the 75th relative to 25th per-
American Journal of Hematology707
Logistic regression models were used to investigate associations of a vari-
ety of patient characteristics with active or past ulcers. Covariates were log-
transformed as necessary to normalize their distributions and to reduce the
influence of outlying values. Significance of individual predictors was deter-
mined using the Wald Chi-square statistic, retaining as significant variables
with P values less than 0.05.
Associations of leg ulcer history with mortality were examined using data
from 448 subjects for whom follow-up information was available and whose
enrollment date was on or before 12/31/2008. Subjects, who were not known
to be dead, were censored at the date of last contact with study staff. Kaplan–
Meier survival curves were calculated to examine mortality over time among
patients with a history of leg ulcers compared with patients without a history of
leg ulcers, using the log-rank test and a P value of 0.05 to determine signifi-
cant differences in mortality between the two groups. Cox Proportional Haz-
ards regression models were used to examine the association of leg ulcer his-
tory with mortality while adjusting for significant covariates as determined in
prior mortality analyses of this cohort. The likelihood ratio test was used to
determine the significance of individual regression coefficients. All analyses
were performed using SAS version 9.1.3 (SAS Institute, Cary, NC) and Stata
version 9.0 (StatCorp LP, College Station, TX).
Caterina P. Minniti wrote the manuscript, designed the research and ana-
lyzed the data. James G. Taylor VI enrolled subjects, performed laboratory
experiments and collected the data. Mariana Hildesheim analyzed the data
and prepared the tables. Patricia O’Neal enrolled subjects. Jonathan Wilson
analyzed data and prepared figures. Oswaldo Castro enrolled subjects and
contributed to the design and interpretation of the data. Victor R. Gordeuk
enrolled subjects, contributed to the interpretation of the data and to the writ-
ing of the manuscript. Gregory J. Kato designed the research, contributed to
the analysis of the data and to the writing of the manuscript. The authors
are grateful to Mary K. Hall, CIP for expert protocol management. We thank
Darlene Allen for data collection and Britny Hall for manuscript preparation.
We thank all the patients who participated in this study.
1Cardiovascular and Pulmonary Branch, Sickle Cell Vascular Disease Section,
NHLBI, National Institutes of Health, Bethesda, Maryland;2Center for Sickle Cell
Disease, Department of Medicine, Howard University, Washington, District of
Contract grant sponsor: Division of Intramural Research (NIH, NHLBI)
*Correspondence to: Caterina P. Minniti, National Institutes of Health, National
Heart, Lung, and Blood Institute, Pulmonary and Vascular Medicine Branch,
Building 10CRC Room 5-5140, Bethesda, MD 20892
Conflict of interest: No conflicts of interest.
Published online 27 April 2011 in Wiley Online Library
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708 American Journal of Hematology
Myeloablative allogeneic stem cell transplantation for
lymphoblastic lymphoma in Sweden: A retrospective study
Vladimir Lj Lazarevic,* Mats Remberger,Hans Ha ¨gglund,Helene Hallbo ¨o ¨k,Gunnar Juliusson,Eva Kimby,
Claes Malm,Anders Wahlin,Hamdy Omar and Jan-Erik Johansson
The World Health Organization from 2008 classifies B- or T-cell lympho-
blastic leukemia/lymphoma as a precursor lymphoid neoplasm . Conven-
tionally, when a lymph node mass is present and 25% or more of the
nucleated cells in the bone marrow are lymphoblasts, a diagnosis of lympho-
blastic leukemia is preferred to lymphoblastic lymphoma. Data about the
patients (?16 years of age), who had undergone allogeneic stem cell trans-
plantation (ASCT) due to T- and B-cell lymphoblastic lymphoma (LBL) in
Sweden, were collected from patient files, local registries, and the The Euro-
pean Group for blood and marrow transplantation (EBMT) data base in all
participating centers (Stockholm, Gothenburg, Lund, Linko ¨ping, Uppsala,
and Umea ˚). Nineteen patients were transplanted in four Swedish centers
(Stockholm, Uppsala, Lund, and Gothenburg) between August 1987 and
February 2005. A patient who underwent syngeneic stem cell transplantation
and two others who were transplanted with reduced intensity conditioning
regimens were excluded from this study. All the patients had a lymph node
biopsy-proven histologic diagnosis of a precursor B or T lymphoblastic lym-
phoma. The cut-off value of 30% for blast cells in the bone marrow was
used for distinguishing between leukemia and lymphoma, because the
majority of patients were diagnosed before 2004. The study protocol was
approved by the Regional Ethical Review Board in Gothenburg and the
patients gave their informed consent before transplantation. They were
treated according to different chemotherapy protocols (Wollner protocol,
NOPHO, HD Ara-C and Mitoxantrone, Riehm, HyperCVAD, DexaBEAM,
Swedish National therapy program for adult ALL, and Norwegian national
program for ALL). Decisions about the transplantations were made, respec-
tively, at the transplantation centers. Among the patients in complete remis-
sion (CR), only one patient was transplanted in CR2. human leurocyte anti-
gen (HLA)-identical related donors were used in 14 patients (73%), HLA-A,
B, and Dr locus (DR) identical unrelated donors were used in four patients
(23%) and mismatched unrelated in one patient (4%). Other initial patient
characteristics are shown in detail in Table I. Probabilities of overall survival
and relapse-free survival were estimated using Kaplan–Meier method and
compared with the log-rank test . The incidence of graft versus hoct dis-
ease (GVHD), transplantation related mortality (TRM), and relapse were
estimated using a cumulative incidence curves estimator . All the tests
were two sided. Analyses were performed using a cmprsk package (devel-
oped by Gray, June 2001), Splus 6.2 software (S-plus 6.2, Insightful, Seattle,
WA) and Statistica software (Statistica, StatSoft, Tulsa, OK). Transplantation
outcome of the 19 patients is presented in Table I. Acute GVHD Grades II–
IV developed in six patients with a cumulative incidence of 32%. The inci-
dence of acute GVHD was higher in patients who received peripheral-blood
stem cells (PBSC) than bone marrow (BM) (67% vs. 15%; P 5 0.02).
Chronic GVHD developed in 4/16 patients with a cumulative incidence of
25% within 2 years. The cumulative incidence of TRM at 1 and 3 years was
32%. The incidence of TRM was higher in the unrelated donor transplants
than in the sibling transplants (80% vs. 14%, P 5 0.002) and in T- compared
with B-LBL (42% vs. 0%; P 5 0.06, ns). The cumulative incidence of relapse
was 42%. The incidence of relapse was higher in the patients with >1 year
between diagnosis and ASCT than in the patients with ASCT <1 year after
diagnosis (71% vs. 25%, P 5 0.06, ns). The 5-year overall survival (OS)
and the progression-free survival (PFS) were 30% and 32%, respectively.
The OS and PFS were longer in the patients who were transplanted <1
year after the diagnosis versus patients transplanted >1 year after diagnosis
(P 5 0.06, ns). There are not many reports of the results of allogeneic stem
cell transplantation for lymphoblastic lymphoma in the literature. We report
the results of the patients who were transplanted at the time when lympho-
blastic lymphoma was treated predominantly as high-grade lymphoma rather
than ALL. All the patients were transplanted in chemosensitive disease,
most in CR1 and with myeloablative conditioning regimen, preferentially with
cyclophosphamide and TBI. The incidence of acute and chronic GVHD does
not differ significantly from that of other studies done in similar circumstan-
ces. The high incidence of TRM, especially in the matched unrelated donor
(MUD) transplants, and the high relapse risk (42%) contribute to low PFS
and OS. The results of our study are comparable to previous studies in lym-
phoblastic lymphoma [4–6]. The PFS and OS were almost identical to the
previous report of the Swedish ALL group . However, our study shows
better results for the patients transplanted <1 year after the diagnosis com-
pared with those transplanted >1 year after diagnosis. A year after trans-
plant, the survival curve reaches plateau. To analyze whether there was a
time related outcome improvement, we divided the patients into two groups:
those transplanted between 1987 and 1998 and those transplanted between
1998 and 2005. Surprisingly, the group transplanted between 1987 and
TABLE I. Myeloablative Allogeneic Transplantation for Lymphoblastic Lymphoma in Sweden 1987–2005
Dg-Tx < 1
SCT ATGaGVHD cGVHD
American Journal of Hematology 709
1998 had a significantly superior PFS and OS compared with later trans-
plants (P 5 0.01) (Fig. 1). Upon a closer look at the two groups we noticed
that most of the patients (91%) transplanted before 1998 were transplanted
<1 year after the diagnosis. Only 25% of the patients transplanted after
1998 were transplanted a year or longer after the diagnosis (Table I). The
patients transplanted between 1987 and 1998 had received less chemother-
apy before the transplantation. Another contributor to the better results of
this group may be the fact that more patients were in CR at ASCT and that
fewer received PBSC than in the other group. The use of PBSC led to more
acute GVHD and higher TRM. Furthermore, among those transplanted
between 1998 and 2005, more patients had unrelated donors. In this group,
there were significantly more acute GVHD and TRM compared with that of
the matched related donors. Although this group is profoundly heterogene-
ous, this retrospective analysis confirms that a selection of patients and
donors plays a crucial role for the outcome. We have shown that myeloabla-
tive allogeneic stem cell transplantation for lymphoblastic lymphoma led to a
long-term survival in a third of the patients. Finally, when the transplantation
was done <1 year after the diagnosis in CR, using matched sibling donors
and BM as a stem cell source, with fewer chemotherapy cycles before the
transplant, better results for ASCT were achieved.
Department of Hematology
University Hospital Lund
Lund University, Lund, Sweden
*Correspondence to: Vladimir Lj Lazarevic, Department of Hematology, University
Hospital Lund, Lund University, Lund SE-221 38, Sweden
Conflict of interest: Nothing to report.
Published online 5 May 2011 in Wiley Online Library
1. World Health Organization. In:Swerdlow ,SH,Campo ,E,Harris ,NL, et al., edi-
tors. Classification of Tumours of Haematopoietic and Lymphoid Tissues.
Lyon:IARC Press; 2008
2. Kaplan E, Meier P. Nonparametric estimation from incomplete observations. J
Am Stat Assoc 1958;53:457–481.
3. Gooley T, Leisenring W, Crowley J, Storer B. Estimation of failure probabilities
in the presence of competing risks: New representations of old estimators.
Stat Med 1999;18:695–706.
4. Levine JE, Harris RE, Loberiza FR Jr, et al. A comparison of allogeneic and
autologous bone marrow transplantation for lymphoblastic lymphoma. Blood
5. Bouabdallah R, Xerri L, Bardou VJ, et al. Role of induction chemotherapy and
bone marrow transplantation in adult lymphoblastic lymphoma: A report on 62
patients from a single center. Ann Oncol 1998;9:619–625.
6. Goldstone AH, Richards SM, Lazarus HM, et al. In adults with standard-risk
acute lymphoblastic leukemia, the greatest benefit is achieved from a
matched sibling allogeneic transplantation in first complete remission, and an
autologous transplantation is less effective than conventional consolidation/
maintenance chemotherapy in all patients: Final results of the International
ALL Trial (MRC UKALL XII/ECOG E2993). Blood 2008;111:1827–1833.
7. Hallbo ¨o ¨k H, Ha ¨gglund H, Stockelberg D, et al.; Swedish Adult ALL Group. Autol-
ogous and allogeneic stem cell transplantation in adult ALL: The Swedish Adult
ALL Group experience. Bone Marrow Transplant 2005;35:1141–1148.
patients with lymphoblastic lymphoma transplanted (1987–1998) and (1999–2005).
Overall survival after myeloablative conditioning and ASCT in 19
Acquired factor V inhibitor complicating warfarin therapy
Factor V is a procofactor that exists in both the plasma (80%) and within
platelet alpha-granules (20%) . Factor V is converted into its active
form by thrombin. Factor Va serves as cofactor for factor Xa in the pro-
thrombinase complex, which forms on the platelet surface and produces
thrombin via limited proteolysis of prothrombin. Acquired factor V inhibi-
tors are rare with an annual incidence reported between 0.09 and 0.29
cases per million persons [2,3]. Factor V inhibitors may be alloantibodies
that develop following exposure to bovine thrombin-derived fibrin seal-
ant . In patients exposed to topical thrombin preparations, the inci-
dence of factor V inhibitors may be significantly higher . Factor V
inhibitors may also develop as autoantibodies with a number of reported
associations including antibiotics [6,7], sepsis , malignancies , and
autoimmune illnesses [10–12]. The clinical presentation of factor V inhib-
itors ranges from abnormal coagulation studies with no associated
bleeding to life threatening hemorrhage. Here, we report an unusual case
of a factor V inhibitor developing in an elderly woman anticoagulated
with warfarin who had persistently abnormal coagulation studies after its
discontinuation. This raises significant issues regarding differential diag-
nosis, work-up, and therapeutic intervention.
An 80-year-old African-American woman was admitted to our hospital with
respiratory complaints and was found to have abnormal coagulation. Past
medical history was significant for diabetes mellitus, chronic renal insuffi-
ciency, coronary artery disease, heart failure, and atrial fibrillation for which
she was anticoagulated with warfarin.
The patient had recent hospital admissions for a suspected gastrointesti-
nal bleed, lower extremity cellulitis, and hypoglycemia. Endoscopy revealed
no gastrointestinal bleeding. A course of doxycycline and cephalexin was
given for cellulitis during the second hospitalization. There was no surgery
or exposure to bovine thrombin during these hospitalizations. At the time of
the third discharge, which was 1 month after the episode of cellulitis and the
exposure to antibiotics, the prothrombin time (PT) was 16.2 s (normal, 11.5–
15.5 s) and the international normalized ratio (INR) was 1.3. The patient
was discharged on warfarin and 24 days later, as an outpatient, the PT was
noted to be 75.2 s. Warfarin was held and vitamin K was given, but the PT
remained 59.2 s 20 days later.
Forty-seven days following her most recent discharge, the patient was
readmitted with dyspnea. At the time of admission, the PT was noted to be
54.6 s with an INR of 5.8. The activated partial thromboplastin time (APTT)
was found to be >150 s (normal, 23.6–35.7 s). The platelet count was 169
3 103/mL. She was treated with antibiotics, steroids, diuretics, and broncho-
dilators. Her respiratory status worsened and intubation and mechanical
ventilation were required on the third hospital day. Although vitamin K defi-
ciency usually prolongs the PT more than the APTT, the coagulopathy was
initially attributed to depletion of vitamin K-dependent factors secondary to
antecedent warfarin use. A total of 10 mg of Vitamin K was given orally and
another 10 mg given subcutaneously without significant improvement in the
PT or APTT. There was no bleeding.
710 American Journal of Hematology
There was no evidence of disseminated intravascular coagulation. The D-
dimer was 0.31 mcg/mL (normal, <0.5 mcg/mL), and the fibrinogen was
334 mg/dL (normal, 175–450 mg/dL. A mixing study was ordered (Table I).
Neither the PT nor the APTT corrected suggesting a circulating anticoagu-
lant. Factor levels were obtained (Table II). The factor V level was 3%
(normal, 50–150%). Factor II was 68% (normal, 50–150%). Factors VII, VIII,
and X demonstrated circulating anticoagulant effect with recovery to normal
levels with serial dilutions (from 16, 65, and 22% to 60, 205, and 66%
respectively). The factor V inhibitor level was found to be 17 Bethesda units/
mL. The thrombin time was 20.6 s (normal, <21.0 s). A review of all recent
hospitalizations was conducted, and there was no surgical procedure or
exposure to bovine thrombin. Interestingly, the mixing study suggested time
dependence as the PT and APTT were longer after 1 h of incubation. Time
dependence is often associated with factor VIII inhibitors but has been
reported with factor V inhibitors as well .
There was no bleeding but a recommendation was made to transfuse one
unit of single donor platelets before invasive procedures. This recommenda-
tion was based on the use of platelet transfusions to control active bleeding
associated with acquired factor V inhibitors . Using this strategy, central
venous catheters were placed without bleeding.
For inhibitor suppression, prednisone was initiated at 1 mg/kg daily. The
APTT improved from 134.9 to 115.1 s and the PT from 50.6 to 36.8 s after
13 days of prednisone. Unfortunately, the patient died of sepsis before a
repeat factor V level was obtained.
The development of factor V inhibitors, in the absence of bovine thrombin
exposure, has been reported only rarely in the medical literature. In a review
by Kno ¨bl and Lechner, 105 cases were reported over a greater than 40-year
period. Bleeding manifestations occurred in 60% of these patients with surgi-
cal site bleeding, hematuria, gastrointestinal bleeding, and hematomas being
the most common hemorrhagic complications. Only 32.6% of patients with
alloantibodies following exposure to bovine thrombin developed bleeding.
There was a statistically significant association between residual factor V
levels and bleeding .
In another review by Ang et al. in which patients with factor V inhibitors
acquired after exposure to bovine thrombin were not included, 73 cases
were identified from 1950 to 2008 plus an additional three newly reported
cases. Of these patients, 68.4% had documented bleeding with the mucous
membranes being the most common site. Intracranial bleeds occurred in
14% of patients. Fatal bleeding events occurred in 12% of patients. All
cause mortality was 31%. There was a nonsignificant trend toward lower
factor V levels and a higher PT in bleeders versus nonbleeders and a statis-
tically significant prolongation in APTT in bleeders versus nonbleeders.
There was no association between inhibitor level and bleeding risk . This
is reminiscent of the lack of association between the titre of factor VIII inhibi-
tors and bleeding risk .
The optimal treatment of bleeding in patients with factor V inhibitors is
not known. The rarity of the disease has precluded a rigorous evaluation
of treatment strategies. Fresh frozen plasma and prothrombin complex
concentrates (PCC) have generally not been effective in controlling bleed-
ing . A recent report demonstrated the successful use of recombinant
activated factor VII to control bleeding due to a factor V inhibitor .
Similarly, activated PCC has also been used successfully in a factor V
Platelet transfusions have been successfully used to treat active bleeding
caused by factor V inhibitors . Factor V on the platelet surface may serve
as a source of factor V that is protected from inhibitors. This may explain
the utility of platelet transfusions in these patients . One interesting study
evaluated the potential mechanism of bleeding in patients with factor V inhib-
itors. The inhibitors were found to target the C2 domain in all cases but
interfered with prothrombinase activity only in those cases associated with
In the review by Kno ¨bl, the median duration of factor V inhibitors was 9.7
weeks. Those inhibitors associated with known precipitants all resolved by 1
year and had a median duration of 8 weeks. In contrast, idiopathic factor V
inhibitors had only a 61.7% probability of resolution after a median of 22.9
weeks . Various methods have been used to suppress factor V inhibitors
including corticosteroids [9,20], cyclophosphamide , and rituximab .
However, given the transient nature of some factor V inhibitors in the
absence of suppression therapy and the rarity of this condition, it is prema-
ture to evaluate the true benefit of these strategies. In fact, Kno ¨bl et al.
found no difference in terms of disappearance of the inhibitor or survival in
those who received suppressive therapy compared with those who did not
. As with other immune-mediated bleeding disorders, rapid inhibitor
clearance has been achieved with the use of intravenous immunoglobulin
 and plasma exchange .
This is an informative case of an idiopathic factor V inhibitor developing in
an elderly woman on warfarin. She developed strikingly abnormal coagula-
tion studies in the outpatient setting with prolongation of the PT and APTT.
She had no exposure to bovine thrombin, surgery, or a history of malig-
nancy. There was an exposure to antibiotics, but the INR was 1.3, 1 month
following this exposure. The differential diagnosis for combined prolongation
of PT and APTT includes vitamin K deficiency, liver dysfunction, warfarin
use, congenital factor deficiency, the development of an acquired inhibitor of
coagulation, disseminated intravascular coagulation, and a lupus anticoagu-
lant. This case of a factor V inhibitor exemplifies the necessity to consider
other causes of coagulopathy in patients on warfarin when the coagulopathy
does not respond to cessation of warfarin and supplementation with vitamin
K. In such situations, as in this case, the differential diagnosis of abnormal
studies of coagulation must be revisited.
Division of Hematology/Oncology, Department of Medicine, The Tisch Cancer
Institute, The Mount Sinai School of Medicine, New York
*Correspondence to: Benjamin Gartrell, MD, Mount Sinai School of Medicine, One
Gustave L. Levy Place, Box 1079, New York, NY 10029
Conflict of interest: Nothing to report.
Published online 5 May 2011 in Wiley Online Library
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human plasma and platelets. Blood 1982;60:59–63.
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TABLE I. Result of Mixing Studies
Patient plasma (s)
Patient 1 normal plasma: immediate (s)
Patient 1 normal plasma: 1 h (s)
TABLE II. Coagulation Studies
Patient values Reference value
Factor II (%)
Factor V (%)
Factor VII (%)a
Factor VIII (%)a
Factor X (%)a
Thrombin time (s)
aThese factors demonstrated circulating anticoagulant effect and these values rep-
resent the values after 1/40 dilution.
American Journal of Hematology711
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ment of human factor V in a patient with Hashimoto’s disease and bullous
pemphigoid. Thromb Res 1995;77:63–68.
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21. de Raucourt E, Barbier C, Sinda P, et al. High-dose intravenous immunoglo-
bulin treatment in two patients with acquired factor V inhibitors. Am J Hematol
Intra-arterial methylprednisolone for the management of
steroid-refractory acute gastrointestinal and hepatic
graft versus host disease
Laurie A. Milner,1,2,3* Michael W. Becker,2Steven H. Bernstein,2Lauren Bruckner,3
Jonathan W. Friedberg,2George A. Holland,4J.J. Ifthikharuddin,2Jane L. Liesveld,2Edward J. Mathes,4
Heather L. Menchel,5Craig A. Mullen,3Talia Sasson,4and Gordon L. Phillips II2
Steroid-refractory or dependent (SR/D) acute graft versus host disease
(aGVHD) involving the gastrointestinal (GI) tract or liver is a dire com-
plication of allogeneic hematopoietic stem cell transplantation (HSCT)
for which there is no standard therapy [1–3]. Most agents utilized are
highly immunosuppressive and thus associated with high rates of
opportunistic infections . An alternative approach involves direct
infusion of corticosteroids into mesenteric and/or hepatic arteries .
While the mechanism of action remains speculative, a few small series
suggest reasonable efficacy with this approach [5–10]. Here we
describe a retrospective analysis of intra-arterial (IA) methylpredniso-
lone (MePDSL) treatment of 13 patients with SR/D aGVHD involving the
GI tract, including four with concurrent liver GVHD. Various combina-
tions of the celiac, superior and inferior mesenteric, and common hep-
atic arteries were injected with MePDSL, generally 60 mg/vessel, on a
median of 3 consecutive days. There were no major procedural compli-
cations. Six patients experienced GI complete responses (CRs) and
two experienced partial responses. One of four patients with liver
GVHD achieved a durable CR. Three patients remain alive and well.
These results were obtained in a highly refractory group of patients;
we suggest that better results might be obtained with earlier and more
standardized IA therapy.
Acute GVHD is a common complication of allogeneic HSCT that can
result in significant morbidity and mortality [11,12]. Involvement of the GI
tract with aGVHD is particularly problematic; in addition to having a lower
response rate than GVHD involving other systems, compromised GI function
prevents adequate oral nutrition and provides an obvious portal for infection
. When primary therapy with corticosteroids is inadequate, the adminis-
tration of additional highly immunosuppressive agents often contributes to
fatal opportunistic infections. As an alternative, the use of MePDSL infused
into regional mesenteric and/or hepatic arteries offers the potential to direct
therapy to sites of involvement, thus minimizing systemic effects and secon-
dary complications, thereby improving the long-term outcomes of these
The precise mechanism by which IA MePDSL is effective for ‘‘steroid-
refractory’’ disease remains unknown. It is possible that a higher regional con-
centration of glucocorticoid has an enhanced effect on the modulation of local
cellular immune response and production of proinflammatory cytokines. It has
also been postulated that a high local concentration of steroid overcomes
refractoriness caused by down-regulation of steroid receptors in the inflamed
GI mucosa of patients with severe GI GVHD . Evidence for such a mecha-
nism comes largely from studies in patients with inflammatory bowel disease,
which demonstrate that glucocorticoid receptors are down-regulated in
inflamed intestinal mucosa and that this finding correlates with steroid refrac-
During the inclusive period of 26 April 2007 and 15 July 2010, we treated
13 patients with SR/D GI GVHD with IA MePDSL. Four patients had concur-
rent liver GVHD. Patient demographics, GVHD prophylaxis, and GVHD treat-
ment before, and concurrently with, the initiation of IA MePDSL injections,
are summarized in Table I. Details regarding IA MePDSL administration and
results are summarized in Table II. The 13 patients were treated on 15 occa-
sions, comprising a total number of IA injections of 106. Two patients
received injections of IA MePDSL on two separate occasions, patient 1 five
months after the initial treatment and patient 2 two weeks later. All patients
except one received all IA injections planned; the IMA could not be cannu-
lated on one occasion in a small child (patient 1).
As indicated in Table II, MePDSL was given at a flat dose of 60 mg/
vessel in 10 of the 13 patients; two pediatric patients received doses of 1
to 2 mg/kg/vessel and one adult patient received 50 mg/vessel. Most fre-
quently (seven patients), MePDSL was injected into the three main mes-
enteric vessels (celiac, superior, and inferior mesenteric arteries). Five
patients received injections into the common hepatic artery, which also
feeds the celiac axis; when the common hepatic artery was injected, a
separate injection into the celiac artery was deferred. The majority of
patients (eight) received injections on 3 consecutive days. Two pediatric
patients received injections on a single day, one due to clinical concerns
and one due to parental request. One patient received injections on 4
Consistent with previous reports, we found IA infusion of MePDSL to
be safe. Minor complications included three hematomas that did not
require transfusion, one episode of gastrointestinal bleeding that subsided
with supportive care, and one episode of abdominal pain and bacteremia
that resolved with appropriate therapy. One patient suffered a mild myo-
cardial infarction a few days after the procedure; this event was felt to be
unrelated to the procedure. While there were no serious immediate com-
plications, the relative contribution, if any, of this therapy to more delayed
and/or systemic complications (e.g., infection) is more difficult to ascer-
tain. Of particular note in this regard, all but one of our patients received
monoclonal antibody therapy (generally infliximab) either before, or con-
currently with, IA MePDSL. This additional immunosuppressive therapy,
as well as the inherently high mortality rate associated with severe SR/D
712 American Journal of Hematology
GI GVHD, makes it virtually impossible to determine whether or not treat- Download full-text
ment with IA MePDSL contributed to any of the adverse outcomes we
For assessing therapeutic response, we chose only two parameters:
achievement of a CR of GI (and hepatic) signs and symptoms by day 28 fol-
lowing IA treatment and overall survival. Even so, responses may have been
over- or underestimated; to some degree this is inherent in grading aGVHD
of the GI tract, as the signs and symptoms can be affected by other inciting
factors, as well as ancillary therapies. In addition to limiting response param-
eters, we also applied a relatively stringent definition of response: the
absence of GI symptoms while tolerating enteral nutrition and medications.
Of note in this regard, many of our patients were acutely ill at the time of IA
MePDSL treatment, making the assessment of response problematic.
Indeed, two patients died prior to day 128 and four additional patients who
died between day 129 and 143 were so unwell that accurate retrospective
assessments of response were not completely feasible.
Six of our 13 patients had GI CRs. Two have had durable CRs: one is 8
months postprocedure, tapering immunosuppressive therapy; the other is
more than 3 years postprocedure and off all immunosuppression. One patient
(1) had a CR following a single IA infusion, but then suffered a recurrence of
GVHD; he received another infusion 5 months after the first, resulting in a PR.
He survived for 15 months before succumbing to an infection. Two patients
had resolution of GI symptoms at day 128, but died of infection shortly there-
after. One patient had a GI CR, but subsequently developed progressive skin
GVHD and liver dysfunction and died from a complication of therapy. Of the
remaining seven patients, three had no response, two had PRs, and two died
before day 128 and were therefore not evaluable. Of the eight patients achiev-
ing less than a CR, one who achieved a good PR is doing well 8 months post-
IA MePDSL, tapering off immunosuppression. Four patients received hepatic
IA MePDSL for liver GVHD. One of these patients achieved a durable CR, one
had no response, and two expired before day 128.
While the responses and outcomes we observed may be considered mod-
est, our patients were generally treated late in the disease process, dimin-
ishing the likelihood of a good response or of long-term survival. The two
patients who were treated relatively early in the disease process both had
good responses and remain alive and well. Patient 4 received IA MePDSL
as second line therapy and is now more than 3 years post treatment, off all
immunosuppression. Patient 13 was treated with IA MePDSL soon after
institution of other second line agents; he achieved both GI and hepatic CRs
and remains well 8 months post therapy. All of our other patients had
received two to four lines of prior therapy and were receiving multiple other
agents concurrently with IA MePDSL.
Considering our results, as well as those of others, we believe this thera-
peutic strategy warrants follow-up studies. Optimizing patient selection and
administration of IA therapy to permit the aggressive tapering of systemic
immunosuppression would likely be a crucial to the success of such studies.
One approach would be to incorporate IA MePDSL into the primary treat-
ment of aGVHD therapy in selected patients; even if this resulted in treat-
ment of some patients destined to respond to standard therapy, it is possible
that production of a more rapid CR would allow earlier discontinuation of
PatientTransplant day Vessels injectedDose MP mg/vessel Response GI, d28Response liver, d28Current status
C1, SM1, IM1
C3, SM3, IM3
C3, SM3, IM3
C2, SM3, IM3, CH1
C3, SM3, IM3
SM3, IM3, CH3
SM3, IM3, CH3
C3, SM3, IM3
C3, SM3, IM3
SM4, IM4, CH2
SM3, IM3, CH3
1 mg/kg 5 11
2 mg/kg 5 20
1 mg/kg 5 30
1 mg/kg 5 30
Died, d 1444; infection, GvHD
Died, d 1790; infection; min IST
Died, d 174; complic of therapy
Alive, >d 11200; off IST, no GvHD
Died, d 136; sepsis
Died, d 143; GvHD, infection
Died, d 132; GvHD, infection
Died, d 111; relapse, GvHD
Died, d 1370; GvHD relapsed AML
Alive; d 1240 tapering IST
Died, d 129; infection
Died, d 116; bleeding, infection
Alive, >d 1200; tapering IST
aNot applicable: no liver GvHD when treated with IA MePDSL.
bCR by GI biopsy, but no change in symptoms in setting of multiple infections (including CMV).Vessels (number of injections for each vessel follows letter designation): C,
celiac; SM, superior mesenteric; IM, inferior mesenteric; LH, left hepatic; RH, right hepatic; CH, common hepatic. Response: CR, complete response; PR, partial
response; NR, no response; NE, not evaluable. IST: immunosuppressive therapy.
TABLE I. Patient Demographics and Systemic Therapy before Intra-Arterial Methylprednisolone
Patient Age (yrs)Sex DxConditioning regimenTransplant type GVHD prophylaxisGI GVHD grade Systemic Tx before IA MPa
Therapy w/IA MP
1 1½M SCIDNMA 6/6 UCB CsA, MMF III
29M HLH MA10/10 MUD CsA, MTXEtan,Inflix,Ritux
aAll patients received methylprednisolone, MMF, and a calcineurin inhibitor; denoted are additional agents. Conditioning regimen: MA, myeloablative; NMA, non-myeloa-
blative, RIC, reduced intensity. Transplant type: UCB, umbilical cord blood; MUD, matched unrelated donor (degree of HLA match denoted); MSD, matched sibling. GvHD
prophylaxis and treatment: MMF, mycophenolate mofetil; Tac, tacrolimus; IA, intra-arterial; MP, methylprednisolone; Etan, etanercept; inflix, infliximab; dacluz, dacluzu-
mab; Ritux, rituximab; siro, sirolimus. Dx, diagnosis; HLH, hemophagocytic lymphohistiocytosis; MDS, myelodysplastic syndrome; MM, multiple myeloma.
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