Whole blood EBV-DNA predicts outcome in diffuse large B-cell lymphoma

Abstract

An association between Epstein-Barr Virus (EBV) infection and lymphoproliferative diseases has been reported with EBV + diffuse large B cell-lymphoma (DLBCL) of the elderly described as a distinct entity. In a cohort of 218 human immunodeficiency virus (HIV)-negative patients with diffuse large B-cell lymphomas, we detected EBV-DNA in 25% of whole blood (WB) samples at diagnosis. Presence and viral load in WB, mononuclear cells or plasma did not predict the presence of EBV in the tumor biopsy. Positive Hepatitis C virus (HCV) serology was associated with a higher frequency of EBV in WB. Patients with EBV-DNA in WB had a significantly shorter progression-free (p = 0.02) and overall survival (p = 0.05) after immunochemotherapy with R-CHOP (Rituximab, Cyclophosphamide, Doxorubicin, Vincristine, Prednisolone). We conclude that detection of EBV in WB is not a surrogate marker for EBV-association in diffuse large B-cell lymphoma, however it associates with worse outcome.

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Leukemia & Lymphoma
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Whole blood EBV-DNA predicts outcome in diffuse
large B-cell lymphoma
Maria Chiara Tisi, Elisa Cupelli, Rosaria Santangelo, Elena Maiolo, Eleonora
Alma, Manuela Giachelia, Maurizio Martini, Silvia Bellesi, Francesco D’Alò,
Maria Teresa Voso, Maurizio Pompili, Giuseppe Leone, Luigi Maria Larocca &
Stefan Hohaus
To cite this article: Maria Chiara Tisi, Elisa Cupelli, Rosaria Santangelo, Elena Maiolo, Eleonora
Alma, Manuela Giachelia, Maurizio Martini, Silvia Bellesi, Francesco D’Alò, Maria Teresa Voso,
Maurizio Pompili, Giuseppe Leone, Luigi Maria Larocca & Stefan Hohaus (2015): Whole blood
EBV-DNA predicts outcome in diffuse large B-cell lymphoma, Leukemia & Lymphoma, DOI:
10.3109/10428194.2015.1072766
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LEUKEMIA & LYMPHOMA
2015; EARLY ONLINE: 1–7
http://dx.doi.org/10.3109/10428194.2015.1072766
ORIGINAL ARTICLE: CLINICAL
Whole blood EBV-DNA predicts outcome in diffuse large B-cell lymphoma
Maria Chiara Tisi
1
, Elisa Cupelli
1
, Rosaria Santangelo
2
, Elena Maiolo
1
, Eleonora Alma
1
, Manuela Giachelia
1
,
Maurizio Martini
3
, Silvia Bellesi
1
, Francesco D’Alo
`
1
, Maria Teresa Voso
1
, Maurizio Pompili
4
, Giuseppe Leone
1
,
Luigi Maria Larocca
3
* & Stefan Hohaus
1
*
1
Institutes of Hematology, Catholic University S. Cuore, Rome, Italy,
2
Institutes of Microbiology, Catholic University S. Cuore, Rome, Italy,
3
Institutes of Pathological Anatomy, Catholic University S. Cuore, Rome, Italy, and
4
Department of Internal Medicine, Catholic University S.
Cuore, Rome, Italy
ABSTRACT
An association between Epstein–Barr Virus (EBV) infection and lymphoproliferative diseases has
been reported with EBV + diffuse large B cell-lymphoma (DLBCL) of the elderly described as a
distinct entity. In a cohort of 218 human immunodeficiency virus (HIV)-negative patients with
diffuse large B-cell lymphomas, we detected EBV-DNA in 25% of whole blood (WB) samples at
diagnosis. Presence and viral load in WB, mononuclear cells or plasma did not predict the presence
of EBV in the tumor biopsy. Positive Hepatitis C virus (HCV) serology was associated with a higher
frequency of EBV in WB. Patients with EBV-DNA in WB had a significantly shorter progression-free
(p¼0.02) and overall survival (p¼0.05) after immunochemotherapy with R-CHOP (Rituximab,
Cyclophosphamide, Doxorubicin, Vincristine, Prednisolone). We conclude that detection of EBV
in WB is not a surrogate marker for EBV-association in diffuse large B-cell lymphoma, however
it associates with worse outcome.
KEYWORDS
Diffuse large B-cell
lymphoma, EBV, prognosis
HISTORY
Received 20 January 2015
Revised 7 July 2015
Accepted 10 July 2015
Introduction
Exactly 50 years have passed since Epstein and Barr
described a DNA virus in cultured lymphoblastic cells
from a patient with the African variant of Burkitt
lymphoma [1]. Subsequently, the evidence-base show-
ing association between lymphoma and EBV-infection
has increased [2–4]. EBV-infection is a driving oncogenic
event in lymphomas arising in states of immunodefi-
ciency [5], both acquired as in HIV-infection or as a
consequence of immunosuppressive therapy in patients
[6,7]. Defects in the immunological control of EBV
infection is an issue for Hodgkin lymphoma (HL) [8].
The HLA-A1 allele, which is associated with a reduced
cellular response to EBV, leads to an increased risk of
EBV-associated HL [9].
The most recent EBV-related lymphoma type that has
been included as a provisional entity into the WHO
classification of 2008 is the EBV + diffuse large B cell
lymphoma of the elderly [10]. Its frequency appears
to vary according to the geographic region, with
frequencies as high as 8–11% in East Asia, whilst
in Western countries the proportion of EBV + cases
appears to be less frequent, representing 2–4% of all
DLBCLs [11–19].
EBV-DNA can be found in the whole blood of patients
with EBV-associated lymphoproliferative diseases. Au
et al demonstrated that in patients with T/NK lymph-
oma, EBV-DNA is a biomarker to monitor for disease
activity at diagnosis and during therapy [20]. It has also
been shown that in HL, EBV is frequently present at high
levels in the plasma or serum of EBV-associated cases
[21–24]. In EBV-associated HL, EBV copy number
correlated with several parameters of prognostic rele-
vance, reflecting tumor burden and activity of disease.
Stage and IPS score inversely correlated to parameters
that are potential indicators of the immune status, as
lymphocyte counts and antibody titers for the latent EBV
nuclear antigen EBNA1 [23].
*The last two authors shared senior authorship.
Correspondence: Stefan Hohaus, Istituto di Ematologia, Universita’ Cattolica S. Cuore, L.go A. Gemelli, 1, 00168 Roma, Italy. Tel: +39 06
30154180. Fax: +39 06 35503777. stefan.hohaus@rm.unicatt.it
There is an accompanying commentary that discusses this paper. Please refer to the table of contents of the print issue in which this article
appears.
!2015 Taylor & Francis
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This data demonstrated a gap in the evidence-base,
indicating a need to investigate the role of the EBV viral
load as a prognostic marker and as a predictor for
EBV-association in diffuse large B-cell lymphomas.
Materials and methods
Patients
The study included a sequential unselected cohort of 218
HIV-negative patients with diffuse large B-cell lymphoma,
diagnosed and treated at the Institute of Hematology of
the Catholic University of the Sacred Heart (Rome, Italy)
between 2006 and 2013. Seventy-six patients with DLBCL
who were observed in the same time period and for
whom no data on EBV in whole blood was available were
not included into the study. This cohort had fewer
patients aged over 60 years (45%) and hepatitis C virus
(HCV)-positive patients (42%). Other patient characteris-
tics were not different from patients included in the
study. Patient characteristics are outlined in Table I.
Whole blood samples were obtained at the time of initial
diagnosis, and samples were collected using EDTA as the
anticoagulant. Survival analysis was restricted to patients
(n¼179) who received standard treatment consisting of
immunochemotherapy, combining the CD20 monoclonal
antibody Rituximab with anthracycline-based chemo-
therapy (Rituximab, Cyclophosphamide, Doxorubicin,
Vincristine, Prednisolone (R-CHOP) or R-CHOP-like).
HCV-positive patients did not receive antiviral therapy
during lymphoma treatment. Treatment consisted of R-
CHOP in 11/20 HCV-positive patients. No major liver
toxicity was observed in HCV-positive patients treated
with R-CHOP. Follow-up samples were available for 19
patients: after the end of immunochemotherapy in four
patients in complete remission (CR), and during treat-
ment after a median of three cycles (range 1–4) in 15
patients. Informed consent was obtained from patients
according to institutional guidelines, and blood sample
collection and analysis had been approved by our
institutional ethical committee.
EBV serology and EBV-DNA quantification
Serum samples were screened for the presence of IgG
antibodies to EBNA-1 and viral capsid antigen (VCA)
using commercially available enzyme immunoassays
(LiasonÕEBNA IgG and LiasonÕVCA IgG, DiaSorin
S.p.A., Salluggia, Vercelli, Italy), according to the manu-
facturer’s instructions. EBV-DNA was measured in the
whole blood of 218 patients with DLBCL at the time of
initial diagnosis (WB: n¼218; plasma: n¼47; mono-
nuclear cells: n¼44), using a commercial real-time PCR
kit, amplifying a 191 bp region of the EBNA-1 gene
(BioQuant EBV, Biodiversity, Brescia, Italy), and the ABI
PRISM 7300 Sequencer Detection System (Applied
Biosystems, CA, USA). The sensitivity for reliable EBV-
DNA detection was 200 copies/ml WB.
In situ hybridization for EBV
A group of 52 patients with DLBCL were analyzed for the
presence of EBV in the tumor cells. In-situ hybridization
(ISH) of EBV-encoded small RNAs (EBERs) on formalin-
fixed, paraffin-embedded tissue section was performed
following the manufacturer’s instructions (Dako;
Dakopatts, Golstrup, Denmark) [25].
Table I. Patient characteristics and presence of EBV in whole blood (WB).
EBV in WB Patients EBV in WB
Patient characteristics Patients Negative Positive p*(R-CHOP treated) Negative Positive p*
Number 218 164 54 179 140 39
Age Median, range (years) 66 (15–92) 64 (17–90) 70 (15–92) 0.04 63 (16–79) 62 (17–79) 66 (16–79) 0.2
Age 460 years 139 (64%) 100 (61%) 39 (72%) 0.2 106 (59%) 80 (57%) 26 (67%) 0.4
Gender Male 115 (53%) 86 (52%) 29 (54%) 1.0 102 (57%) 79(56%) 23 (59%) 0.9
Histologic subtype DLBCL, NOS 192 (88%) 144 (88%) 48 (89%) 1.0 157 (88%) 124 (89%) 33(85%) 0.4
T/HRLBCL 23 (11%) 17 (10%) 6 (11%) 19 (11%) 13 (9%) 6 (15%)
Leg-type 3 (1%) 3 (2%) 0 (0%) 3 (2%) 3 (2%) 0 (0%)
Stage Advanced (III/IV) 124 (57%) 90 (55%) 34 (63%) 0.3 101 (56%) 78 (56%) 23(59%) 0.9
B-symptoms Yes 68 (31%) 52 (32% 16 (30%) 1.0 54 (30%) 42 (30% 12 (31%) 1.0
ECOG 2–4 56 (26%) 41 (25%) 15 (28%) 0.7 51 (28%) 39(28%) 12 (31%) 0.8
Bulk 46 cm Yes 90 (41%) 68 (41%) 22 (41%) 1.0 79 (44%) 60 (43%) >19 (49%) 0.6
Extranodal 42 56 (26%) 41 (25%) 15 (28%) 0.7 46 (26%) 35 (25%) 11 (28%) 0.7
LDH Elevated 90 (41%) 63 (38%) 27 (50%) 0.2 76 (42%) 54 (39%) 22 (56%) 0.1
IPI 3–5 89 (41%) 63 (38%) 26(48%) 0.2 66 (37%) 49 (35%) 17 (44%) 0.4
HCV Positive 20 (9%) 11 (7%) 9 (17%) 0.05 11 (6%) 7 (5%) 4 (10%) 0.1
HBV HBsAg pos 3 (1%) 2 (1%) 1 (2%) 2 (1%) 1 (1%) 1 (3%)
Anti-HBcAg pos 36 (17%) 26 (16%) 10 (20%) 0.5 28 (16%) 21 (16%) 7 (19%) 0.6
Anti-HBcAg neg 177 (83%) 136 (84%) 41 (80%) 146 (82%) 117 (84%) 29 (74%)
EBER-ISH Positive 9/52(17%) 3/27(11%) 6/25(25%) 0.3 9/46(20%) 3/22(14%) 6/21(29%) 0.3
Significant p-values are in bold.
*p-value of Fisher’s exact test. For comparison of median age Wilcoxon ranked sum test was used. DLBCL, diffuse large B-cell lymphoma; NOS, not otherwise
specified; T/HRLBCL, T-cell/histiocyte-rich large B-cell lymphoma.
2M. C. TISI & S. HOHAUS
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Statistical analysis
Statistical analyses were performed using the Stata 10.0
software (Stata Corp., College Station, TX). Fisher’s exact
test was used to examine for differences in patient
characteristics according to the presence of EBV in whole
blood. Wilcoxon signed rank test was used for two-
sample comparisons of EBV-DNA levels according to
dichotomized patient characteristics. Concentrations of
viral DNA were analyzed as continuous variables follow-
ing logarithmic transformation and as dichotomic vari-
ables using the presence of EBV-DNA as a cut-off point.
Correlations between the various blood parameters
were calculated by Spearman rank correlation. The
primary survival end point was progression-free survival
(PFS), with progression during treatment, lack of com-
plete remission at the end of first-line treatment, relapse
and death from any cause were included as adverse
events. Survival curves were estimated using the Kaplan-
Meier product limit method. Log-rank tests were used to
analyze for differences in PFS. Hazard ratios and 95%
confidence intervals were adjusted for multiple prog-
nostic factors using the Cox proportional hazards model.
Results
Presence of EBV in whole blood compartments
and tumor tissue
EBV was detected in 54 of 218 (25%) WB samples at
diagnosis of DLBCL. The copy number varied between
210
2
and 4.9 10
6
copies/ml. The mononuclear cell
(MNC) fraction and plasma samples of 47 and 44 patients
were also studied. There was a significant association
between presence of EBV in WB and MNC (p¼0.04,
Supplementary Table SIA – online only), while there was
no association between EBV in WB and plasma
(Supplementary Table SIB – online only).
In 17 patients with positive EBV-DNA in WB at
diagnosis, follow-up samples were available. The samples
were collected during treatment after a median of three
cycles (range 1–4) of immunochemotherapy (13 patients:
six CR, three PR, and four with stable or progressive
disease) and after end of treatment in four patients (all in
CR). All sample results were negative for EBV-DNA.
The association between EBV in whole blood and the
tumor tissue was studied by analyzing the lymphoma
tissue of 52 patients. This cohort was selected by
balancing patients with and without EBV and preferen-
tially including histotypes suspicious for EBV-association,
as the T-cell/histiocyte-rich variant of DLBCL (T/HRLBL).
EBER-ISH was positive in the tumor cells of nine of 52
lymphoma biopsies (17%). Four of nine patients with an
EBER-positive lymphoma were younger than 50 years,
the proposed cut-off age to define EBV + DLBCL of the
elderly. A positive staining for EBER was observed in the
reactive cells of the microenvironment of three biopsies,
while no signal for EBER was observed in 40 cases.
The presence of EBER appeared to be more frequent in
the histocyte/T-cell rich subtype (5/15, 33%) compared
to DLBCL NOS (4/37, 11%, p¼0.1). This did not translate
into a higher frequency of EBV in whole blood in
T/HRLBL (6/23, 26%).
We did not find a significant association between the
presence of viral load of EBV-DNA in any blood
compartment and the presence of EBV in the lymphoma
cells [Fig. 1].
Association of EBV in whole blood and patient
characteristics
The presence of EBV-DNA in WB was associated with a
positive serology for Hepatitis C virus (HCV). A total of 20/
216 patients (9%) were HCV-positive of which 9/20
patients (45%) harbored EBV-DNA in WB, versus 44 of
196 (22%) HCV-negative patients (p¼0.05). HCV geno-
type was available for 10/20 HCV-positive patients.
Genotypes were type 2 in five patients, type 1 in four
patients and type 3 in one patient. HCV-RNA was
detected in 17/19 HCV + patients, ranging from 2 10
4
to 1.2 10
7
copies/ml. Copy numbers of EBV and HCV
were not significantly correlated (r ¼0.42, p¼0.07). There
was no association between EBV-positivity in WB and
previous exposure to hepatitis B virus (HBV) [Table I].
Immune status and EBV
Analysis of potential associations between EBV-DNA in
WB and indicators for immune response against EBV were
conducted. All 54 patients with EBV-DNA in whole blood
had a positive serology. In 48 patients the serological
study showed positive titers against both EBNA1 and
VCA, while in six patients serology was partially positive:
five patients had only titers for VCA, and one patient had
only positive EBNA titers. Patients with EBV-DNA in WB
had significantly higher levels of antibody titers against
the VCA antigen (median, 591 U/ml vs. 231 U/ml;
p¼0.007). Titers against the EBNA1 antigen were not
different between patients with or without EBV in WB
(median 265 U/ml and 222 U/ml, p¼0.7). There were no
differences in lymphocyte counts and levels of gamma
globulins, immunoglobulin IgG, IgA and IgM according to
the presence of EBV in WB (data not shown).
EBV viral load and prognosis
Analysis of the prognostic impact of EBV viral load in PB
was restricted to 179 patients. They all followed a
EBV-DNA IN DLBCL 3
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chemotherapeutic regimen consisting of the monoclo-
nal CD20 antibody Rituximab and anthracycline-contain-
ing regimen CHOP, which is considered standard
therapy for DLBCL. Median observation for progres-
sion-free survival was 36 months. Progression-free sur-
vival was significantly lower in the cohort of EBV-positive
patients (n¼39) (58% at 3 years; 95%CI, 41–72%), when
compared to EBV-negative patients (n¼140) (77% at 3-
years; 95%CI, 68–83%; p¼0.02, Fig. 2A). The presence of
EBV in WB was also a negative prognostic marker for
overall survival (p¼0.05) [Fig. 2B]. As presence of EBV
was associated with older age, we also separately
analyzed the impact of EBV according to age. In this
subgroup analysis, the age cut of 60 years did not reveal
differences for younger and older patients. Using the
median age of patients with EBV in whole blood, i.e. 70
years, we found a significant impact of EBV in the elderly
(70 years old) patients (p¼0.03), while there was no
statistically significant difference in the patients younger
than 70 years (570 years).
We analyzed the impact of EBV-DNA in the total
cohort of 218 patients including all patients irrespective
Figure 1. Viral load of EBV in WB (A), in MNC (B) and in plasma (C) according to the presence of EBV in the tumor cells detected by
EBER-ISH. Frequency and viral load in WB, MNC, and plasma were not different between EBV + and EBV- DLBCL (p¼0.3; p¼0.1;
p¼0.7).
4M. C. TISI & S. HOHAUS
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of the type of treatment. The impact on progression-free
survival and overall survival was similar to the analysis of
the R-CHOP-treated patients (p¼0.02 and p¼0.05,
respectively). However, in the multivariate Cox analysis
of progression-free survival including IPI, only the IPI
score retained its prognostic impact (p¼0.003), while
the presence of EBV-DNA was of borderline significance
(HR 1.5; 95%CI, 0.96–2.40; p¼0.07).
Discussion
EBV was detected in the whole blood of 25% of patients
with DLBCL at diagnosis. The presence of EBV was
associated with patient age and HCV infection, but not
with EBV-status of the tumor cells. Additionally, EBV-
positivity was found to be a negative prognostic factor
for survival. This raises important questions on the origin
of the virus in whole blood and the impact on the
potential mechanisms associated with poor prognosis.
Our study was not designed to definitely clarify the
association between EBV in various blood compartments
and in the tumor tissue in DLBCL. DNA from various
blood compartments and tumor tissue for EBER-ISH
were available only for a limited number of patients. This
analysis did not reveal associations between EBV in the
tumor cells and blood compartments. Our data indicate
that the presence of EBV in whole blood, in plasma or
MNC cannot be used as a bona fide test for EBV-
association. Jones et al. showed that cell-free (plasma)
but not MNC EBV-DNA in lymphoma patients can be
found as tumor-specific DNA in a broad-range of EBV-
associated lymphomas with a high-degree of sensitivity
and specificity [26]. The origin of EBV in whole blood of
patients with DLBCL whose tumors are EBV-negative is
unclear. EBV-positive bystander cells in the tumor tissue
were observed in only 3 of 43 patients (7%). All three
patients had EBV in whole blood. However, it appears
unlikely that release of EBV from EBV-positive bystander
cells within the malignant lymph node can explain the
presence of EBV in whole blood in the majority of
patients, as 16 of 19 patients with EBV in whole blood
did not show the presence of EBV in tumor or bystander
cells [Fig. 1A].
We observed a trend between EBER positivity and
morphological characteristics of T/HRLBL. Dojcinov et al.
reported that EBV + DLBCL of the elderly frequently
show features resembling T/HRLBL [27]. EBER positivity
was not restricted to elderly patients. In fact, only 6/11
EBER + patients were older than 50 years. This is in line
with a recent report from Argentina that detected similar
frequencies of EBER positivity in DLBCL in patients
younger and older than 50 years [28].
Among memory B cells in healthy individuals, there is
a small fraction of EBV + cells that accounts for approxi-
mately 1–50/10
6
circulating MNC [29]. Most probably,
this normal B-cell population is expanded in patients
with DLBCL and EBV positivity in whole blood.
The frequency of EBV-positivity that we observed is in
line with data from Marques et al. who reported a
frequency of 21% EBV positivity in MNC from whole
blood or bone marrow in a cohort of 130 patients with
aggressive lymphoma from Portugal [30]. These authors
did not test for EBV-association in the lymphoma tissue.
In their cohort, patients were either treated with CHOP
or R-CHOP. These authors observed a trend for a worse
outcome for CHOP-treated patients when EBV was
present in whole blood or bone marrow, while there
was apparently no difference for R-CHOP treated
patients, although in this subgroup survival was unex-
pectedly high. The higher number of patients included
Figure 2. Survival in 179 patients with DLBCL treated with immunochemotherapy (R-CHOP) according to the presence of EBV-DNA in
WB. Progression-free survival (A) and overall survival (B) were significantly better in patients without EBV-DNA in WB (p¼0.02, and
p¼0.05, respectively).
EBV-DNA IN DLBCL 5
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in our study, unbiased selection, a more homogenous
treatment, and geographical differences may have
contributed to the differences in the results.
EBV was more frequently present in the whole blood
of elderly patients, and in patients with HCV-infection.
Alterations of the immune system, termed immunose-
nescence, later in life and during chronic HCV-infection
could create an immunologic environment that favors
both the development of aggressive B-cell lymphoma
and expansion of EBV-positive cells in whole blood [31].
Associations between EBV-positivity and simple markers
of the immune status were explored. These included
lymphocyte counts, levels of gamma globulins and
antibody titers against EBV antigens. No associations
were found indicating a reduction of immune surveil-
lance. More in-depth research is needed. Studies of T-cell
function focusing on the cytotoxic T-cell response
directed against EBV epitopes may help to solve this
issue. Conversely, we observed increased titers against
the viral capsid antigen in DLBCL patients with EBV
in WB that may indicate increased engagement of
the immune system against EBV. Atypical EBV
serologies prior to diagnosis with increased VCA titers
has already been reported in EBV-associated Hodgkin-
lymphoma [32].
HCV infection, but not infection with HBV, was
associated with the presence of EBV in whole blood.
HCV infection was almost always associated with a high
viral load. This is in line with data from a multi-centered
study from Italy in which 91% of patients with DLBCL
and a positive HCV serology were HCV-RNA positive and
higher viral loads were associated with poorer prognosis
[33]. Conversely, serology of HBV infection in patients of
our study indicated in only three patients (1%) an active
infection, while positive anti-HBcAg titers only in 16% of
patients indicated either a resolved or occult infection.
The failure to eliminate HCV has been reported to lead
to a progressive loss of T-cell functionality [34]. This
attenuation of viral-specific T-cell responses in chronic
viral infections, known as ‘‘exhaustion’’ is characterized
by loss of proliferative capabilities, reduced cytokine
production and expression of inhibitory receptors
including programmed cell death-1 [PD-1], on CD8+
cells [34–37]. Elevated levels of soluble PD-ligand 1 have
been reported to be associated with inferior outcome in
DLBCL [38]. Further studies are needed to clarify whether
this immune dysfunction in chronic HCV infection
creates an immunological environment also favoring
expansion of EBV + memory B cells. We speculate that
EBV in WB could be an indicator for an alteration in
the immune response as a risk factor that favors B cell
proliferation of both normal and malignant B cell
subpopulations. Further studies are needed to clarify
whether this scenario could be the mechanism behind
the association of EBV in WB and inferior outcome. EBV
was cleared from whole blood after standard therapy
that included the B-cell directed monoclonal antibody
Rituximab, in line with previous observation by Kasamon
et al. [39].
In conclusion, the presence of EBV in WB is not a
surrogate marker for the EBV-status in DLBCL, but it
appears to be an indicator for an alteration in the host’s
immune control with a prognostic importance.
Potential conflict of interest: A disclosure form provided
by the author is available with the full text of this article at
www.informahealthcare.com/lal.
This work was supported by grants from AIRC (Associazione
Italiana per la Ricerca sul Cancro), the Nadia Salcini Foundation
and Fondi d’Ateneo, Linea D1, Universita
`Cattolica del Sacro
Cuore. M.G. was supported by Fondazione Roma - Progetto
Cellule Staminali.
The authors thank Ilaria Pignatelli for helpful comments on
the manuscript.
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Supplementary materials are available online
EBV-DNA IN DLBCL 7
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