Download Flow Cytometric Detection of Immunophenotypically Normal Plasma

MOKSLO DARBAI • SCIENTIFIC PAPERS
Laboratorinë medicina.
2011, t. 13, Nr. 2(50), p. 59–64.
Flow Cytometric Detection of
Immunophenotypically Normal
Plasma Cells in Multiple
Myeloma Patients Provides
Clinically Important Information
1, 2
Valdas Peèeliûnas
3
Auðra Janiulionienë
3, 4
Rëda Matuzevièienë
1, 2
Laimonas Griðkevièius
1
Hematology, Oncology and
Transfusion Medicine Center,
Vilnius University Hospital
Santariðkiø Clinics, Santariðkiø 2,
08661 Vilnius, Lithuania
E-mail: [email protected]
2
Clinics of Internal, Family Medicine
and Oncology, Faculty of Medicine,
Vilnius University,
M. K. Èiurlionio 21, 03101 Vilnius,
Lithuania
Summary
Background. With the advent of multiparameter flow cytometry (FC)
it became possible simultaneously analyze normal and malignant
plasma cells (PC) in multiple myeloma (MM) patients. There is some
data that residual normal bone marrow plasma cells (nBMPC) is of prognostic significance in plasma cells dyscrasias. There is no data about
clinical value of immunophenotypically normal peripheral blood circulating plasma cells (nCPC).
Material and methods. In this study we evaluated clinical significance of normal plasma cells detection by FC in peripheral blood (PB) and
bone marrow (BM) compartments of refractory or relapsed (RR) multiple
myeloma (MM) patients. For sample analysis we used 2 tubes stained
with antibody combinations CD56/CD138/CD45/CD19/CD38/CD20 and
cLambda/cKappa/CD138/CD19/CD38/CD56.
Results. PB and BM samples taken at the same time from 40 relapsed/refractory MM patients and 11 healthy controls (HC) were analyzed. We were able to detect nCPC and nBMPC subpopulation in 57.5%
and 90% of MM patients, respectively. We did not find any clinical significance of nCPC detection at baseline, however increase of nCPC in response to treatment was associated with prolonged time to progression (TTP). We also showed that proportion of residual nBMPC to all
BMPC is of prognostic significance in advanced MM patients.
Conclusions. Our finding supports clinical value of nPC identification for advanced MM patients, and use of FC rather than light microscopy for BMPC compartment analysis before treatment.
Keywords: myeloma, immunophenotyping, prognosis.
INTRODUCTION
3
Laboratory Diagnostics Centre,
Vilnius University Hospital
Santariðkiø Clinics, Santariðkiø 2,
08661 Vilnius, Lithuania
4
Physiology, Biochemistry and
Laboratory Medicine Department,
Faculty of Medicine, Vilnius University
M. K. Èiurlionio 21, 03101 Vilnius,
Lithuania
In the last few years flow cytometry
(FC) became an important tool for
evaluation of multiple myeloma in especially in diagnosis, prognosis and response to treatment [1]. This method
was shown to be valuable in analysis
of minimal residual disease (MRD) after treatment [2–4]. Growing interest
to flow cytometry is determined by re-
LABORATORINË MEDICINA Ÿ 2011, t. 13, Nr. 2(50)
cent re mark able im prove ments in
myeloma treatment efficacy. There is
an increasing number of high quality
remissions beyond sensitivity level of
con ven tional methods used for response definition. Immunophenotypical characteristic of malignant plasma
cells (PC) also was shown to be of prognostic significance [5, 6]. With the improvement of FC technology became
possible to simultaneously identify
59
Valdas Peèeliûnas, Auðra Janiulionienë, Rëda Matuzevièienë, Laimonas Griðkevièius
normal and malignant PC in the same
sample [7–9]. This was shown to have
prognostic value in such plasma cells
disorders as MGUS, asymptomatic
and symptomatic myeloma [10, 11].
Until now BM compartment of PC was
the main object of investigations, PB
however was uncommon target for investigations mainly due to very low
counts of cir cu lating plasma cells.
Nevertheless several studies were performed and results showed that presence and increased amount of circulating in peripheral blood plasma cells
(CPC) are associated with more aggressive disease course and worse
prognosis [12–15]. In clinical studies
evaluating PB compartment of PC
mostly 3 to 4 colors FC was used and
con cen trated on the anal y sis of
immunophenotypically aberrant CPC
(aCPC) only. There is little known
about normal PC population in PB of
myeloma patients, also only few
clinical studies analyzed normal bone
marrow plasma cells (nBMPC).
Main purpose of this study was to
assess normal PC subpopulation of
MM patients in PB and BM compartments and evaluate its clinical significance.
Number of patients included
40
Age years median (range)
57 (39–74)
Males/females n
26/14
IgG n (%)
IgA n (%)
IgD n (%)
LCH only n (%)
Non/oligosecreating n (%)
25 (62.5)
6 (15)
2 (5)
3 (7.5)
4 (10)
Time from diagnosis median months (range)
24.5 (3–172)
Treatment lines median (range)
2 (1–5)
Previous HDT n (%)
15 (37.5)
b2mg mg/L median (range)
4.1 mg/L (2.0–45.4)
Albumin g/L median (range)
43.5 (24.4–53.0)
Hemoglobin g/L median (range)
114 (72.1–152.8)
Extramedullary relapse n (%)
5 (12.5)
LCH – light chains, HDT – high dose chemotherapy, Ig – immunoglobulin, b2mg –
beta 2 microglobulin
Patients were prospectively included
if they met the following inclusion criteria: were 18 years of age or older,
had relapsed or refractory multiple
myeloma according to EBMT criteria
[16] after at least one prior line of therapy and were scheduled to receive either a Bortezomib containing regimen
or VAD (vincristin, doxorubicin and
dexamethasone). Clinical and laboratory data including age, sex, type of
paraprotein, b2 microglobulin levels,
time from diagnosis, the number of
lines of therapy, previous high-dose
therapy were recorded. All patients
signed informed consent according to
Declaration of Helsinki. The study
was ap proved by the Lith u a nian
Bioethics Committee.
APC/PE-Cy7/APC-Cy7). All samples
were prepared by erythrocyte lysed
whole blood/bone marrow technique.
For intracytoplasmic immunoglobulin
l i g h t c h a i n de t e c t i o n we u s e d
Fix/Perm (BD, San Jose) cell permeabilization solution. Choice of markers
was in agreement with most recent
European Myeloma Network (EMN)
and In ter na tional Myeloma Workgroup (IMWG) consensus recommendations and our own experience [1].
Data was ac quired on 6 col our
multiparameter FACSCanto (BD, San
Jose, CA) flow cytometer equiped with
FACSDiVa data acquisition and analysis software.
In order to achieve better specificity (avoiding false positive results)
minimal threshold for positive PC population in PB was defined as more
than 20 events. PC population less
than minimal threshold was defined
as “0” [1, 8].
“Normal” PC population was defined as CD138+, CD38+, CD19+,
CD56-, normal kappa/lambda ratio,
CD45 variable. “Aberrant” immunophenotype was defined as CD138+,
CD38+, CD19-, CD56+/-, abnor mal
kappa/lambda ratio, CD45 variable
[1].
Data is presented as percentage of
total living cells (TLC) in the sample
analyzed.
Flow cytometry
Statistical analysis
MM patients BMPC
subpopulations
6 colour FC used in this study was previously described in detail elsewhere
[17]. In short: BM and PB samples
were stained in two tubes with
following an ti body com bi na tions:
CD56/CD138/CD45/CD19/CD38/CD20
and cLambda/cKappa/CD138/CD19/
CD38/CD45 (FITC/PE/PerCP-Cy-5.5/
OS was defined as the duration between the date of entry into the study
and death, with those alive censored
at the last follow-up date. TTP was defined as the interval from entry into
the study to disease progression. Survival was analyzed by Kaplan and
Meier method [18]. Differences be-
We were able to confirm the presence
of plasma cells in bone marrow of patients by morphology under the light
microscope (LM) in all but one patient
with predominant extramedulary disease. The proportion of plasma cells in
BM by LM was 12.5% (median, range
0–89%).
MATERIAL AND METHODS
Patient selection and
characteristics
60
Table 1. Characteristics of myeloma patients included in study
tween survival curves were evaluated
using the log-rank test. For multivariate analysis Cox regression proportional hazard model (stepwise regres sion) was used. Com par i sons
among subgroups were performed using Mann-Whitney Test. A two-tailed
P value less than 0.05 was considered
as significant.
RESULTS
Patients
Blood and bone marrow samples of
40 adult relapsed/refracory (RR) multiple mieloma (MM) patients admitted
to sin gle cen ter and fulfiling the
Blade (C) criteria for relapse/refractoriness were analyzed. Samples of peripheral blood and bone marrow were
taken simultaneously before starting
treatment of relapse. PB samples were
also analyzed before second treatment
course to evaluate kinetics of normal
circulating plasma cells (nCPC) in response to treatment. Patients characteristics are presented in table 1.
We also included 11 healthy bone
marrow donors as a controls. Median
age of healthy con trols (HC) was
28 years (19–74), 6 males and 5 females.
LABORATORINË MEDICINA Ÿ 2011, t. 13, Nr. 2(50)
Flow Cytometric Detection of Immunophenotypically Normal Plasma Cells in Multiple Myeloma Patients Provides Clinically Important Information
Table 2. Size of immunophenotypically normal plasma cell subpopulation in peripheral blood and bone marrow of MM (multiple myeloma) patients and healthy
controls. Shown as a proportion of TLC (total living cells) in a sample
MM patients
N40
nCPC
CD19+CD56-
nBMPC
CD19+CD56-
Mean
0.015%
0.11%
Median
0.006%
0.04%
p=0.48
p=0.001
Mean
0.01%
0.21%
Median
0.003%
0.20%
Mann-Whitney Test
Healthy
N11
nCPC – immunophenotypically normal plasma cells
nBMPC – immunophenotypically normal bone marrow plasma cells
Flow cytometry revealed plasma
cells in all 40 patients’ bone marrow
samples. Majority of these cells had
aberrant phenotype and proportion of
such cells in the sample was 8.0% (median, range 0.05–87.9%).
32 MM patients (80%) had detectable nBMPC subpopulation. Median
proportion of detected nBMPC subpopulation was 0.04% (0–1.51) of TLC
in the sample. nBMPC subpopulations
were de tect able by FC in all HC.
nBMPC proportion in MM patients was
decreased as compared to HC (Table 2).
proximately 1 log smaller proportions,
than aberrant phenotype aCPC: mean
0.025±0.034% and mean of 0.72±1.2%.
nCPC subpopulation were identifiable
in similar proportion of HC as in MM
patients – 7 of 11 cases (63.6%). Size of
detected nCPC subpopulations in MM
patients and HC were also similar (Table 2).
Peripheral blood PC in
MM patients
nCPC
Peripheral blood circulating plasma
cells were detected in 33 cases out of
40 (82.5%). In 23 (57.5%) cases we
were able to identify immunophenotypically nCPC. Besides nCPC in some
MM patients we were also able to detect immunophenotypically aberrant
circulating PC (aCPC). Proportion of
both CPC subpopulations was very
low, however, in patients with detectable CPC, nCPC were observed in ap-
Prognostic value of
immunophenotypically normal
PC detection in RR MM patients
nCPC population, whether detected or
not at the begining of treatment had
no significant associations neither
with disease burden (defined as bone
marrow plasma cells by microscopy or
FC) nor biological properties (reflected
by b2mg, albumin, hemoglobin levels
and renal function). Detection of baseline nCPC was also not prognostic for
TTP and OS. We were able to identify
only weak inverse correlation of nCPC
pro por tion with MM pa tients age
(r=-0.34, p=0.03).
We also looked at kinetics of MM
patients’ nCPC in response to one
treatment course. We found proportion of nCPC significantly lower at
baseline than after one treatment
course – median 0.006% vs 0.013% respectively (p=0.014). Patients with increased nCPC had prolonged TTP in
comparison to patients with decreased
or absent nCPC: median of 339 vs
105 days (p=0.038) respectively. OS
was not different in both groups.
nBMPC
We evaluated prognostic impact of residual bone marrow nPC in refractory
or relapsed MM patients. Five patients with pre dom i nantly extramedullar relapse and low BM involvement were excluded from this analysis, as BM analysis in these cases was
considered to be not informative in respect to disease activity. To test prognostic value of aBMPC/nBMPC ratio
in univariate fashion we dichotomised
patients in two groups according to the
median value of aBMPC/nBMPC ratio, which was found to be 225. TTP
and OS both were significantly longer
in patients with higher nBMPC propor tion me dian 414 vs 230 days
(p=0.019) and median not reached vs
533 days (p=0.038), re spec tively
(Fig. 2d). Markers related to disease
activity and prognosis have more favorable profile in patients with higher
nBMPC pro por tion: lower b2mg
(p=0.006), BMPC by LM (p=0.019) and
calcium (p=0.045), higher hemoglobin
(p=0.03) and plate lets (0.002). We
built Cox re gres sion model us ing
aBMPC/nBMPC ratio as continuous
variable for multivariate analy sis.
aBMPC/nBMPC ratio was found to be
only significant independant predictor
for TTP (p=0.014) and OS (p=0.003).
DISCUSSION
Fig. 1. Kinetics of immunophenotypically normal circulating plasma cells
(nCPC) in response to one chemotherapy course
Error bars represents 95% confidence interval for mean.
LABORATORINË MEDICINA Ÿ 2011, t. 13, Nr. 2(50)
Very low numbers of polyclonal circulating plasma cells (CD138+/CD19+/
CD56-) are detectable in the peripheral blood of healthy people. They
have been shown to be early, transitional stage plasma cells (pre-plasma
cells) differentiating from B lymphocytes that migrate from peripheral
lymphoid organs to the bone marrow
[7, 19] or reactive short living immunoglobulin secreting plasmacytes [7,
20]. The numbers of immunophenotypically normal CPCs tend to increase in reactive conditions (e.g. infection, vaccination). Reactive plasmacytosis has also been reported during
hematopoietic recovery in MM patients post chemotherapy [19, 21]. The
lifespan of normal CPCs in peripheral
61
Valdas Peèeliûnas, Auðra Janiulionienë, Rëda Matuzevièienë, Laimonas Griðkevièius
Fig. 2. Prognostic impact of bone marrow plasma cell content analysis by different methods
Bone marrow samples from myeloma patients were analyzed for plasma cells content by light microscopy (LM) or flow cytometry
(3 parameter were measured: aBMPC – immunophenotypically aberrant bone marrow plasma cells, nBMPC – immunophenotypically
normal bone marrow plasma cells, aBMPC/nBMPC – ratio of detected aBMPC and nBMPC calculated for every patient). Patients were
assigned to one of two groups by the established value of evaluated parameter – above or below median value, represented by doted line or
solid line respectively. Univariate analysis was done for time to progression (TTP) and overall survival (OS) in each pair by ploting
Kaplan Meyer curves. Log-rank test was used for calculation of p value.
blood is usually short as they undergo
apoptosis or migrate into the bone
marrow [7, 20, 22]. To the best of our
knowledge, this is the first report of
the prognostic impact of nCPC population in MM patients.
In PB of healthy people we were
able to detect only immuno phenotypically normal, polyclonal PC populations with spo radic CD19- events,
without formation of any population.
Proportions of detected nCPC were
identical in MM patients and HC.
Moreover there were no significant associations of nCPC with any of disease
burden or activity parameter. nCPC
proportion had only weak inverse correlation with patients’ age. This finding is inline with recent publication of
Caraux et al. that amount of circulating PC have a tendency to decrease
during ageing in otherwise helthy
people [23].
The detection of nCPC population
before treatment did not demonstrate
prognostic significance. However, patients with increasing nCPCs in response to treatment had longer TTP
compared to those with decreasing or
undetectable nCPC. Increasing nCPC
after treatment could be related to
haematopoietic reconstitution after
chemotherapy [21] and may be a feature of better residual immune reactivity. This observation is compatible
with data from other studies indicating that higher normal plasma cell
proportion in BM after treatment is
associated with prolonged progression
free survival as well as better immune
62
reconstitution [2, 10]. Unique biological role of nCPC subpopulation could
be emphasized by our findings that PB
PC of MM patients carrying aberrant
immunophenotype (aCPC) and attributable to malignant clone had inverse
clinical significance to nCPC: increase
of aCPC after chemotherapy was
demonstrated to be an extremely poor
prognosis factor [24].
It is difficult to draw unequivocal
conclusions from this small study, but
we could speculate that peripheral
normal PC production is not impaired
in MM patients and immunoparesis is
a consequence of depressed nPC population in BM of MM patients.
Recently B. Paiva et al. presented
re sults of large study in volv ing
594 newly diagnosed MM patients.
They demonstrated that more than 5%
residual nBMPC of all BMPC were associated with better prognosis, and
more favorable disease characteristics
[25]. How ever, as pro por tion of
nBMPC de pends equally on both
nBMPC and malignant BMPC, authors did not clearly indicate whether
prognostic impact of detected nBMPC
proportion was independent from malignant BMPC. We analyzed advanced
MM pa tients and pro por tion of
nBMPC found in our cohort was much
l o we r. T hus we c on s i d er ed t o
dichotomize patients’ cohort into two
groups using median level of nBMPC
detected, which was approximately at
one log lower than the 5% level discussed in paper of B. Paiva et al. We
found out that higher proportion of re-
sidual nBMPC was predictive for prolonged TTP and OS (Fig. 2d). Thereafter using the same dichotomisation
method we grouped patients according
to the me dian val ues of nBMPC,
aBMPC, and BMPC detected by LM
then checked newly formed patient
groups for differences in TTP and OS
(Fig. 2b, 2c). There were no significant
differences in these groups. aBMPC/
nBMPC seems to present summarized
prognostic impact factor of both involved parameters. aBMPC/nBMPC
ratio was shown to be of higher prognostic value than BMPC detected by
LM (Fig. 2a). Multivariate analysis
con firmed in de pend ent prog nos tic
value of aBMPC/nBMPC ratio for TTP
and OS.
In sum mary, we dem on strated
that peripheral blood nCPC (immunophenotypically normal plasma cells)
remains rather intact and independent of disease burden in advanced
myeloma patients. Moreover, better
immune reactivity as reflected by increase of nCPC in response to chemotherapy is associated with prolonged
time to pro gres sion. Pro por tion of
nBMPC of total BMPC is of prognostic
significance in RR MM patients. Our
findings together with data from study
of B. Paiva et al indicate that baseline
anal y sis of aBMPC and nBMPC
subpopulations using FC should be
prefered over LM in both newly diagnosed and RR MM patients. u
Gauta:
Priimta spaudai:
2011 04 14
2011 06 27
LABORATORINË MEDICINA Ÿ 2011, t. 13, Nr. 2(50)
Flow Cytometric Detection of Immunophenotypically Normal Plasma Cells in Multiple Myeloma Patients Provides Clinically Important Information
Santrauka
NORMALIØ PLAZMINIØ LÀSTELIØ NUSTATYMAS MIELOMINE LIGA
SERGANTIEMS PACIENTAMS TAIKANT TËKMËS CITOMETRIJÀ SUTEIKIA
KLINIÐKAI REIKÐMINGOS INFORMACIJOS
Valdas Peèeliûnas, Auðra Janiulionienë, Rëda Matuzevièienë,
Laimonas Griðkevièius
Ávadas. Tobulëjanti tëkmës citometrijos
metodologija ágalino viename mëginyje
atskirai analizuoti normalias ir piktybines plazmines làsteles. Yra duomenø,
kad nustatytas didesnis normaliø plazminiø làsteliø (nPL) kiekis kaulø èiulpuose yra prognostiðkai palankus veiksnys pacientams, sergantiems plazminiø
làs te liø ne op la zi jo mis. Iki ðiol në ra
skelbta duomenø apie mielomine liga
(ML) serganèiø pacientø periferiniame
kraujyje cirkuliuojanèiø nPL klinikinæ
reikðmæ.
Tiriamieji ir metodai. Taikydami
6 spalvø tëkmës citometrijà tyrëme nPL
kieká recidyvavusia ar atsparia gydymui
ML serganèiø pacientø periferiniame kraujyje ir kaulø èiulpuose, analizavome jø klinikinæ reikðmæ. Mëginiai buvo nudaþyti
2 antikûnø kombinacijomis CD56/CD138/
CD45/CD19/CD38/CD20 ir cLamb da/
cKappa/CD138/CD19/CD38/CD45.
Rezultatai. Periferinio kraujo ir kaulø
èiulpø mëginiai buvo paimti ið 40 ML serganèiø pacientø ir 11 sveikø donorø. Mes
nustatëme nPL populiacijas 57,5 % ir 90 %
ML pa cien tø ati tin ka mai pe ri fe ri nio
kraujo ir kaulø èiulpø mëginiø prieð gydymà. nPL radimas periferiniame kraujuje neturëjo prognostinës reikðmës, taèiau nPL kiekio padidëjimas periferiniame kraujuje po pirmojo gydymo kurso buvo susijæs su prailgëjusiu laiku iki ligos
progresijos. Mes taip pat nustatëme, kad
nPL santykis su visomis kaulø èiulpuose
nustatytomis plazminë mis làstelë mis
yra reikðmingas ir nepriklausomas paþengusia ML serganèiø pacientø prognostinis þymuo.
Iðvados. Mûsø tyrimas parodë, kad
nPL nustatymas recidyvavusia ar atsparia gydymui mielomine liga sergantiems
pacientams teikia kliniðkai reikðmingos
informacijos. ML pacientø kaulø èiulpø
mëginio tyrimas prieð gydymà taikant
TC teikia prognostiðkai svarbesnës informacijos nei mikroskopavimas.
Reikðminiai þodþiai: mielominë liga,
imunotipavimas, prognozë.
REFERENCES
1. Rawstron AC, Orfao A, Beksac M,
Bezdickova L, Broo imans RA,
Bumbea H, et al. Report of the European Myeloma Network on multiparametric flow cytometry in mul ti ple
m y e lo m a a nd r e l a t e d d i s o r d e r s .
Haematologica 2008; 93(3): 431–8.
2. Rawstron AC, Davies FE, DasGupta R,
Ashcroft AJ, Patmore R, Drayson MT,
et al. Flow cytometric disease monitoring in multiple myeloma: the relationship between normal and neoplastic
plasma cells predicts outcome after
transplantation. Blood 2002;
100(9): 3095–100.
3. Sarasquete ME, Garcia-Sanz R, Gonzalez D, Martinez J, Mateo G, Martinez P,
et al. Minimal residual disease monitoring in multiple myeloma: a comparison between allelic-specific oligonucleotide real-time quantitative polymerase chain reaction and flow cytometry.
Haematologica 2005; 90(10): 1365–72.
4. Paiva B, Vidriales MB, Cervero J,
Mateo G, Perez JJ, Montalban MA, et
al. Multiparameter flow cytometric remission is the most relevant prognostic
factor for multiple myeloma patients
who un dergo autologous stem cell
transplantation. Blood 2008;
112(10): 4017–23.
5. M a t e o
G,
M o n t a l ba n
MA,
Vidriales MB, Lahuerta JJ,
Mateos MV, Gutierrez N, et al. Prognostic value of immunophenotyping in
mul ti ple myeloma: a study by the
PETHEMA/GEM co op er a tive study
groups on patients uniformly treated
with high-dose therapy. J Clin Oncol
2008; 26(16): 2737–44.
6. M o r e a u P , R o b i l l a r d N , A v e t Loiseau H, Pineau D, Morineau N,
Milpied N, et al. Patients with CD45
negative multiple myeloma receiving
high-dose therapy have a shorter survival than those with CD45 positive
mul ti ple myeloma. Haematologica
2004; 89(5): 547–51.
7. Pellat-Deceunynck C, Bataille R. Normal and ma lig nant hu man plasma
cells: proliferation, differentiation, and
expansions in relation to CD45 expression. Blood Cells Mo l Dis 2004;
32(2): 293–301.
8. Rawstron AC. Immunophenotyping of
plasma cells. Curr Protoc Cytom 2006;
Chapter 6: Unit 6 23.
9. Rawstron A, Fenton JA, Gonzalez D,
Davies FE, O’Connor SJ, Richards S, et
al. Accurate prediction of outcome for
patients with IgG and IgA monoclonal
gammopathy of unknown significance
using flow cytometry. Haematologica
2005; 90(Suppl No. 1): 196.
10. San Miguel JF, Almeida J, Mateo G,
Blade J, Lopez-Berges C, Caballero D,
et al. Immunophenotypic evaluation of
the plasma cell compartment in multiple myeloma: a tool for comparing the
efficacy of different treatment strategies and predicting outcome. Blood
2002; 99(5): 1853–6.
11. Perez-Per sona E, Vidriales MB,
Mateo G, Garcia-Sanz R, Mateos MV,
de Coca AG, et al. New criteria to identify risk of progression in monoclonal
gammopathy of uncertain significance
and smol der ing mul ti ple myeloma
base d on mult iparame ter fl ow
cytometry analysis of bone marrow
plasma
cells.
Blood
2007;
110(7): 2586–92.
12. Rawstron AC, Owen RG, Davies FE,
Johnson RJ, Jones RA, Richards SJ, et
al. Circulating plasma cells in multiple
myeloma: characterization and correlation with disease stage. Br J Haematol
1997; 97(1): 46–55.
13. Nowakowski GS, Witzig TE, Dingli D,
Tracz MJ, Gertz MA, Lacy MQ, et al.
Circulating plasma cells detected by
flow cytometry as a predictor of survival in 302 patients with newly diag-
LABORATORINË MEDICINA Ÿ 2011, t. 13, Nr. 2(50)
nosed multiple myeloma. Blood 2005;
106(7): 2276–9.
14. Mateo G, San Miguel JF, Orfao A.
Immunophenotyping of plasma cells in
multiple myeloma. Methods Mol Med
2005; 113: 5–24.
15. Kumar S, Rajkumar SV, Kyle RA,
Lacy MQ, Dispenzieri A, Fonseca R, et
al. Prog nos tic value of cir cu lat ing
plasma cells in monoclonal gammopathy of undetermined significance.
J Clin Oncol 2005; 23(24): 5668–74.
16. B l a d e J , S a m s o n D , R e e c e D ,
Apperley J, Bjorkstrand B, Gahrton G,
et al. Criteria for evaluating disease response and progression in patients
with multi ple myeloma treated by
high-dose therapy and haemopoietic
stem cell transplantation. Myeloma
Subcommittee of the EBMT. European
Group for Blood and Marrow Transplant. Br J Haematol 1998,
102(5): 1115–23.
17. Peceliunas V, Janiulioniene A, Matuzeviciene R, Griskevicius L. Six color flow
cytometry detects plasma cells expressing ab er rant immunophenotype in
bone marrow of healthy donors. Clinical Cytometry; Part B 2011, In press.
18. Kaplan EL, Meier P. Nonparametric
estimation from incomplete observations. Journal of the American Statistical Association 1958; 53: 457–81.
19. Medina F, Segundo C, Campos-Caro A,
Gonzalez-Garcia I, Brieva JA. The heterogeneity shown by human plasma
cells from tonsil, blood, and bone marrow reveals graded stages of increasing
maturity, but local profiles of adhesion
mol e cule ex pres sion. Blood 2002;
99(6): 2154–61.
20. Bataille R, Jego G, Robillard N,
Barille- Nion S, Harousseau JL,
Moreau P, et al. The phenotype of normal, reactive and malignant plasma
cells. Identification of “many and multiple myelomas” and of new targets for
63
Valdas Peèeliûnas, Auðra Janiulionienë, Rëda Matuzevièienë, Laimonas Griðkevièius
myeloma ther apy. Haematologica
2006; 91(9): 1234–40.
21. Jego G, Avet-Loiseau H, Robillard N,
Moreau P, Amiot M, Harousseau J, et
al. Reactive plasmacytoses in multiple
myeloma during hematopoietic recovery with G- or GM-CSF. Leuk Res 2000;
24(7): 627–30.
22. Jego G, Robillard N, Puthier D,
Amiot M, Accard F, Pineau D, et al. Reactive plasmacytoses are expansions of
plasmablasts retaining the capacity to
64
differentiate into plasma cells. Blood
1999; 94(2): 701–12.
23. Caraux A, Klein B, Paiva B, Bret C,
Schmitz A, Fuhle r GM, e t al.
Circulating human B and plasma cells.
Age-associated changes in counts and
detailed characterization of circulating
normal CD138- and CD138+ plasma
cells.
Haematologica
2010;
95(6): 1016–20.
24. Peceliunas V, Janiulioniene A, Matuzeviciene R, Griskevicius L. Circulating
plasma cells (CPCs) predict the outcome of relapsed or refractory multiple
myeloma (RR MM). Haematologica
2010; 95(Suppl. 2): 1.
25. Paiva B, Vidriales MB, Mateo G,
Perez JJ, Montalban MA, Sureda A, et
al. The persistence of immunophenotypically normal residual bone marrow
plasma cells at diagnosis identifies a
good prognostic subgroup of symptomatic multiple myeloma patients. Blood
2009; 114(20): 4369–72.
LABORATORINË MEDICINA Ÿ 2011, t. 13, Nr. 2(50)