DEPARTMENT OF IMMUNOLOGY & HISTOCOMPATIBILITY – UNIVERSITY OF THESSALY
TNFRSF13B/TACI and TNFRSF13C/BAFFR in B cell chronic lymphocytic leukemia
DEPARTMENT OF IMMUNOLOGY & HISTOCOMPATIBILITY – UNIVERSITY OF THESSALY
Introduction
TACI:
• Receptor of BAFF and APRIL
participates in:
• Survival and differentiation of B cells
• T cell-independent immune respon-ses
• Antibody production
• Isotype switching
• Homeostasis of B cells
BAFF-R:
• Receptor of BAFF
• Expressed on naïve B cells triggering their survival and maturation
Mackay et al, Cytokines & Growth Factor Reviews 2008
DEPARTMENT OF IMMUNOLOGY & HISTOCOMPATIBILITY – UNIVERSITY OF THESSALY
Novak et al, Blood 2002Hait et al., Immunology 2006
Jian et al., Medical Journal 2008
TACI and BAFFR might participate in the survival of B-CLL
cells, protecting them from apoptosis through NFκΒ
activation
Introduction
DEPARTMENT OF IMMUNOLOGY & HISTOCOMPATIBILITY – UNIVERSITY OF THESSALY
Anti-TACI, anti-BAFF and anti-APRIL therapeutic
approaches are now available, for the treatment of
patients with autoimmune diseases and might be useful
in patients with B-CLL • Phase I/II study of TACI-Ig to neutralize APRIL and BlyS (R) in patients with refractory or relapsed multiple myeloma or active previously-treated Waldenstrom’s macroglobuliemia.
Rossi et al, Blood 2005
• Synthetic anti-BR3 antibodies that mimic BAFF binding and target both human and murine B cells.
Lee et al, Blood 2006
• Phase I clinical study of Atacicept in patients with relapsed and refractory B-cell Non-Hodgkin’s Lymphoma. Ansell et al, Clin Cancer Res, 2008
• Ib trial of Atacicept, a recombinant protein binding BlyS and APRIL, in patients with chronic lymphocytic leukemia (B-CLL) Kofler et al, Leukemia 2011
• Development and characterization of APRIL antagonistic monoclonal antibodies for treatment of B-cell lymphomas.
Guadagnoli et al, Blood 2011
Introduction
DEPARTMENT OF IMMUNOLOGY & HISTOCOMPATIBILITY – UNIVERSITY OF THESSALY
The determination of TNFRSF13B/ TACI and
TNFRSF13C/ BAFFR expression on B-CLL cells and
their contribution to the phenotype and the
prognosis of the disease
Aim of the study
DEPARTMENT OF IMMUNOLOGY & HISTOCOMPATIBILITY – UNIVERSITY OF THESSALY
Bone marrow
104 patients with B-CLL (M/F: 59/45, mean age: 68.5 y)
30 patients with low-grade NHL
(M/F: 18/12, mean age: 77.3 y) &
145 healthy donors (M/F: 84/61, mean age: 67.8 y)
Peripheral blood
Materials & Methods
DEPARTMENT OF IMMUNOLOGY & HISTOCOMPATIBILITY – UNIVERSITY OF THESSALY
1. Hematologic, serologic and flow cytometric
analyses (for diagnostic purposes), including in
B-CLL a further analysis of the most powerful
prognostic factors (ZAP-70, CD38 and IgH
mutational status).
1. TACI and BAFFR protein expression was detected
by flow cytometry using monoclonal
antibodies (TACI clone 1a1, Abcam and BAFFR
clone 11C1 Biolegend, respectively) in all
samples.
2. TNFRSF13B and TNFRSF13C mRNA expression were
estimated by qRT-PCR in 29 samples (19 with B-
CLL, 8 with NHL and isolated B cells from 2
healthy donors).
Materials & Methods
DEPARTMENT OF IMMUNOLOGY & HISTOCOMPATIBILITY – UNIVERSITY OF THESSALY
Materials & Methods
4. B-CLL cells exhibiting high or absent TACI, were
stimulated by PKW (pokweed) 2.5 ng/mL, BAFF 1
μg/mL and APRIL 200 ng/mL and the apoptosis
status was estimated by flow cytometry using an
Annexin V-FITC/ 7-AAD (7-AAD) kit.
5. BAFF and APRIL serum levels were estimated by
ELISA in 37 samples (16 with B-CLL, 11 with NHL
and 10 healthy donors).
6. The statistical analysis was performed using the
SPSS software ver.16.0.
DEPARTMENT OF IMMUNOLOGY & HISTOCOMPATIBILITY – UNIVERSITY OF THESSALY
Results - 1
TACI=2.1% TACI=55% TACI=90.1%
Variable TACI mRNA and protein expression on B-CLL cells
BAFFR protein expression
B-CLL cells normal B cells
MFI=6 MFI=11
DEPARTMENT OF IMMUNOLOGY & HISTOCOMPATIBILITY – UNIVERSITY OF THESSALY
Results - 2
TACI=2.1% TACI=55% TACI=90.1%
Variable TACI mRNA and protein expression on B-CLL cells
TACI mean expression ± STDEV
B-CLL patients
13.4 ± 22% p < 0.05
NHL patients 36.8 ± 29% p = 0.340
Healthy donors
27.6 ± 14.2%B-CLL NHL Healthy
TACI
expression (%)
DEPARTMENT OF IMMUNOLOGY & HISTOCOMPATIBILITY – UNIVERSITY OF THESSALY
TACI expression was not
asso-ciated with
(p>0.05) :
•the presence of
autoimmunity,
•hypogammaglobulinemia,
•monoclonal gammapathy,
•the infection risk and
•the known prognostic
factors of B-CLL
Results - 3
Pts Disease TACI (%)
Hypermutation
status(%)
CD38 (%)
1. B-CLL 0.1 99.6 5.4
2. B-NHL 1.1 98.2 3.6
3. B-CLL 2.2 99.6 73
4. B-CLL 2.2 90.1 1.4
5. B-CLL 2.5 98.8 23.2
6. B-CLL 16.9 95 1
7. B-CLL 21.9 96 18.5
8. B-CLL 29.8 99.6 63.8
9. B-CLL 43.5 94.8 1.9
10. B-CLL 54.3 96.2 0.3
11. B-CLL 78.5 99.6 24.1
12. B-NHL 84 88 6
Low TACI
Medium
TACI
High TACI
hypermutation status ≤ 98% good prognosis CD38 > 20%
bad prognosis
DEPARTMENT OF IMMUNOLOGY & HISTOCOMPATIBILITY – UNIVERSITY OF THESSALY
CONTROL BAFF APRIL
APRILCONTROL BAFF
TACI protects B-CLL cells from apoptosis
TACI=2.1%
TACI=50.1%
alive Early apoptotic
Late apoptotic&dead
alive Early apoptotic
Late apoptotic&dead
alive Early apoptotic
Late apoptotic&dead
alive Early apoptotic
Late apoptotic&dead
alive Early apoptotic
Late apoptotic&dead
alive Early apoptotic
Late apoptotic&dead
Results - 4
DEPARTMENT OF IMMUNOLOGY & HISTOCOMPATIBILITY – UNIVERSITY OF THESSALY
B-CLL NHL Healthy
BAFFR_MFI
BAFFR_MFI Mean ± STDEV
B-CLL patients
6.71 ± 2.9% p < 0.05
NHL patients 6.7 ± 3.7% p = 0.05
Healthy donors
10.7 ± 5.9%
BCLL cells
BCLL cells
Normal B cells
At the relapse of the disease
Results - 5
DEPARTMENT OF IMMUNOLOGY & HISTOCOMPATIBILITY – UNIVERSITY OF THESSALY
Serum BAFF was low and APRIL high in the majority of B-CLL
patients compared to NHL patients and healthy donors
BAFF pg/ml Median (Range)
APRIL pg/ml Median (Range)
B-CLL patients
0 (0 – 55.0) 5.11 (0.80 - 64.06)
NHL patients 9.0 (0 – 152.0) 2.21 (0 - 5.11)
Healthy donors
49.5 (21.8 – 92.0) 4.78 (2.47 - 18.55)
Soluble BAFF levels was low to undetectable in B-CLL patients. Kreuzaler et al, J Immunol
2012
APRIL serum levels predict time to first treatment in patients affected by B- cell chronic lymphocytic leukemia. Tecchio et al, Eur. J. Haematol 2011
Results - 6
DEPARTMENT OF IMMUNOLOGY & HISTOCOMPATIBILITY – UNIVERSITY OF THESSALY
Conclusions
TACI expression is:
• low in the majority of B-CLL patients
• not associated with disease prognosis
• accompanied by very low or absent serum BAFF and high serum APRIL levels
BAFFR MFI expression is:
• low in the B-CLL and NHL patients
These findings should be taken into account in the
case of anti-TACI and anti-BAFFR therapeutic
approaches.
DEPARTMENT OF IMMUNOLOGY & HISTOCOMPATIBILITY – UNIVERSITY OF THESSALY
THANK YOU
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