Transcript Слайд 1 - Univerzita Karlova v Praze

Belarusian Center for
Primary
Immunodeficiencies
Belarusian Research
Center for Pediatric
Oncology and Hematology
Severe reduced level of memory B cells
– is it diagnostic tool for CVID?
ESID – Prague
May 2009
Svetlana Sharapova
Sasha Migas
Galina Kachan
Mihael Belevtsev
Immunology Department
Molecular Biology Department
Clinical Department
Immunology Department
Common Variable Immunodeficiency
Common variable immunodeficiency (CVID) is a heterogeneous disorder that is associated with low
serum immunoglobulin concentrations, defective specific-antibody production and an increased
susceptibility to bacterial infections of the respiratory and gastrointestinal tracts.
(Ochs H. et alPrimary Immunodeficincy Disease: Oxford University Press 2007)
CVID has two picks of manifestation: from 6 to 10 and from 26 to 40 years.
Gastrointestinal complications are fairly common in CVID —up to 50% of patients with CVID have
chronic diarrhoea with malabsorption. Other gastrointestinal diagnoses in patients with CVID include
Crohn’s disease, intestinal granulomatous disease, intestinal parasitic bacterial or viral infections,
coeliac sprue, and intestinal lymphangiectasia.
Autoimmune thrombocytopenic purpura and autoimmune haemolytic anaemia are the most common
autoimmune consequences, occurring in 5–8% of all patients with CVID.
(Lancet, 2008)
Patients with CVID often have defects in post-antigenic B-cell differentiation:
-- a reduction of CD27+ memory Bcells;
-- impaired class switching;
-- expansion of CD21low immature Bcells;
-- poor diferentiation to plasma cells. (Warnatz et al.)
Date
1999
Clinical case
History
Male was born to healthy unrelated parents. Delivery at term.
Body weight: 3 530g. Early neonatal period was without
features. At his first year he grown up and developed according
to his age, get over some respiratory infections (not severe
disease course).
December (4 years 9 months) Pt had an infectious mononucleosis
(diagnose was made on clinical data: fever, intoxication,
2003
lymphadenopathy, hepatosplenomegaly, in blood there were
found atypical mononuclear cells – 28%).
On the background of which appears:
pneumonia in left S5,
thrombocytopenia 12,000 per liter,
agranulocytosis 100* 10(9)/L.
That was the reason for admittance to our hospital.
Date
Clinical case
In our hospital pneumonia progressed
February
(despite antibacterial therapy);
2004
It was supposed to be tuberculosis;
The patient was sent for clinical examination
and treatment at the Institute of Tuberculosis,
where specific treatment was not effective.
The effect was reached at single dosing IV Ig
(0,8g/kg) + Zienam (Imipenem), prednisolone
2mg/kg.
Tuberculosis was excluded;
March
2004
Consequently appeared hemolysis on the basis
of antituberculous therapy.
Lab Evaluation
Thr-penia: 12,000/L
Leu-penia:
3,3*10(9)/L
Agran-cytosis:
100* 10(9)/L,
Anemia: Hbg 92 g/L
NORMAL:
Urine analysis,
electrolyte levels,
lymphocyte subsets,
Ig concetration, blood
coagulation, Liver
enzymes.
Date
Patient’s data
Autoimmune Manifestation
From
Immune thrombocytopenia (ITP) is mediated by
platelet autoantibodies that accelerate platelet
December
destruction and inhibit their production.
2003
to April
2004
Was stopped by prednisolone, IV Ig
Autoimmune haemolytic anaemia(AIHA) is an
From
April 2004
to
December
2004
immune disorder is an immune disorder
caused by antibodies directed against
unmodified autologous red cells.Most AIHA
are caused by warm antibodies, whereas cold
antibodies are less commonly detected.
Wien Klin Wochenschr. 2008;120(5-6):136-51.
Repeated hemolytic crises (at the attempt to
reduce the dose of corticosteroids)
Lab Evaluation
BM cytology:
a lot of
megakaryocyte,
absent
functioning it’s
Reticulocytosis:
140 ‰,
norma 5-10‰;
Haptoglobin:
<13g/L
norma > 30
Anemia:
Hbg: 40 g/L;
Coombs test:
direct positive 1:8
Date
December
2004
Intractable hemolysis
Severe hemolytic crisis was the reason for
admittance to hospital (intensive care unit)
Our patient with severe autoimmune hemolysis
with both cold and warm antibodies was
previously refractory to conventional treatments
was treated with weekly infusions of
Rituximab.
The only treatment with steroids, was allowed
during the period of rituximab administration.
Rituximab reacts specifically with the CD20
antigen and induces B-cell depletion. This could
interfere with the production of autoantibodies
in some immune diseases.
Treatment
IV Ig 0,2 g/kg
Prednisolone
2mg/kg
Rituximab
375mg/m2,
for 2 weeks
Date
Patient Data
Treatment Result
April
2005
Complete hematologic response (+90)
Hbg: 112 g/L
Hemolysis was stopped, Hb became normal (+10). Coombs test:
The hematologic improvement was prompt,
negative
appearing by the second infusion of rituximab.
June
2005
(8-12 times ) watery stool with negative
bacterial seeding.
Skin: hyperemic spotswith desquamation
(atopy) possible reaction to medicines.
Despite intensive antibiotic therapy, IVIg the
patient is constantly ill (sinusitis, ethmoiditis)
Secondary malabsorbtion syndrome with
secondary lactase deficiency was revealed;
Clinically important electrolytic disturbances;
Physical development delay,
Osteoporosis (as manifestation of secondary
hyperparathyroidism)
Hypoelectrolytemia
(K, Ca, Ph, Mg,)
Patient’s Lab Data
Recovery humoral immunity
IV Ig replacement 0,2g/kg
1,2
1
0,8
0,6
0,4
0,2
0
m
on
th
-4
m
on
th
-3
m
on
th
-2
m
on
th
-1
m
on
th
+
1
m
on
th
+
5
m
on
th
+
9
m
on
+
th
12
m
on
+
th
24
m
on
+
th
36
m
on
+
th
48
m
on
th
IgM
IgA
-6
Ig g/L
Evolution of Ig concentration
Period of time before Rituximab and after
2004
2005
2007
2009
Recovery humoral immunity
Evolution of B cells % in PB
25
20
15
10
5
0
m
on
th
-4
m
on
th
-3
m
on
th
-2
m
on
th
-1
m
on
th
+
1
m
on
th
+
5
m
on
th
+
9
m
on
+
th
12
m
on
+
th
24
m
on
+
th
36
m
on
+
th
48
m
on
th
CD19+
-6
% frome CD45+CD14-
Patient’s Lab Data
Period of time before Rituximab and after
2004
2005
2007
2009
CVID
Diagnosis
Humoral immunity has not recovered during 4 years and 4 months
after Rituximab treatment!!
After absence of recovery of humoral immunity was suggested CVID as
diagnosis
The well-accepted definition of CVID includes
three key features:
1) the presence of hypogammaglobulinaemia
of two or more immunoglobulin isotypes
(low IgG, IgA, or IgM),
2)recurrent sinopulmonary infections,
3) impaired functional antibody responses
In addition to these date, there can be other clinical findings
including autoimmunity, granulomatous disease, and
neoplasia.
Common variable immunodefi ciency: a new look at an old disease, Lancet, 2008
Pt has decreasing of
three isotypes
Pt has constant sinusitis,
ethmoiditis
Pt (I-0):
α-antibodies - very low
β-antibodies - absent
Pt – Evans syndrome
Date
Diagnostics procedures
Patient’s Bone Marrow Investigation
Healthy Donor
Pro-B cells
April
2008
B-cell
precursors
4,7%
4,9%
Pre-B
cells
+52months
after
treatment
55,9%
Patient
Pro-B cells
0%
0.09%
B-cell
precursors
0%
Pre-B cells
0,05%
CD19
HANDBOOK OF DIAGNOSTIC
HEMATOPATHOLOGY TESTS, 2001
CD34
Gated on CD45+CD14-
CD34
Date
Diagnostics procedures
Patient’s Bone Marrow Investigation
Healthy Donor
Data.011
Pro-B cells
February
7,5%
2009
+62months
after
treatment
17,1%
Pre-B1 cell
100
101
R1
102
103
CD19 Per CP
75,4%
104
Patient
Pro-B cells
3%
28,2%
2,3%
Pre-B1 cell
69,1%
CD19
CD34
Patient data
Extended Immunological Investigations
Memory B cells (CD19+CD27+IgD-) detection
Healthy
Donor
57,3%
11,1%
95,9%
4,02%
18,6%
Patient
97,3%
1,9%
51,4%
48,6%
0%
IgD
CD21
Patient data
Mutational analysis
Resequencing of 1-5 exons of TNFRSF13B gene has not revealed any
mutations;
The only nucleotide substitution was synonymous homogenic SNP in
exon 2 : rs8072293 C/T;
Patient data
Extended Immunological Investigations
Phenotyping of naïve and memory CD4 T cells
(gated on CD3+)
Patient
CD4+CD45RA+
CD4+CD45RO+
14,5%
CD45RA
35,9%
CD45RO
The immune dysregulation in our patient was extended to a reduction
of naive CD4+CD45RA+ T cells
Summary
At present moment our Pt’s intestinal problems (severe malabsorbtion)
occupy the first place.
Since 2005 Pt has had 8-12 times stool in a day.
Most days of his life he spends in our Hospital.
The Pt moves in hospital room with great difficulties (severe osteoporosis).
He has stopped to grow since 2005 and has poor quality of life.
-- According to literature data our patient may have CVID (severe
phenotype) but usually such patient’s respond to IV Ig replacement therapy,
treatment with steroids etc.
-- Severe clinical data is a full manifestation of CVID in first age-specific
period or complication of rituximab administration?
-- We observed B-cells populations disturbance in BM of patien’t with CVID
before but have never seen absolute absence of CD19+ in BM ?
-- We do not know if it is possible to regarded that synonymous homogenic
SNP in exon 2 : rs8072293 C/T formed such severe phenotype?
-- It is possible to consider BMT as curable therapy?
Thank you for your attention
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