Transcript T cells - The American Society of Pediatric Hematology/Oncology

Immunology and Immunodeficiency
Sung-Yun Pai MD
Some points
• Rest assured, this course (in my biased opinion)
prepares you well for boards
• The content in this talk only ~5% of the questions
• Focuses on IMPORTANT cells of the blood—the
LYMPHOCYTES!
• Unfortunately the board questions seem to lag behind
the content specs so some “old” content specs are
included
• * in the corner means there are some content specs on
that slide
• I have tinkered with this lecture in response to previous
years’ feedback
Objectives
 Identify the cellular basis of immune compromise in
patients undergoing chemotherapy or HSCT, the
classes of infections associated with various cellular
deficiencies, and general principles of management
of infections in immunocompromised hosts
 Identify the normal pattern of development of the
adaptive immune system in children and the
presentation and diagnostic features of specific
congenital deficiencies of adaptive immunity
Content to be reviewed
 Infections in the immunocompromised host
 Normal T and B cell development and function and
evaluation of recurrent infection
 Congenital immunodeficiency states made easy
 (2 slides on AIDS)
2
I
N
N
A
T
E
thymus
A
D
A
P
T
I
V
E
CD16/
56
neutrophils
 Phagocytosis
 Produce bactericidal
reactive oxygen species
macrophages
 Phagocytosis
 Antigen presentation
 Clearance of debris
dendritic cells
 Antigen presentation
 Phagocytosis
natural killer
(NK) cells
CD4
CD3/CD4 T
cell
CD8
CD3/CD8
T cell
CD19
B cells
 Kill virally infected cells
 Kill tumor or foreign cells
 Cellular immunity
 Help B cells class switch
 Help CD8 T cells kill
 Cellular immunity
 Cytolytic activity
 Make antibody
 Antibodies opsonize bacteria
 Antigen presentation
3
neutrophils
I
N
N
A
T
E
thymus
A
D
A
P
T
I
V
E
macrophages
BACTERIA
FUNGI
contribute to all
dendritic cells
CD16/
56
CD4
CD8
CD19
natural killer
(NK) cells
CD3/CD4 T
cell
CD3/CD8
T cell
B cells
VIRUSES
(esp DNA viruses)
VIRUSES
PNEUMOCYSTIS
FUNGI
BACTERIA (esp
encapsulated)
4
What host factors cause immunocompromise?
• Loss of physical barriers
• Medication
• Acquired or congenital defects in cell number
• Acquired or congenital defects in cell function
5
Chemotherapy and HSCT cause
multiple hits to the immune system
• Loss of physical barriers
mucosal impairment
central venous catheters etc.
• Medication
corticosteroids (ALL, GVHD)
specific immunosuppressants (CsA, others)
antibody therapy (ATG, rituximab, alemtuzumab)
• Acquired or congenital defects in cell number
neutropenia, CD4 lymphopenia
• Acquired or congenital defects in cell function
6
Timing of infections post-allogeneic HSCT
BACTERIA
Pre-engraftment
(day 0-30)
Post-engraftment
(early day 30-100)
Post-engraftment
(late day >100)
Gram +
Gram Anaerobes
Same
Esp catheter
associated
Encapsulated
organisms*
FUNGI
Candida
Aspergillus
Aspergillus*
Aspergillus*
VIRUSES
HSV
Respiratory
CMV*
Respiratory
EBV-LPD
VZV
CMV*
Respiratory
EBV-LPD
OTHER
DEFECT
Pneumocystis
Toxoplasma
• profound
neutropenia
• catheter
• lymphopenia esp CD4
• T cell suppressive meds
• aGVHD
• catheter
* risk enhanced or associated with GVHD acute or chronic and its treatment
*
Pneumocystis
Toxoplasma
• lymphopenia esp CD4
• T cell suppressive meds
• cGVHD
• Poor Ig production
7
Management of infection in
immunocompromised hosts
Bacterial infection
Fungal infection
Pneumocystis
Viral infection (especially DNA viruses)
Note: these are not meant to be the absolute most up to date recommendations and are geared
towards what I think are testable principles supported by evidence
8
Bacterial infection
*
In chemo/HSCT patients bacteria are derived from endogenous
flora, in context of mucosal and skin barrier disruption
Epidemiology has shifted from GNR to GPC
GNR: E. coli, Pseudomonas, Enterobacter, Klebsiella, Serratia, other
GPC: Staph aureus, coagulase negative Staph, alpha-hemolytic strep (S. mitis, S.
sanguis), Enterococcus
anaerobes: Bacteroides, Clostridium
Association of Ara-C and alpha-hemolytic strep with ARDS and
rapidly evolving sepsis syndrome
Keep in mind resistant organisms and local pattern
methicillin-resistant S. aureus, vancomycin-resistant enterococcus
inducible plasmids encoding extended spectrum beta lactamases causing resistance
after exposure to cephalosporins in Enterobacter and other GNR
9
*
Bacterial infection
Prophylaxis:
Randomized controlled trials and meta-analyses demonstrate decrease in infection and
possibly improved survival with prophylactic antibiotics in neutropenic cancer
patients
Benefits of use must be weighed against risk of developing resistance
Principles of empiric therapy:
Fever and neutropenia in setting of chemotherapy is an emergency
Instituting immediate therapy without awaiting culture results is standard
Physical examination to identify localizing source must be performed but rarely
identifies cause
Regimen should cover enteric GNR, S. aureus, other gram positives
monotherapy is the standard recommendation (3rd gen cephalosporin,
carbapenem, piperacillin/tazobactam)
Additional agents (aminoglycoside, quinolone, more GPC coverage) upfront or delayed
depending on clinical circumstances
Stopping antibiotics after 7 days in persistently neutropenic patients leads to about
40% of patients developing fever or hypotension
10
*
Random interlude about the spleen
o Several immunological functions of the spleen
o Site of filtration and innate immunity
o Opsonization of encapsulated organisms
o Consumption by macrophages
o Site of primary and secondary adaptive immune responses
o T cells get primed by APC
o T cells interact with B cells, which make specific IgM
o Activated follicular B cells mature, class switch, undergo
affinity maturation
o Site of specialized B cell population
o Marginal zone B cells that respond to polysaccharide
antigens and are generated by 2 years of age
11
Anatomic/functional asplenia
*
Prophylaxis: oral penicillin twice a day for <5 years old
Lifelong for anyone who has had post-splenectomy sepsis
Consider for >5 years old for first 1-2 years after splenectomy
Principles of empiric therapy:
Give antibiotics within 2 hours, at home if >2h from hospital
For hospital: parenteral 3rd or greater generation cephalosporin
For home: amoxicillin 45 mg/kg (max 2g) or levofloxacin 10 mg/kg (max 750 mg)
Principles of vaccination:
Special schedules for pneumococcus, HiB, meningococcus
ACIP recommendations for immunocompromised (functional or anatomic asplenia,
including sickle cell disease, HIV+, CSF leak, cochlear implant) 2012
Catch up everyone with PCV13 if PCV13 naïve
Catch up everyone with PPSV23 if PPSV23 naïve
If naïve to both, give PCV13 then 8 weeks later PPSV23
2nd dose of PPSV23 after 5 years
If PPSV23 in past, but naïve to PCV13, give PCV13 >8 weeks from last PPSV23
12
*
Fungal infection
Two major populations at risk, with propensity for different fungi
• profound and long neutropenia
(Candida, Aspergillus, others)
• defective cellular immunity
(Cryptococcus, Histoplasma, Coccidiodes)
Other rarer fungi include:
• Fusarium, Mucor, Trichosporon, dematiaceous molds
Unlike bacterial infection, these are not endogenous but patients
become colonized due to exposure and modulation of normal
flora
13
*
Fungal infection
Prophylaxis:
Large trials have shown decrease in fungal infections in adults during leukemia
induction or undergoing HSCT
Current IDSA recommendations are to use an azole (fluconazole, itraconazole,
voriconazole, posaconazole) or echinocandin (micafungin, caspofungin) for
prevention of Candidiasis during induction or HSCT (neutropenia >7 days)
Patients with CGD benefit from prophylactic treatment
Principles of empiric therapy:
Empiric anti-fungal therapy is standard for persistent fever and neutropenia (4 to 7
days)
Other possible signs of fungal disease include:
nasal or sinus tenderness, painful swallowing, pulmonary infiltrates
Therapy should be directed against Candida and Aspergillus, and no one agent is
specifically recommended.
Choice of empiric agent should take into account prophylactic regimen, known
colonization, individual patient profile and local epidemiology
14
*
Pneumocystis jirovecii
(formerly called Pneumocystis carinii)
risk factors for Pneumocystis pneumonia include:
HIV infection with CD4 count <200
HSCT recipients
cancer especially leukemia
solid organ transplant recipients
glucocorticoids, chemo or other immunosuppressants
primary immunodeficiency especially SCID
(renamed because P. jirovecii infects humans while P. carinii infects other
mammals, like rodents)
15
*
Pneumocystis jirovecii
Prophylaxis:
Trimethoprim-sulfamethoxazole
150 mg TMP/m2/day (5 mg/kg/day) divided BID TIW consecutive days most tested
Alternatives to TMP/SMX:
atovaquone, dapsone, aerosolized pentamidine, intravenous pentamidine,
clindamycin plus primaquine, sulfadoxine plus pyrimethamine
few if any studies in non-HIV patients comparing efficacy
Treatment: Trimethoprim-sulfamethoxazole
15-20 mg/kg/day divided TID-QID, PO or IV for 14-21 days
adjunctive corticosteroids shown to be effective in HIV, less certain in non-HIV
Alternatives to TMP/SMX:
mild disease: atovaquone, severe disease: pentamidine IV, clindamycin plus
primaquine
16
*
Viral infection
Patients with poor cell mediated immunity are at high risk for
infection due to DNA viruses
• post-BMT
• solid organ transplant
• HIV
• primary immunodeficiency
CMV, HSV, VZV, EBV can all cause disease from primary exposure,
or from reactivation of latent infection
17
*
Viral infection
Cytomegalovirus
Herpes family DNA virus that causes asymptomatic or self-limited flu/mononucleosis
illness in competent host
Can be passed through organs, blood, breastmilk, saliva, urine
Disease manifests as viremia, interstitial pneumonitis, colitis, retinitis, hepatitis,
esophagitis/gastritis
Asymptomatic shedding in urine and saliva can occur
Organ disease and viremia/blood antigenemia are not always correlated
Prophylaxis
using only blood products from CMV negative donors OR leukofiltration
in BMT setting, treatment with ganciclovir prevents interstitial pneumonia
Treatment
ganciclovir 5 mg/kg/dose IV twice a day for 2 weeks
valganciclovir orally, dosing is not well established in children
foscarnet 180 mg/kg/day divided twice or three times a day for 2 weeks
cidofovir does have activity against CMV
18
*
Viral infection
Varicella zoster (Herpes zoster)
Herpes family DNA virus that causes primary varicella and shingles
Primary or reactivation disease can be severe in immunocompromised patients,
causing hepatitis, pneumonitis, encephalitis
Prophylaxis
universal vaccination began in US in 1995
acyclovir, valacyclovir, famciclovir can all be used to prevent VZV reactivation
in the absence of Varicella specific Ig (VZIG), pooled IVIg can be used to prevent
disease after exposure in non-immune children up to 96h post
Treatment
high dose acyclovir, 1500 mg/m2/day IV divided every 8 hours, adjust for renal
insufficiency and hydrate
oral acyclovir or other anti-virals
19
Content to be reviewed
 Infections in the immunocompromised host
 Normal T and B cell development and function and
evaluation of recurrent infection
 Congenital immunodeficiency states made easy
 (2 slides on AIDS)
20
Lymphocytes contribute to both
innate and adaptive immune systems
CD16/56
NK cell
thymus
*
INNATE
CD4
helper T
cell
CD8
cytolytic
(killer)
T cell
CD19
B cell
ADAPTIVE
21
Major arms of the adaptive immune system *
Cellular immunity
T cells (most are a/b T cells)
Humoral immunity
Express TCR with CD3 complex (TCRa,
TCRb, CD3zz, CD3ed, CD3eg)
B cells
-CD4 T cells
Produce antibody
see antigens complexed to MHC class II on APC
helper function
cooperate with B cells to induce class switching
control opportunistic infection
cancer surveillance
regulatory functions
IgM and IgD first
IgG, IgA, IgE only after class switching
additional mutations in Ig locus occur—B cells
carrying receptors with higher affinity are
selected (affinity maturation)
-CD8 T cells
see antigen complexed to MHC class I on APC
cytolytic function
kill virally infected cells
control opportunistic infection
cancer surveillance
Antibodies opsonize bacteria
particularly important for
•respiratory pathogens
•skin flora
•encapsulated organisms
22
*
What holes are generated in the immune system when
lymphocytes are absent or do not function?
Cellular immunity
Humoral immunity
T cells
B cells
-CD4 T cells
Produce antibody
loss of Ig production
opportunistic infections
fungal infections
viral infections
-CD8 T cells
lack of killer function
viral infection
especially DNA viruses
cancer surveillance
Antibodies opsonize bacteria
inability to opsonize bacteria
especially respiratory,
skin, encapsulated organisms
23
How does the adaptive immune system develop?
Blood
Bone marrow
Thymus
TCR
TCR rearrangement
HSC
CD3
HSC
T
T
PreT
ProB
TCR
T
BCR (Ig) rearrangement
PreB
CD4
preBCR
CD3
proliferation
CD8
T
BCR signaling
IgM
Spleen
IgM
CD19
immB
B
24
IgD
How does the adaptive immune
system adapt?
Antigen
TCR
TCR
rearrangement
Helper function:
B cell help
T cell help
CD4
T
T
TCR
HSC
T
ProB
BCR (Ig)
rearrangement
proliferation
CD8
Killer function:
Lyse infected cells
T
IgM
PreB
immB
Antigen
Antigen
IgM
CD19
immB
Ig class
switch
Affinity
maturation
IgG
IgA
B
IgD
Spleen &
Lymph node
IgM
25
25
*
How to measure immunologic function?
Enumeration
total lymphocytes, subsets
absolute lymphocyte count
CD3 = T cells
CD4 = helper T cells
CD8 = cytolytic T cells
CD19 = B cells
CD16 or CD56 = natural killer (NK) cells
26
Normal values for lymphocyte subsets in
children and adults
*
• ALC, CD3 and CD4
counts are higher in
young children
• B cell numbers are
maximal 2 months to
2 years then contract
• NK cell numbers are
pretty stable
27
Infant versus adult lymphocyte subsets
Adult
Infant (1 wk to 2 mo)
ALC
1800 (1000-2800)
ALC
6700 (3500-13100)
CD3
1200 (700-2100)
CD3
4600 (2300-7000)
CD4
700 (300-1400)
CD4
3500 (1700-5300)
CD8
400 (200-900)
CD8
1000 (400-1700)
CD19
200 (100-500)
CD19
1000 (600-1900)
CD56
300 (90-600)
CD56
500 (200-1400)
ALC of less than 1000-2000 in an infant is highly abnormal.
values from Comans-Bitter J Peds 1997
28
*
How to measure immunologic function?
Non-specific function
T cells
Proliferation to mitogens
B cells
Total IgG, IgA, IgM
phytohemagglutinin (PHA)
pokeweed mitogen (PWM)
concanavalin A (ConA)
IgG subclasses
IgG1, IgG2, IgG3, IgG4
Remember that antibody production also requires intact T
cell function
29
*
*
How to measure immunologic function?
Specific function
T cells
Proliferation to antigen
tetanus, Candida
Skin testing
Candida control
B cells
Antigen-specific antibody
protein Ag
• (tetanus, hepBsAg)
carbohydrate antigen
• (blood group, pneumovax)
Remember that antibody production also requires intact T
cell function
30
Immunoglobulins for dummies
*
Class
IgM
IgG
IgA
Structure
Pentameric
Monomeric
Mono-, dimeric
Affinity
Low affinity
High affinity
Low affinity
Placental
transfer?
No
Yes
No
%
5-10%
75-85%
5-15%
* IgD does not require class switching, no clear function
* Levels of IgE not necessarily part of an immunodeficiency work-up
31
*
Normal immunoglobulin for age (IgG)
1800
1600
physiologic nadir
2-6 months
1400
1200
1000
800
600
400
200
0
low
NB
636
high 1606
1 mo 2 mo 3 mo 4 mo 5 mo 6 mo 9 mo
251
206
176
196
172
215
217
906
601
581
558
814
704
904
12
mo
2 yr
3 yr
4 yr
6 yr
8 yr
294
345
424
441
463
633
10 yr 18 yr
608
639
1069 1213 1051 1135 1236 1280 1572 1349
32
350
Normal immunoglobulin for age (IgA)
300
250
200
150
100
50
0
NB
1 mo 2 mo 3 mo 4 mo 5 mo 6 mo 9 mo
12
mo
2 yr
3 yr
4 yr
6 yr
8 yr
10 yr 18 yr
low
1.4
1.3
2.8
4.6
4.4
8.1
8.1
11
16
14
14
22
25
33
45
70
high
3.6
53
47
46
73
84
68
90
84
106
123
159
154
202
236
312
33
*
*
Normal immunoglobulin for age (IgM)
400
350
300
250
200
150
100
50
0
NB
1 mo 2 mo 3 mo 4 mo 5 mo 6 mo 9 mo
12
mo
2 yr
3 yr
4 yr
6 yr
8 yr
10 yr 18 yr
low
6.3
20
17
24
27
33
35
34
41
43
48
47
43
48
52
56
high
25
87
105
89
101
108
102
126
149
173
168
200
196
207
242
352
34
Take home points about normal lymphoid
development
 infants and children have higher ALC and T cell numbers than
adults
 IgG crosses the placenta
 physiologic nadir of IgG occurs around 2-6 months
35
*
Content to be reviewed
 Infections in the immunocompromised host
 Normal T and B cell development and function and
evaluation of recurrent infection
 Congenital immunodeficiency states made easy
 (2 slides on AIDS)
36
*
Congenital Immunodeficiency in one slide
Absent
Present/Broken
T cells
SCID
Ex: WAS
B cells
XLA
Ex: CVID
Neutrophils
SCN
Ex: CGD
37
Note: patients lacking NK cells have been described but are very rare
Congenital Immunodeficiency
 Failure of T cell development
SCID (severe combined immunodeficiency)
 Failure of B cell development
XLA (X-linked agammaglobulinemia)
 Functional T, B, or combined defects
T and B cells present but not fully functional
Wiskott-Aldrich syndrome (WAS)
X-linked Hyper-IgM syndrome (CD40LG)
X-linked lymphoproliferative disease (XLP)
Common variable immunodeficiency (CVID)
Autoimmune lymphoproliferative syndrome (ALPS)
38
*
Congenital Immunodeficiency
 Failure of T cell development
SCID (severe combined immunodeficiency)
 Failure of B cell development
XLA (X-linked agammaglobulinemia)
 Functional T, B, or combined defects
T and B cells present but not fully functional
Wiskott-Aldrich syndrome (WAS)
X-linked Hyper-IgM syndrome (CD40LG)
X-linked lymphoproliferative disease (XLP)
Common variable immunodeficiency (CVID)
Autoimmune lymphoproliferative syndrome (ALPS)
39
*
*
Definition of SCID
“genetically heterogeneous group of syndromes having in common
a profound disturbance in the development and function of both
T and B cells”
• Absence of functional autologous T cells is uniform
• B cells can be present or absent
• B cells even if present usually don’t function, i.e. no specific
antibody production
• generally fatal before age 2 due to overwhelming infection
40
*
When to suspect SCID
Symptoms
Signs/Lab findings
• Failure to thrive
• thrush
• Chronic diarrhea
• absence of lymphoid tissue
• Recurrent infections
• opportunistic infection ex: PCP
Associated findings
• low or absent T cells
• FH ex: X-linked
• lack of immunoglobulins
• neurologic (ADA)
• lack of thymic shadow
• cardiac (DiGeorge)
41
*
How to diagnose SCID
Check ALC, send lymphocyte subsets
T cells very low or absent
Send lymphocyte proliferation studies
Absent or very low proliferation of T cells to mitogens
Total IgG, IgA, IgM and specific Ab if vaccinated
42
*
SCID with normal ALC?
SCID with T cells present?
4 month old boy with chronic cough and FTT.
Diagnosed with PCP.
CBC:
WBC 12
60% polys
35% lymphs
ANC = 7200
ALC = 4200
CD3 = 400
CD4 = 8
CD8 = 300
(2300-6500)
CD19 = 3595
(600-3000)
CD56= 15
(100-1300)
(1500-5000)
(500-1600)
(3700-9600)
FISH XX/XY = 2% XX
proliferation to mitogens: flat
IgG 87 (196-558)
43
*
How to diagnose SCID
Check ALC, send lymphocyte subsets
Because B cells can be present, normal ALC does not rule out SCID
Because maternal engraftment can occur, presence of T cells does not rule out
SCID
Send lymphocyte proliferation studies
Maternal T cells do not proliferate. Normal proliferation rules out classic SCID.
Total IgG, IgA, IgM and specific Ab if vaccinated
Molecular defect can be narrowed down based on
lymphocyte phenotype:
T-B+ (can be NK - or +)
T-B- (can be NK - or +)
44
*
T- B+ SCID
Molecular defects:
cytokine common gamma chain (IL2RG, gc, CD132)
JAK3 (JAK3, Janus kinase 3)
IL-7 receptor alpha chain (IL7R, IL-7Ra, CD127)
B cells are present but receive no help, generally do not
produce antibody
gc has X-linked inheritance, accounts for about 1/3 of all SCID
JAK3 and IL-7Ra are autosomal recessive
45
*
Lack of growth factor signaling leads to absence of T cells
gc
IL-7Ra
gc
46
Normal adaptive immune system development
Blood
Bone marrow
Thymus
TCR
TCR rearrangement
HSC
CD3
HSC
T
T
PreT
ProB
TCR
T
BCR (Ig) rearrangement
PreB
CD4
preBCR
CD3
proliferation
CD8
T
BCR signaling
IgM
Spleen
IgM
CD19
immB
B
47
IgD
Lack of cytokine signaling causes loss of T cells
Blood
Bone marrow
Thymus
TCR rearrangement
HSC
HSC
ProB
BCR (Ig) rearrangement
PreB
preBCR
X
proliferation
BCR signaling
IgM
Spleen
IgM
CD19
immB
B
48
IgD
*
T- B- SCID
Molecular defects:
Adenosine deaminase (ADA)
Recombinase activating gene 1, gene 2 (RAG1, RAG2)
Artemis (DCLRE1C, DNA crosslink repair 1C), other
DNA repair enzymes
All autosomal recessive inheritance
RAG1/2, Artemis have NK+ phenotype
ADA deficiency generally causes NK- phenotype
49
*
TCR/BCR
RAG
Artemis
Lack of rearrangement machinery leads to absence
of both T and B cells
50
Lack of rearrangement machinery leads to absence of
both T and B cells
Blood
Bone marrow
Thymus
X
TCR rearrangement
HSC
HSC
ProB
X
BCR (Ig) rearrangement
proliferation
Spleen
51
*
Accumulation of toxic metabolites leads to
absence of all lymphocytes, T, B, NK
dAdenosine
ADA
dATP
dInosine
PNP
Hypoxanthine
Xanthine
Uric acid
52
gc
TCR/BCR
RAG
Artemis
gc
dAdenosine
ADA
dATP
dInosine
PNP
Hypoxanthine
Xanthine
Uric acid
53
Diagnosis and management of SCID

supportive management
protection from infection
IgG replacement
aggressive diagnosis and prompt treatment of infection
prophylaxis against Pneumocystis

ALL BLOOD PRODUCTS MUST BE IRRADIATED to avoid fatal
transfusion-associated GVHD

Consider enzyme replacement with PEG-ADA for ADA deficient
patients
Immediate referral for curative treatment
i.e. HSCT*
* except complete DiGeorge, which is often treated with thymic transplant
54
*
SCID and HSCT for boards purposes
*
• a variety of donors and conditioning approaches can be used
to cure SCID with HSCT
donor
conditioning?
T cell depletion?
*matched sibling
no
no
haploidentical
parent
no
YES
unrelated
YES
to prevent lethal GVHD
either
55
Take home points about SCID
 definition of SCID is lack of FUNCTIONAL AUTOLOGOUS T cells
an ALC of <2000 in an infant is very abnormal
maternal engraftment is common
i.e. presence of T cells does NOT rule out SCID
maternal T cells will not proliferate
 B cells even if present do not function
i.e. SCID is in differential of very low IgG
 inheritance may be X-linked or AR
 lack of all lymphocytes (T, B, NK) should make you think of
ADA
56
*
Congenital Immunodeficiency
 Failure of T cell development
SCID (severe combined immunodeficiency)
 Failure of B cell development
XLA (X-linked agammaglobulinemia)
 Functional T, B, or combined defects
T and B cells present but not fully functional
Wiskott-Aldrich syndrome (WAS)
X-linked Hyper-IgM syndrome (CD40LG)
X-linked lymphoproliferative disease (XLP)
Common variable immunodeficiency (CVID)
Autoimmune lymphoproliferative syndrome (ALPS)
57
*
*
X-linked agammaglobulinemia (XLA)
 X-linked disorder of B cell development due to mutation of
the BTK (Bruton’s tyrosine kinase) gene which signals
downstream of the pre-B cell receptor
 B cell development is arrested in the bone marrow at the
pre-B cell stage resulting in complete absence of peripheral
B cells
 IgG levels fall after maternal IgG naturally declines, pyogenic
infections with bacteria occur starting 6-9 months
 prone to chronic enteroviral meningoencephalitis
 avoid live virus vaccines
58
Normal adaptive immune system development
Blood
Bone marrow
Thymus
TCR
TCR rearrangement
HSC
CD3
HSC
T
T
PreT
ProB
TCR
T
BCR (Ig) rearrangement
PreB
CD4
preBCR
CD3
proliferation
CD8
T
BCR signaling
IgM
Spleen
IgM
CD19
immB
B
59
IgD
Loss of BCR signaling abolishes B cell development
Blood
Bone marrow
Thymus
TCR
TCR rearrangement
HSC
CD3
HSC
T
T
PreT
ProB
TCR
T
BCR (Ig) rearrangement
PreB
CD4
preBCR
CD3
proliferation
CD8
T
X
BCR signaling
IgM
Spleen
60
*
X-linked agammaglobulinemia (XLA)
 laboratory shows absence of CD19 cells on lymphocyte
subsets, low to absent IgA, low to absent IgM, low to absent
IgG depending on age (placental transfer)
 T cell immunity (T cell numbers, skin reaction, proliferation
to mitogens, control of opportunistic infections) is intact
 mainstay of treatment is IgG replacement with dramatic
decrease in incidence of infections
61
Congenital Immunodeficiency
 Failure of T cell development
SCID (severe combined immunodeficiency)
 Failure of B cell development
XLA (X-linked agammaglobulinemia)
 Functional T, B, or combined defects
T and B cells present but not fully functional
Wiskott-Aldrich syndrome (WAS)
X-linked Hyper-IgM syndrome (CD40LG)
X-linked lymphoproliferative disease (XLP)
Common variable immunodeficiency (CVID)
Autoimmune lymphoproliferative syndrome (ALPS)
62
*
*
Wiskott-Aldrich syndrome (WAS)
 X-linked disorder due to mutation of the WAS protein (WAS,
WASP) gene
 classic triad of immunodeficiency, eczema and
thrombocytopenia with small platelets (ex: MPV 4-5
compared to normal about 7-8 fl)
 not all patients especially when young manifest all parts of
the triad, in particular, immune defect tends to worsen with
time
 most boys die by young adulthood from bleeding, infection,
autoimmunity or lymphoma
63
Wiskott-Aldrich syndrome (WAS)
 immune defect is combined T and B
typically normal to low T and normal B cell numbers
T cells can be functionally abnormal
abnormal proliferation to mitogen
abnormal proliferation to antigen
B cell function more often abnormal than T
poor specific antibody production
classically lacks carbohydrate responses
i.e. isohemagglutinins, pneumovax
Total IgG declines over time
Allergic manifestations including high IgE also seen
64
*
*
Wiskott-Aldrich syndrome (WAS)
 not all WASP mutations lead to WAS
 complete lack of WAS protein usually leads to WAS
 missense mutations with detectable protein may lead to
X-linked thrombocytopenia (XLT)
small low platelets
no or mild eczema
normal immune function
some patients may develop autoimmunity
 missense activating mutations in GTPase binding domain
X-linked neutropenia (XLN)
normal platelets, lymphocytes, no eczema
severe neutropenia, with variable other findings
(myelodysplasia, large platelets, reversal of CD4/CD8 ratio)
65
Wiskott-Aldrich syndrome (WAS)
 diagnosis in the old days made by non-random X-linked
inactivation in appropriate clinical setting, now WAS protein
levels and gene sequencing
 splenectomy can correct thrombocytopenia for years
 late manifestations include immune thrombocytopenia (may
occur even after splenectomy), autoimmune arthritis, vasculitis,
lymphoma
 supportive therapy includes Ig replacement, platelet
transfusion, management of immune complications, treatment
of infections
 curable by early well-matched HSCT
66
*
Activated CD4+ T cell
Hyper-IgM syndromes
Proliferation
Cytokines
Activation of DC
 share in common defects in switching from
IgM to other isotypes
 process is dependent on CD40L on T cells
(expressed on activated CD4+ T cells)
interacting with CD40 on B cells (leads to
class switching)
 most common form is an X-linked disorder
due to defect in CD40 ligand (CD40LG,
CD40L, CD154) gene
 hence boys have combined B and T cell
defect with primary manifestation of
hypogammaglobulinemia, low IgG, low IgA
and normal or high IgM
CD40L
CD40
Maturation
Class switching
Somatic hypermutation
Naïve IgM+ B cell
67
*
Hyper-IgM syndromes
(X-linked, CD40LG deficiency)
 infectious profile of recurrent bacterial infections (low
IgG), opportunistic infections such as PCP,
cryptosporidium, histoplasma (poor CD4 cell function)
 lab tests show normal T and B cell numbers, low IgG, low
IgA, normal to high IgM, low CD40L expression after
stimulation of T cells in vitro, no production of specific IgG
after vaccination
 curable by HSCT
68
*
*
X-linked lymphoproliferative disease (XLP)
 X-linked disorder due to mutations in the SAP gene (SLAM associated
protein, SH2D1A), an adaptor protein that associates with the SLAM family of
receptors on T, B, NK and other cells
 pleomorphic presentation with certain clinical scenarios being “classic”
 fulminant often fatal acute mononucleosis due to EBV
 lymphoproliferation usually nonclonal
 recurrent infections with dysgammaglobulinemia or
hypogammaglobulinemia
 typically have normal T and B cell numbers, normal proliferation, may have
high IgA or IgM and low IgG, defective B cell memory generation
69
*
X-linked lymphoproliferative disease (XLP)
 diagnosis requires high index of suspicion, sometimes due to careful review
of family history with X-linked pattern of illness
 detection of SAP protein by intracellular FACS or sequencing of SH2D1A
gene can make the diagnosis
 fulminant EBV infection has an extremely poor outcome
 patients who develop hypogammaglobulinemia should be replaced
 HSCT may be considered in selected patients
70
*
Common variable immunodeficiency (CVID)
 heterogeneous group of disorders with largely unknown
molecular defects, typical onset in adults 20-40 years
 clinical definition is “a genetic immune defect characterized
by significantly decreased levels of IgG, IgA and/or IgM with
poor or absent antibody production, with exclusion of
genetic or other causes of hypogammaglobulinemia”
 unlike XLA, B cells detectable and patients often have
defects in T cell number or function, predisposing to
autoimmunity, granulomatous disease (lungs, intestine), and
lymphoma
71
Common variable immunodeficiency (CVID)
 Like other antibody deficiencies, sinopulmonary infections,
chronic enteroviral meningoencephalitis are common
 Autoimmune disease especially ITP/AIHA is prominent*
ITP
Evans
AIHA
RA
34%
12%
10%
7%
anti-IgA
5%
SLE
4%
alopecia, DM, IBD, 3% each
pernicious anemia,
myasthenia gravis,
autoimmune neutropenia,
primary biliary cirrhosis,
immune urticaria
 treatment is IgG replacement and supportive treatment of
infectious and autoimmune complications
* Cunningham-Rundles. Autoimmunity in primary immune deficiency: taking lessons from our patients. Clin
Exp Immunol (2011) vol. 164 Suppl 2 pp. 6-11.
72
*
Autoimmune lymphoproliferative syndrome (ALPS)
 Lymphoproliferative disease caused by defects in pathways
controlling apoptosis of lymphocytes
 Molecular defects increasingly understood, with majority
due to germline heterozygous defects in FAS or somatic
heterozygous defects in FAS (rarer: FASLG, CASP10, CASP8,
NRAS)
 Chronic nonmalignant noninfectious lymphadenopathy
and/or splenomegaly and elevated CD3+ CD4- CD8- TCRab
cells (DN T cells) in blood (and also typically in LN biopsy)
 Autoimmune cytopenia very common and may be
presenting symptom
73
*
Take home points about non-SCID
immunodeficiencies
*
• Boys with XLA lack B cells but have normal T cell function
• WAS is a triad of small low platelets, eczema, combined T and
B cell dysfunction
• Boys with WAS are at risk for bleeding, infection, lymphoma,
autoimmunity (i.e. these things can be elicited on family history)
• A boy that is super sick with EBV--think XLP
• Patients with antibody deficiencies (i.e. CVID) and ALPS
frequently have autoimmune cytopenias
74
Explanation of the Tables that follow
o These have been abstracted from the publication by
International Union of Immunological Societies that is
updated every several years
o Diseases in bold not already covered in the previous slides
o are in other sections’ content specifications (see arrow)
o were in previous content specifications (board
questions still might occur, see arrow)
o are disorders I could see coming up
o Diseases not in bold are generally rarer and are not in
content specifications, but
o you could potentially see in clinical practice or
o they are just too cool and interesting for me to skip
Modified from Al-Herz W et al. Primary immunodeficiency diseases: an update on the classification from the international union of
immunological societies expert committee for primary immunodeficiency. Front Immunol. 2014;5:162.
75
Severe Combined Immunodeficiency
(B+ subtypes)
Deficiency or
Disease name
Gene name
XL/AR/AD
T
B
Other features
gc (common
gamma chain)
IL2RG
X-linked
-
+
NK -
JAK3
JAK3
AR
-
+
NK -
IL7Ra
IL7RA
AR
-
+
NK +
CD45
PTPRC
AR
-
+
NK -
CD3d
CD3D
AR
-
+
NK +
CD3e
CD3D
AR
-
+
NK +
CD3z
CD3Z
AR
-
+
NK +
Coronin-1-A
CORO1A
AR
-
+
Migration defect,
therefore thymus
present
76
Severe Combined Immunodeficiency
(B- subtypes)
Deficiency or
Disease name
Gene
name
XL/AR/AD
T
B
Other features
Recombinase
activating gene 1
RAG1
AR
-
-
NK +
Recombinase
activating gene 2
RAG2
AR
-
-
NK +
Artemis
DCLRE1C
AR
-
-
NK +, radiation sensitivity
Adenosine deaminase ADA
AR
-
-
NK -, rib flaring, hearing loss,
neuro findings, pulmonary
alveolar proteinosis
DNA PKcs
DNAPK
AR
-
-
Radiation sensitivity, microcephaly
Reticular dysgenesis
(adenylate kinase 2)
AK2
AR
-
-
Severe neutropenia, hearing loss
77
Combined Immunodeficiency
(generally less severe than SCID)
Deficiency or
Disease name
Gene name XL/AR/A Cell profile
D
Ig
profile
Other features
ZAP70
ZAP70
AR
Nl CD4, CD8 T
cell number, Nl B
cell number
Normal
May have
autoimmunity
MHC class I
TAP1, TAP2,
TAPBP
AR
Nl CD4, CD8 T
cell number
Normal
Vasculitis, pyoderma
gangrenosum
MHC class II
CIITA, RFX5,
RFXAP, RFXANK
 CD4, nl CD8 T
cell number
Nl or 
Features like SCID,
FTT, diarrhea,
respiratory
infections,
liver/biliary disease
Purine nucleoside
phosphorylase
PNP
AR
Progressive 
Nl or 
Neurologic
impairment
CD3g
CD3G
AR
Nl with  TCR
expression
Nl
CD8
CD8A
AR
CD8 -, CD4 +
Nl
78
Combined Immunodeficiency
(generally less severe than SCID)
Deficiency or
Disease name
Gene
name
XL/AR/
AD
Cell profile
Ig profile
Other features
Wiskott-Aldrich
syndrome
WASP
X-linked
 T with age
IgG 
with age,
poor specific
Ab, low
isohem
Thrombocytopenia,
small platelets,
eczema,
autoimmunity,
lymphoma
SLAM-associated
protein (SAP, XLP
type 1)
SH2D1A
X-linked
Normal T and May have
B #, reduced hypogamm,
memory B
poor specific
Ab
EBV related
lymphoproliferation,
fatal mono, HLH,
absent NK-T cells
X-linked inhibitor of
apoptosis (XLP type 2)
BIRC4
X-linked
Normal T and B
#
EBV related
lymphoproliferation,
HLH, colitis, IBD
Ataxiatelangiectasia
mutated
ATM
X-linked
 T with age, May have 
Nl B #
May have
hypogamm,
poor specific
Ab
79
Ataxia,
radiosensitivity,
telangiectasia,
lymphoproliferation
Combined Immunodeficiency
(with other major anomalies)
Deficiency or
Disease name
Gene
name
XL/AR/A Cell profile
D
Ig
profile
Other features
Digeorge (lack of
thymus organ
development)
Deletion
of
22q11.2
incl TBX1
AD or
sporadic
 T cell #, in
complete can be
like SCID, nl B
cell #
Variably

Cardiac, jaw,
hypocalcemia;
autoimmunity
CHARGE (coloboma,
heart, atresia choanae,
retarded growth,
genital, ear)
CHD7,
some
unknown
AD or
sporadic
 T cell #, in
complete can be
like SCID, nl B cell #
Variably

Hearing loss,
hypocalcemia,
swallowing defect
CID with multiple
intestinal atresia
TTC7A
AR
 T cell #, in
complete can be
like SCID

Multiple intestinal
atresias,
polyhydramnios
Cartilage-hair
hypoplasia (RNA
component of
mitochondrial RNA
processing
endoribonuclease)
RMRP
AR
 T cell #, in
complete can be
like SCID, nl B
cell #
Variably

Short-limbed
dwarfish, sparse
hair, anemia, BM
failure, lymphoma
80
Hyper-IgM syndromes
Deficiency or
Disease name
Gene
name
XL/AR/A Cell profile
D
Ig
profile
Other features
CD40 ligand (Xlinked hyper-IgM)
CD40LG
X-linked
Nl T and B #, lack
of isotype
switching
IgM nl
or ,
other
isotypes

Neutropenia,
opportunistic
infections (T cell
functional defect),
liver and biliary
tract disease
CD40
CD40
AR
Nl T and B #, lack of
isotype switching
IgM nl or
, other
isotypes

Ectodermal dysplasia
with immunodeficiency
IKGKB
(NEMO)
X-linked
Nl T and B #, lack of
isotype switching,
susceptible to
mycobacteria
IgM nl or
, other
isotypes

Conical teeth, lack of
sweat glands and skin
appendages, frontal
bossing
Activation-induced
cytidine deaminase
(AID)
AICDA
AR
Nl T and B #, lack of
isotype switching;
normal T cell
function
IgM nl or
, other
isotypes

Lymphadenopathy
due to hyperplastic
germinal centers
Uracil-N-glycosylase
UNG
AR
Like AID
Like AID
Like AID
81
Predominantly antibody deficiency
Deficiency or Disease
name
Gene name
XL/AR/AD
B cells
Ig profile
Bruton’s tyrosine kinase
BTK
X-linked
-
All isotypes 
AR
-
All isotypes 
Components of pre-BCR and BCR
(m heavy chain, l5, Iga, Igb)
Other components of pre-BCR
and BCR signaling (BLNK, PI3
kinase, E47 transcription factor)
BLNK, PIK3R1,
TCF3
AR
-
All isotypes 
MDS with
hypogammaglobinemia
May be
associated with
monosomy 7,
trisomy 8 DC
variable
variable
One or more isotypes 
Thymoma associated
Unknown
variable
One or more isotypes 
82
Predominantly antibody deficiency
(CVID and other syndromes)
Deficiency or Disease
name
Gene
name
XL/AR/A
D
Features
Common variable
immunodeficiency
Unknown
Unknown
Low IgG and either low IgA and/or
low IgM
Cell surface molecules involved
in BCR signaling
CD19, CD81,
CD20, CD21
AR
Presents as CVID
Growth factor receptors for B
cells
TACI, BAFF
receptor
AR
Presents as CVID
Selective IgA deficiency
Unknown
Variable
Usually asymptomatic, may have
infections, poor Ab response to
carbohydrate, may have allergies or
autoimmune disease
Transient
hypogammaglobulinemia
of infancy
Unknown
Variable
Low IgG and IgA, may be an
exaggerated “nadir,” typically make
responses to vaccines, usually not
associated with significant infections
Isolated IgG subclass
deficiency
Unknown
Variable
Usually asymptomatic, a minority may have
poor Ab response and infections
83
Complement deficiencies
I truly know next to nothing about these but people keep asking me to include them
Deficiency
Gene
XL/AR/ Pathway
AD
Infections? Autoimmunity?
C1q, C1r, C1s
C1QA,
C1QB,
C1QC,
C1R, C1S
AR
Classical
Encapsulated
SLE
C4
C4A. C4B
AR
Classical
Encapsulated
SLE
C2
C2
AR
Classical
Encapsulated
SLE
C3
C3
AR or AD
Central
Yes
glomerulonephritis
C5, C6, C7,
C8a-g, C8b, C9
C5, C6, C7,
C8A, C8G,
C8B, C9
AR
Terminal
Neisseria
-
Other
atherosclerosis
84
Complement pathway defects
I truly know next to nothing about these but people keep asking me to include them
Protein
Gene
XL/AR/ Effect
AD
Infections? Autoimmunity?
Other
C1 inhibitor
SERPING1
AD
Activation,
consumption
of C4/C2,
bradykinin
-
-
Hereditary
angioedema
Factor B
CFB
Gain of
function
AD
alternative
pathway
-
-
Atypical HUS
Factor D
CFD
AR
Defect in
alternative
Neisseria
-
Properdin
CFP
X-linked
Defect in
alternative
Neisseria
-
Factor I
CFI
AR
alternative,
consumption
of C3
Neisseria,
other
Membranoproliferative
glomerulonephritis
Atypical HUS,
preeclampsia
Factor H
CFH
AR
alternative,
consumption
of C3
Neisseria,
other
Membranoproliferative
glomerulonephritis
Atypical HUS,
preeclampsia
85
Finally--2 slides on AIDS
*
• 90% of infants/children acquire perinatally, typically no
symptoms
• virus preferentially infects CD4 T lymphocytes, progressively
depleted by direct toxicity, thymic atrophy, destruction of
infected CD4 T cells by the immune system
• viral entry is mediated by the CD4 protein itself along with the
chemokine receptor CXCR4
• can also infect monocytes, which may act as a reservoir
• screening of infants of mothers with HIV is problematic
because
1)Maternal antibody is placentally transferred (may circulate up to 18 months)
2)CD4 T cell depletion is not always apparent in the face of normally increased
CD4 counts in infants
• Diagnosis much be made on the basis of antigen testing (p24),
HIV NAT (DNA PCR) and/or viral load (RNA levels)
86
Finally--2 slides on AIDS
Hematologic/immunologic manifestations:
• anemia (multifactorial including viral suppression, drugs, drugs, drugs,
anemia of chronic disease, autoimmune, nutritional)
• thrombocytopenia (less multifactorial including ITP, drugs, TTP, bone
marrow infections such as MAI)
• immune
Depletion of CD4 T cells
B cell dysregulation including excess numbers of B cells and extreme
hypergammaglobulinemia
Malignancies in children with HIV:
• NHL systemic or primary CNS
large cell, small noncleaved, MALT, plasmacytoid
• leiomyosarcoma/leiomyoma (children > adults)
• ALL
• Kaposi’s sarcoma (children < adults)
87
*
Selected References
Koh AY, Pizzo PA. Fever and Granulocytopenia, and Infections in Children with Cancer. In: Long SS, Pickering LK, Prober
CG, editors. Principles and Practice of Pediatric Infectious Diseases. 2009.
Koh AY, Pizzo PA. Infectious Disease in Pediatric Cancer Survivors. In: Nathan and Oski 7th edition. 2009.
Pappas et al. Clinical practice guidelines for the management of candidiasis: 2009 update by the Infectious Diseases
Society of America. CLIN INFECT DIS (2009) vol. 48 (5) pp. 503-35
Patrick CC. Opportunistic infections in hematopoietic stem cell transplantation. In: Feigin, Cherry, Demmler-Harrison,
Kaplan, editors. Textbook of Pediatric Infectious Diseases, 6th edition. 2009.
Walsh TJ, Anaissie EJ, Denning DW, Herbrecht R, Kontonyiannis DP, Marr KA, Morrison VA, Segal BH, Steinbach WJ,
Stevens DA, Van Burik JV, Wingard JR, Patterson TF. Treatment of Aspergillosis: Clinical Practice Guidelines of
the Infectious Diseases Society of America. Clin Infect Dis. 2008. 46 (3): 327-360.
Bonilla FA, Geha RS. Primary Immunodeficiency Diseases. In: Nathan DG, Orkin SH, Ginsburg D, Look AT, editors.
Hematology of Infancy and Childhood. Philadelphia: WB Saunders; 2003. p. 1043-1100. (also new edition
2009)
Comans-Bitter WM, de Groot R, van den Beemd R, Neijens HJ, Hop WCJ, Groeneveld K, Hooijkaas H, van Dongen JJM.
Immunophenotyping of blood lymphocytes in childhood. 1997. Journal of Pediatrics 130 (3): 338-393.
Al-Herz W, Bousfiha A, Casanova J-L, Chatila T, Conley ME, Cunningham-Rundles C, et al. Primary immunodeficiency
diseases: an update on the classification from the international union of immunological societies expert
committee for primary immunodeficiency. Front Immunol. 2014;5:162. Cunningham-Rundles. How I treat
common variable immune deficiency. Blood (2010) vol. 116 (1) pp. 7-15
Cunningham-Rundles. Autoimmunity in primary immune deficiency: taking lessons from our patients. Clin Exp
Immunol (2011) vol. 164 Suppl 2 pp. 6-11
Oliveira et al. Revised diagnostic criteria and classification for the autoimmune lymphoproliferative syndrome (ALPS):
report from the 2009 NIH International Workshop. Blood (2010) vol. 116 (14) pp. e35-40
Key to slides and content specifications
II. Leukocytes
C. Lymphocytes
1. Normal morphology and age related values
3,21,26,27,28,35
2. Surface membrane antigens and gene rearrangements
3,21,24,26,27
3. Kinetics
24,25
4. Biochemistry
46
5a.1. B cells Immunoglobulins
3,11,22, 25,29-35
5b.1. T cells and T cell receptor
3,21,22,29,30
5b.2. NK cells
3,21,27
5b.3. Major Histocompatibility/Antigen presentation
3,11,22,78
6. Lymphocytosis
27,28
7. Lymphopenia
27,28,40-56
8. Mononucleosis
Did not include
Key to slides and content specifications
VI: Immunologic Abnormalities
A. Infections in Immunocompromised patients
1. Prophylaxis (Bacterial, Fungal, Viral, Pneumocystis
7, 9-19
2. Treatment of infection in immunocompromised patients (Bacterial, Fungal,
Viral, Pneumocystis)
7, 9-19
B. Immunodeficiency states
1. Congenital immunodeficiencies
a. Clinical features and inheritance
See each disorder
and tables
b. Relationship with cancer
22,23,63,6972,77,79
c.1. Wiskott-Aldrich syndrome
63-66, 74,79
c.2. Disorders of immunoglobulin production
58-61, 67-68, 7172,81-83
c.3. Severe Combined Immunodeficiency
50-56, 76-77,80
c.4. X-linked lymphoproliferative disease
69-70, 74,79
2. Acquired immunodeficiency syndrome (AIDS)
86-87
C. Autoimmune lymphoproliferative syndrome
73, other lectures
Immunology and Immunodeficiency for the
Pediatric Hematologist-Oncologist
Thank you for the invitation
Questions? E-mail:
[email protected]