Selective IgG Deficiency Associated with Profound Neutropenia and Anemia

1
Selective IgG Deficiency Associated with Profound
Neutropenia and Anemia

• Soheil Chegini, MD

2
Case History

• 30 year-old male with a long-standing history of
  hypogammaglobulinemia and neutropenia
• Presenting Symptoms progressive fatigue and
  weakness, dyspnea on exertion, and malaise
• Failed empiric out-patient treatment for a
  suspected respiratory infection with oral
  amoxicillin-clavulanate for 17 d and
  clarithromycin for 8 d
• Dry cough at the onset of this episode that
  resolved
• No documented fevers or chills and no other
  symptoms

3
Past Medical History

• Unremarkable pre- and early postnatal course
• Healthy until age 18 month
• Recurrent OM starting in early childhood PET at
  age 3
• Chronic neutropenia with ANC 50-600 since age 3
• Ruled out for cyclic neutropenia
• Pancytopenia (Hb 4.5, WBC 2.7, ANC 135 plt 85K)
  and splenomegaly at age 7 following a viral
  syndrome
• Negative Coombs and autoantibodies
• Negative work-up for leukemia BM Bx that
  revealed myeloid arrest

4
Past Medical History

• Hospitalization for Pneumonia 4x between ages 3
  and 5 and at age 8 treated with Abx and received
  IM Ig
• Recovered from chicken pox uneventfully
• No history of thrush, sepsis, meningitis,
  osteomyelitis or skin infections
• Inconclusive work up at a university hospital
  prompted a referral to a pediatric
  immunodeficiency center at age 9
• Immunizations DTPx4, OPV x3 and MMRx1
  Pneumococcal polysaccharide vaccination
  (Pnemovax) complicated by severe Arthus reaction
  in 1980

5
Family History

• Healthy parents (Fa Caucasian / Mo Japanese)
• An older brother and sister both healthy
• A healthy 7 month-old daughter
• No known immune disorders
• No frequent or unusual infections
• No unexplained infant deaths

Allergy History

• No symptoms of aeroallergen sensitization with
  documented negative skin tests
• Sulfa antibiotics Vague history of an adverse
  reaction

6
Immune Workup at age 9

• Normal B and T cell count
• IgG 170 (531-1421) IgM 176 (27-112) IgA 660
  (85-335)
• Failed to respond to dT booster
• Impaired in vitro IgG secretion by his
  lymphocytes that normalized by co-incubation with
  normal T cells
• Bacteriophage FX174 vaccination
• Slightly decreased primary response, peak Kv 10
• No immunologic memory after 2 boosters
• Modest secondary response, entirely IgM with peak
  secondary and tertiary Kv of 2.2 and 0.6
• DTH to candida 13 mm induration
• In vitro lymphocyte proliferation was normal for
  mitogens and slightly low but in normal range for
  candida and tetanus

7
Interval Course

• Dx Selective IgG deficiency an unusual CVID
  variant
• Tx IVIG 0.4 g/Kg/mo
• H. influenzae Pneumonia in 1993 while on IVIG
• IVIG discontinued due to loss of insurance in
  1998
• No apparent deterioration without IVIG
• No documented pneumonia since
• Received IVIG infusion twice for symptomatic
  exacerbations similar to his current presentation
• Recurrent oral ulcers Repeatedly HSV negative

8
Physical Exam

• Afebrile, HR 89, RR 18, BP128/83, O2 sat 99 RA
• Pale, well developed, athletic, young man in NAD
• HEENT No rhinorrhea, pharyngeal erythema or
  thrush
• Resp Non-labored, CTA with good AE bilaterally
• CV RRR no m/g/r
• Abd Soft, NT, spleen tip palpable, Liver 10 Cm
• Lymph No palpable LAD
• Extr No C/C/E
• Neuro Intact
• Skin No rash

9
Laboratory Data

• Hemoglobin 5.6 normocytic/normochromic, normal
  RDW
• LDH 1524 (313-618), haptoglobin was lt20
• No serologic evidence of autoimmune anemia
• WBC 2.2 ANC 200, ALC 1600
• Platelets 48K
• ESR 23
• Normal electrolytes, renal function bili and
  liver enzymes
• Albumin 3.4 (3.5-5.0), total protein 5.6
  (6.3-8.2)
• IgG lt 35, IgE lt 2, IgM 144 and IgA 195

10
Imaging Results

• Chest CT Mild bronchiectasis in the medial basal
  segment of the left lower lobe
• Abdominal CT Hepatosplenomegaly with the spleen
  15.5 cm and the liver 18 cm
• Sinus CT No evidence of sinusitis. Thickening
  of the right tympanic membrane with destructive
  changes in the right mastoid air cells

11
Immune Workup

• Lymphocyte subtypes Normal CD3, CD4, CD8, CD19
  and CD16/CD56 cell counts
• Tetanus antibody titer lt 0.1
• Pneumococcal titers lt 0.5 for all 14 serotypes

Hematology Workup

• Bone marrow biopsy Marked hypocellularity
• Small clusters of erythroid precursors, rare
  megakaryocytes
• Scattered lymphocytes, many of which were large,
  atypical and blast-like in appearance
• Highly suggestive of peripheral T-cell lymphoma

12
Hospital Course

• PRBC transfusion 2 units
• Hemoglobin increased to 7.5
• No clear diagnosis
• In the past single IVIG infusions had promptly
  resulted in improvement
• IVIG 400mg/Kg after blood samples were obtained

13
Confirmatory Diagnostic Tests

• Parvovirus DNA detected by PCR in blood sample
• PCR was negative for EBV, HHV6 and HHV8
• CD40L expression
• 0.8 on unstimulated CD4 cells (normal14-52)
• No upregulation after stimulation with PMA
• CD40L (CD154) gene mutation analysis
• Missense mutation at position 506 AgtG
• Conversion of tyrosine 169 to cysteine (Y169C)
• Previously reported mutations at this position
  Y169D and Y169N cause X-HIGM, but Y169C has not
  been reported

Thusberg J, Vihinen M. The structural basis of
hyper IgM deficiency - CD40L mutations. Protein
Eng Des Sel. 2007 20133-41
14
Clinical Course at 6-Month Follow Up

• No infection without any further doses of IVIG or
  correction of neutropenia
• Professional tennis player winning tournaments
• WBC 2.9 with ANC 464 and ALC 1,798
• Hb 15.2 with normal MCV/RDW and Plt 225
• DTH Tetanus 13 mm, Candida 7 mm
• Normal hip and lumbar bone mineral density

Lopez-Granados E, Temmerman ST, Wu L et al.
Osteopenia in X-linked hyper-IgM syndrome reveals
a regulatory role for CD40 ligand in
osteoclastogenesis. Proc Natl Acad Sci U S A.
20071045056-61
15
Just Another Typical Patient with X-HIgM?
16
Queries Relevant to Clinical Management

• IgG deficiency Functionality of IgA and IgM and
  need for IgG replacement
• T cell immunity Susceptibility to opportunistic
  infections and necessity of preventive measures
• Neutropenia Need for G-CSF
• Chronic parvovirus infection Need for
  eradication and ability to achieve and sustain it

17
Review of Literature1. Neutropenia2.
Parvovirus-Induced Anemia3. IgA in X-HIgM
patients
18
Neutropenia in X-HIgM

• About 2/3 of X-HIGM patients have neutropenia
• Neutropenia was present in 38 (68) of 56
  European X-HIgM patients
• Chronic in 25 (45)
• Oral ulcers were present also in 25 patients
  (45) and in 88 of cases were associated with
  neutropenia
• In a US cases  50 of 79 (63) had neutropenia
• Chronic in 27 (34)
• Oral ulcers in 24 of the neutropenic patients
  (48)

Levy J, Espanol-Boren T, Thomas C et al. Clinical
spectrum of X-linked hyper-IgM syndrome. J
Pediatr. 199713147-54 Winkelstein JA, Marino
MC, Ochs H et al. The X-linked hyper-IgM
syndrome clinical and immunologic features of 79
patients. Medicine (Baltimore). 200382373-84
19
Anemia Due to Parvovirus in X-HIgM

• Hasle and colleagues reported the first
  recognized case of chronic anemia due to
  parvovirus infection in an 8 y.o. boy who was
  subsequently diagnosed with X-HIgM (R11X)
• A case series reported this and 2 additional
  patients (ages 14 and 17 yrs) who presented with
  severe anemia as the initial manifestation of
  X-HIgM
• No prior history of increased susceptibility to
  infections
• IgA level was normal (71 mg/dL) in one of these
  patients

Hasle H, Kerndrup G, Jacobsen BB et al. Chronic
parvovirus infection mimicking myelodysplastic
syndrome in a child with subclinical
immunodeficiency. Am J Pediatr Hematol Oncol.
199416329-33 Seyama K, Kobayashi R, Hasle H et
al. Parvovirus B19-induced anemia as the
presenting manifestation of X-linked hyper-IgM
syndrome. J Infect Dis. 1998178318-24
20
Anemia Due to Parvovirus in X-HIgM

• A more recent case report of an X-HIgM with
  initial presentation of parvovirus-induced anemia
• CD40L gene mutation resulted in cytoplasmic
  truncation of CD40L (R11X)

• All immunoglobulin levels were initially normal,
  but IgM and IgG decreased to slightly below normal

Blaeser F, Kelly M, Siegrist K, Storch GA et al.
Critical function of the CD40 pathway in
parvovirus B19 infection revealed by a
hypomorphic CD40 ligand mutation. Clin Immunol.
2005117(3)231-7
21
X-HIgM with Normal or Elevated IgA

• In a European cohort of 55 X-HIGM patients, who
  had a documented IgA, it was very low or
  undetectable in 51, but elevated in 3 cases up to
  292 mg/dL
• In a US case series 63 patients, IgA was normal
  for age in 10 and elevated in 4 patients up to
  165 mg/dL
• In an Argentinean series of 11 patients with
  milder variants IgA level was normal in 3 cases
  up to 204 mg/dL

Levy J, Espanol-Boren T, Thomas C et al. Clinical
spectrum of X-linked hyper-IgM syndrome. J
Pediatr. 199713147-54 Winkelstein JA, Marino
MC, Ochs H et al. The X-linked hyper-IgM
syndrome clinical and immunologic features of 79
patients. Medicine (Baltimore).
200382373-84 Danielian S, Oleastro M, Eva Rivas
M et al. Clinical Follow-Up of 11 Argentinian
CD40L-Deficient Patients with 7 Unique Mutations
Including the So-Called "Milder" Mutants. J Clin
Immunol. 200727455-9
22
Case Report with Similar Features

• AA male initially presented at age 2 with
• Recurrent upper and lower respiratory infections
• Low serum IgG and IgA and elevated IgM

• Chronic neutropenia
• Serum IgA normalized in early childhood but was
  markedly elevated later on
• IgA was polyclonal 93 IgA1 and 7 IgA2

• CD40L gene mutation 554 AgtG in exon 5
• Y170C in the extracellular domain

Atkinson TP, Smith CA, Hsu YM et al. Leukocyte
transfusion-associated granulocyte responses in a
patient with X-linked hyper-IgM syndrome. J Clin
Immunol. 199818430-9
23
Potential Mechanisms1. Neutropenia2. Leaky IgA
Class Switch3. T Cell Immunity
24
Role of CD40L in Hematopoiesis

• Ligation of CD40 on bone marrow stromal cells
• Induces both membrane-bound and secreted Flt-3
  ligand (FL)
• Effects of FL on CD34 hematopoietic stem cells
• Increased proliferation and generation of
  clonogenic cell
• Enhancement of granulocytopoiesis and
  megakaryocytopoiesis
• Differentiation into functional dendritic cells
• FL serum levels inversely correlate with ANC in
  patients with chronic idiopatic neutropenia and
  after chemotherapy or BMT
• In X-HIgM patients FL levels have not been
  investigated

Solanilla A, Dechanet J, El Andaloussi A et al.
CD40-ligand stimulates myelopoiesis by regulating
flt3-ligand and thrombopoietin production in bone
marrow stromal cells. Blood. 2000953758-64 Haida
r JH, Bazarbachi A, Mahfouz R et al. Serum Flt3
ligand variation as a predictive indicator of
hematopoietic stem cell mobilization. J
Hematother Stem Cell Res. 200211533-8 Papadaki
HA, Damianaki A, Pontikoglou C et al. Increased
levels of soluble flt-3 ligand in serum and
long-term bone marrow culture supernatants in
patients with chronic idiopathic neutropenia. Br
J Haematol. 2006132637-9
25
T Cell Independent CSR Signaling

• Do involve not CD40L-CD40 interaction
• Dendritic cells activated by IFN-a, IFN-g, or LPS
  express BAFF (B-cell activating factor) and APRIL
  (a profileration-inducing ligand) that can
  activate B cells
• BAFF and APRIL are also members of TNF family
• Other signals involved in CSR
• Cytokines (TGF-b)
• Engagement of B-cell receptor by cognate antigen
• TLR-9 engagement by CPG

Litinskiy MB, Nardelli B, Hilbert DM et al. DCs
induce CD40-independent immunoglobulin class
switching through BLyS and APRIL. Nat Immunol.
20023822-9 Castigli E, Wilson SA, Scott S et
al. TACI and BAFF-R mediate isotype switching in
B cells. J Exp Med. 200520135-9 He B, Qiao X,
Cerutti A. CpG DNA induces IgG class switch DNA
recombination by activating human B cells through
an innate pathway that requires TLR9 and
cooperates with IL-10. J Immunol. 20041734479-91
26
T Cell Independent CSR Signaling

• TACI is a member of the TNF receptor family that
  serves as a receptor for BAFF and APRIL
• IgA production can be induced by coligation of
  TACI and heparan sulfate proteoglycans bypassing
  APRIL
• CpG (TLR-9) and LPS (TLR-4) strongly up-regulate
  TACI expression
• IgA and IgG secretion stimulated by BAFF or APRIL
  and CpG can be blocked by TACI neutralizing
  antibody and is absent in TACI-/- mouse

Sakurai D, Hase H, Kanno Y et al. TACI regulates
IgA production by APRIL in collaboration with
HSPG. Blood. 20071092961-7 Katsenelson N,
Kanswal S, Puig M et al. Synthetic CpG
oligodeoxynucleotides augment BAFF- and
APRIL-mediated immunoglobulin secretion.Eur J
Immunol. 2007371785-95
27
CD40-Independent CSR Activation

• In vitro stimulated T cells of some X-HIgM
  patients can induce class switch to IgE
• IgE was detectable in 7 of 27 Eropean X-HIgM
  patients and was markedly elevated (gt1000 KU/L)
  in one
• IgE class switch can be induced in highly
  purified tonsilar B cells without involvement of
  CD40
• This is achieved by ligation of CD58 (LFA-3) on
  B-cells by activating antibodies or CD2 on T-cells

Life P, Gauchat JF, Schnuriger V et al. T cell
clones from an X-linked hyper-immunoglobulin
(IgM) patient induce IgE synthesis in vitro
despite expression of nonfunctional CD40 ligand.
J Exp Med. 19941801775-84. Diaz-Sanchez D,
Chegini S, Zhang K, Saxon A. CD58 (LFA-3)
stimulation provides a signal for human isotype
switching and IgE production distinct from CD40.
J Immunol. 199415310-20.
28
CHa Specific CSR

• Homeodomain proteins HoxC4 and Oct-1 in a complex
  with the Ku70/Ku86 heterodimer bind to ATTT
  switch regulatory elements in the Ig and Ie
  promoters
• This dampens basal germ line Ig-Cg and Ie-Ce
  transcriptions and represses CSR to Cg and Ce
• The complex does not bind to the Ia1/Ia2
  promoters
• CD40 signaling dissociates the complex and
  relieves the IH-CH transcriptional repression to
  allow CSR
• CD153 engagement inhibits this dissociation

Schaffer A, Kim EC, Wu X et al. Selective
inhibition of class switching to IgG and IgE by
recruitment of the HoxC4 and Oct-1 homeodomain
proteins and Ku70/Ku86 to newly identified ATTT
cis-elements. J Biol Chem. 200327823141-50
29
Negative Regulatory Pathways

• CD153 (CD30 ligand) is a member of the TNF family
  expressed on the surface of human B cells
• T cells activated by antigen and IL-4 express
  CD30
• IgD IgM B cells up-regulate CD153 on exposure
  to CD154, IL-4, and B cell Ag receptor engagement
• CD153 engagement inhibits IgG, IgA, and IgE
  production
• CD30 T cells inhibit CSR by inducing reverse
  signalling through B-cell CD30L

Cerutti A, Schaffer A, Goodwin RG et al.
Engagement of CD153 (CD30 ligand) by CD30 T
cells inhibits class switch DNA recombination and
antibody production in human IgD IgM B cells. J
Immunol. 2000165786-94
30
Role of CD40L in T Cell Immunity

• CD154 reverse signal to T cells is not essential
  for T cell function
• Normal T cell function can be restored by
  providing B7 costimulation via CD28

Lobo FM, Scholl PR, Fuleihan RL. CD40
ligand-deficient T cells from X-linked hyper-IgM
syndrome carriers have intrinsic priming
capability. J Immunol. 20021681473-8 Grewal IS,
Foellmer HG, Grewal KD et al. Requirement for
CD40 ligand in costimulation induction, T cell
activation, and experimental allergic
encephalomyelitis. Science. 19962731864-7
31
Alternative Roles of CD40L in T Cells

• Besides CD40, CD154 can interact with integrins
• a5b1 (fibronectin receptor)
• aMb2 (CD11b/CD18, Mac-1)
• aIIbb3 the (CD41/CD61, platelet GPIIb/IIIa)
• Binding of CD154 to integrins can stabilize
  adhesion and strengthen co-stimulation between T
  cells and antigen presenting cells

Leveille C, Bouillon M, Guo W et al. CD40 ligand
binds to alpha5beta1 integrin and triggers cell
signaling. J Biol Chem. 20072825143-51  Zirlik
A, Maier C, Gerdes N et al. CD40 ligand mediates
inflammation independently of CD40 by interaction
with Mac-1. Circulation. 20071151571-80