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Blood, Vol. 94 No. 4 (August 15), 1999:
pp. 1488-1490
CORRESPONDENCE
The 782C  T (T254M) XHIM Mutation: Lack of a Tight
Phenotype-Genotype Relationship
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LETTER |
To the Editor:
In a recent report in Blood, Seyama et al1
described 28 unique mutations of the CD40 ligand
(CD40L) gene in 45 X-linked hyper IgM syndrome (XHIM)
patients from 30 unrelated families. Generally, peripheral blood
mononuclear cells (PBMC) are screened for CD40L expression after
activation with phorbol myristate acetate (PMA) and
ionomycin to diagnose XHIM. Activated PBMC of most XHIM patients did
not express functional CD40L (CD154): they failed to bind a CD40-Ig
fusion protein (bCD40-Ig). The investigators discerned 5 different
CD40L staining patterns on cultured T cells, using a polyclonal
antiserum (pAb), 4 different monoclonal antibodies (McAb), and
bCD40-Ig. The type 1 pattern showed weak staining of CD40L with all
reagents mentioned; the type 2 through 5 patterns successively showed
loss of functional activity (bCD40-Ig binding) and protein epitope
expression (loss of 1 to 4 McAb and finally pAb binding). A
relationship between genotype and phenotype was suggested, with
"milder genotypes" (resulting in staining pattern type 1 or 2)
showing milder clinical phenotypes: in 5 of 10 patients from 9 families
with staining pattern type 1 or 2, symptoms started relatively late,
and none of them suffered from opportunistic infections. Two patients
with the 782C  T mutation, a missense mutation in exon 5, fit into
this group.
We found the 782C T XHIM mutation in 4 patients from 2 different
families. Although they showed a favorable clinical course after Ig
replacement therapy was started, the 2 index patients had symptoms from
an early age onwards, including 1 who presented with Pneumocystis
carinii pneumonia (PCP). The other 2 children were diagnosed
shortly after birth because of the XHIM index patient in the family.
The first index patient, patient no. 1 (of family A), is a young Dutch
man, now 19 years of age, who was treated with Ig replacement therapy
from the age of 2 years onward because of hypogammaglobulinemia and
frequent respiratory tract infections. Since then, the frequency of
infections has normalized. He never had an infection with an opportunistic pathogen and never received prophylaxis with
Co-trimoxazole. Recently, stimulation of his PBMC with PMA and
ionomycin induced expression of the early activation marker CD69
(Leu-23; Becton Dickinson, San Jose, CA) on his CD3+ T
lymphocytes, whereas CD40L (LL48; Schering-Plough, Dardilly, France)
expression was absent (Fig 1). Fluorescent
sequencing of the CD40L gene showed a 782C T mutation
(expected protein alteration T254M). The family was screened, and in
patient no. 2, his cousin now 2 years of age, XHIM was identified
shortly after birth (Fig 2). At 9 months of
age, the serum IgG level had decreased to less than 0.5 g/L. Although
there were no clinical symptoms, Ig replacement therapy was started
because of the increased risk of serious infections.
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| Fig 1.
CD69 and CD40L expression on CD3+ T
lymphocytes after stimulation of PBMC with PMA and ionomycin. (A)
Normal control. (B) Patient no. 1.
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| Fig 2.
Single-strand conformation polymorphism (SSCP) analysis
of XHIM family A with T254M mutation. PCR products (198 bp) of exon 5 of the CD40L gene were denatured and run on a 12.5%
polyacrylamide gel at 150 V for 14 hours. The 2 control (C) lanes show
the positions of the DNA strands of the unaffected X-chromosome. The C
T mutation leads to a shift in the position of these bands, as is
shown in lane 1 (XHIM patient no. 2). Carriers (lanes 2 and 3) show
both mutated and unmutated DNA strands. Noncarriers (lane 4) only show
the unmutated DNA strands.
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The second index patient, patient no. 3 (of family B), is a British
boy, now 11 years of age, who was diagnosed as having XHIM at 6 months
of age when he presented with PCP. After successful treatment of this
infection, he has remained asymptomatic on Ig replacement therapy and
Co-trimoxazole prophylaxis except for 1 episode of a diarrhoeal illness
at 7 years of age that was of unknown etiology, lasted for 2 months,
and was associated with weight loss. Recent studies show no evidence of
cholangiopathy on liver function tests, liver histology, and endoscopic
retrograde cholangio-pancreatogram. His younger brother (patient no.
4), now 8 years of age, was diagnosed soon after birth based on the family history. He has remained completely well on Ig replacement therapy and Co-trimoxazole prophylaxis. His liver is also normal. The
mutation in these boys has been previously reported (case 13 in Katz et
al2).
In our opinion, the issue of a phenotype-genotype relation in XHIM
remains debatable. Not all patients with a staining pattern type 1 or 2 described by Seyama et al1 showed a mild clinical phenotype. Comparison of our 2 index patients with the 782C T
mutation with the 2 mild patients described by Seyama et
al1 does not support a tight phenotype-genotype relation in
XHIM patients, although our 4 patients all showed a favorable course
after Ig replacement therapy. We agree that symptomatic PCP may occur
in nonimmunocompromised infants, although this is very unusual.
Nevertheless, we do not believe that the life-threatening episode of
PCP in our patient no. 3 is compatible with a label of mild phenotype.
Furthermore, our XHIM cases also show that a favorable clinical course
on Ig replacement therapy is compatible with a diagnosis of XHIM. This
implies that testing of CD40L expression on activated PBMC
should be considered in all patients with unexplained
hypogammaglobulinemia.
Esther de Vries
Jeroen G. Noordzij
E. Graham Davies
Nico Hartwig
Mart n H. Breuning
Jacques J.M. van Dongen
Maarten J.D. van Tol
Department of Pediatrics
Centre for Clinical Genetics
Leiden
University Medical Centre
Leiden, The Netherlands
Department of
Immunology
Erasmus University Rotterdam
Rotterdam, The
Netherlands
Department of Pediatrics
Sophia Children's
Hospital
Rotterdam, The Netherlands
Department of Immunology
Great Ormond Street Hospital
London, UK
| |
REFERENCES |
1.
Seyama K, Nonoyama S, Gangsaas I, Hollenbaugh D, Pabst HF, Aruffo A, Ochs HD:
Mutations of the CD40 ligand gene and its effect on CD40 ligand expression in patients with X-linked hyper IgM syndrome.
Blood
92:2421, 1998[Abstract/Free Full Text]
2.
Katz F, Hinshelwood S, Rutland P, Jones A, Kinnon C, Morgan G:
Mutation analysis in CD40 ligand deficiency leading to X-linked hypogammaglobulinemia with hyper IgM syndrome.
Hum Mutat
8:223, 1996[Medline]
[Order article via Infotrieve]
Response
In a recent report,1 we have described 27 unique mutations
of the CD40 ligand (CD40L) gene in 30 unrelated
families with X-linked hyper IgM syndrome (XHIM). Assessment of CD40L
expression by activated lymphocytes identified a subgroup of XHIM
patients whose lymphocytes could be stained with all monoclonal
antibodies to CD40L tested and with a CD40-Ig construct (type I) or
only with the monoclonal antibodies but not with CD40Ig (type II). We
further reported that 5 of the 9 patients with type I or type II
staining had a mild disease with late onset, few infections, and no
history of Pneumocystis carinii pneumonia (PCP). Two of these
XHIM patients with mild disease had a T254M mutation. In their letter,
de Vries et al report 2 pairs of unrelated brothers with the same T254M
mutation. They point out that the older boy of family A, now doing well
at 19 years of age, presented with PCP. The others, now 2, 8, and 11 years of age, are on IVIG and have also shown a consistently benign
course. The observation by de Vries et al confirms our findings that,
although mild, some of the patients with type I or type II staining (2 of 9 in our group) may develop PCP. As we reported in a subsequent
communication,2 3 of the 5 mild cases initially presented
with parvovirus B19-induced anemia at 8, 14, and 17 years of age,
respectively. All were asymptomatic before parvovirus B19 infection and
again after IVIG therapy was initiated. Thus, patients with type I and
type II staining may develop opportunistic infections such as PCP and
persistent parvovirus B19 infection, suggesting that any mutation
affecting the function of CD40L3 renders T cells less
efficient. In addition, abnormal T-helper function was demonstrated in
every patient with type I or II staining immunized with the
T-cell-dependent antigen, bacteriophage  X174. However, antibody
titers to phage were higher in patients with type I/II staining than in
classic XHIM.1
Long-term follow-up of XHIM patients with mutations affecting CD40L
suggests that classic XHIM is not a benign disease.1,4,5 Many patients report chronic diarrhea, progressive liver disease often
associated with cryptosporidium infection, ascending cholangiolitis, chronic neutropenia, autoimmunity, lymphohyperplasia, and increased incidents of neoplasms, including lymphomas and biliary tract tumors. A
recent European study found that 23% of their study population with
XHIM had died (mean age at death, 11.7 years).4
Based on the grim diagnosis of classical XHIM, bone marrow
transplantation has been advocated.6 Long-term follow-up of patients with mild and classic phenotype is required to determine if
those patients with type I and type II staining have a better long-term
prognosis. Two registries have begun collecting data, one in Europe and
one in the United States, with emphasis on mutation analysis and
clinical phenotype. These long-term observations will provide the
details needed for optimal clinical management of mild and classic XHIM
phenotypes.
Hans D. Ochs
Division of Infectious
Diseases, Immunology and Rheumatology
Department of Pediatrics
University of Washington School of Pediatrics
Seattle, WA
Kuniaki Seyama
Department of Respiratory Medicine
Juntendo
University, School of Medicine
Tokyo, Japan
| |
REFERENCES |
1.
Seyama K, Nonoyama S, Gangsaas I, Hollenbaugh D, Pabst HF, Aruffo A, Ochs HD:
Mutations of the CD40 ligand gene and its effect on CD40 ligand expression in patients with X-linked hyper IgM syndrome.
Blood
92:2421, 1998
2.
Seyama K, Kobayashi R, Hasle H, Apter AJ, Rutledge JC, Rosen D, Ochs HD:
Parvovirus B19 induced anemia as presenting manifestation of X-linked Hyper IgM syndrome.
J Infect Dis
178:318, 1998[Medline]
[Order article via Infotrieve]
3.
Bajorath J, Seyama K, Nonoyama S, Ochs HD, Aruffo A:
Classification of mutations in the human CD40 ligand, gp39, that are associated with X-linked hyper IgM syndrome.
Prot Sci
5:531, 1996[Medline]
[Order article via Infotrieve]
4. Levy J, Espanol-Boren T, Thomas C, Fischer A, Tovo P, Bordigoni
P, Resnick I, Fasth A, Baer M, Gomez L, Sanders AM, Tabone M-D, Plantaz
D, Etzioni A, Monafo V, Abinun M, Hammarstrom L, Abrahamsen T, Jones A,
Finn A, Klemola T, DeVries E, Sanal O, Peitsch MC, Notarangelo LD: The
clinical spectrum of X-linked hyper-IgM syndrome. J Pediatr (in
press)
5.
Hayward AR, Levy J, Facchetti F, Notarangelo L, Ochs HD, Etzioni A, Bonnefoy JY, Cosyms S, Weinberg A:
Cholangiopathy and tumors of the pancreas, liver, and biliary tree in boys with X-linked immunodeficiency with hyper-IgM.
J Immunol
158:977, 1997[Abstract]
6.
Thomas C, de Saint Basile G, Le Deist F, Theophile D, Benkerrou M, Haddad E, Blanche S, Fischer A:
Brief report: Correction of X-linked hyper-IgM syndrome by allogeneic bone marrow transplantation.
N Engl J Med
333:426, 1995[Free Full Text]
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