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“Complex” receptor for
vitamin B12-intrinsic
factor
Patients with Imerslund-Gräsbeck syndrome
(I-GS, megaloblastic anemia 1 [MGA1], hereditary megaloblastic anemia, Online Mendelian
Inheritance in Man [OMIM] no. 2611001)
usually present with megaloblastic anemia
between 1 and 5 years of age. They have decreased levels of serum vitamin B12 (cobalamin) in the presence of normal levels of intrinsic factor (IF), and many patients have
proteinuria of the tubular type. The Schilling
test result is characteristic of the inability of
enterocytes to absorb the intrinsic factor–cobalamin complex. Patients in the original studies
were described as being Finnish or Norwegian.
Currently, more than 250 patients have been
identified, many of whom are of Middle
Eastern descent. I-GS has locus heterogeneity;
in most Finnish families, the disease is caused
by mutations in the cubilin (CUBN) gene on
chromosome 10p12.1.2 However, mutations in
CUBN were not found in Norwegian patients
showing the same phenotype. Through linkage
studies, a candidate gene was located on the
long arm of chromosome 14q32, and mutations
were found in the AMN gene in the Norwegian
patients.3 This gene emerged as a candidate
because of an expression pattern similar to that
of CUBN, with high levels of expression in
both small intestine and kidney.
Ever since the unexpected discovery of mutations in either the CUBN or the AMN gene in
patients with I-GS, it has become essential to
address the interaction of the 2 gene products:
BLOOD, 1 MARCH 2004 䡠 VOLUME 103, NUMBER 5
cubilin and amnionless. Fyfe and colleagues
(page 1573) show colocalization of cubilin and
amnionless proteins in the apical membranes
and endocytic apparatus of renal proximal
tubule cells. They also demonstrate physical
interaction between these 2 proteins following
sodium dodecyl sulfate–polyacrylamide gel
electrophoresis (SDS-PAGE) affinity purification. Cotransfection of Chinese hamster ovary
(CHO) cells with CUBN and AMN constructs
alters the exclusively intracellular locations seen
with transfection of CUBN alone, to include the
plasma membrane, and allows the endocytosis
of IF-cobalamin. These data reinforce findings
in the canine model of I-GS in which cubilin is
not expressed on the surface of intestinal and
renal cells and is retained in an early biosynthetic compartment. Canine I-GS maps to a
region orthologous to human chromosome 14
and presumably mutations in AMN will soon
be found in affected dogs.
A number of questions are still unanswered. Megalin, a member of the lowdensity lipoprotein (LDL) receptor family,
has been postulated to be involved in
cubilin function. Previous work has shown
the colocalization of cubilin with megalin,
and megalin-deficient mice have decreased
cubilin expression and uptake of cubilin
ligands. The case for amnionless has been
made much stronger than that for megalin
because of the finding of mutations in AMN
in I-GS and by the studies of Fyfe and colleagues. What, if anything, then is the physiologic role of megalin in cobalamin absorption? Also, both cubilin and amnionless are
implicated in early embryonic development
in rodents. Little is known about the mechanisms involved; the only known phenotype
resulting from mutations in the human CUBN
and AMN genes is I-GS. Finally, which
ligands other than cobalamin–intrinsic factor
interact with the new complex of cubilin
and amnionless for which Fyfe and colleague have coined the name “cubam”?
—David Rosenblatt
McGill University
1.
Online Mendelian Inheritance in Man, OMIM
[database online]. Baltimore, MD: Johns Hopkins
University; 2003. Available at: http://www.ncbi.
nlm.nih.gov/omim/. Accessed February 20, 2003.
2.
Aminoff M, Carter JE, Chadwick RB, et al. Mutations in CUBN, encoding the intrinsic factorvitamin B12 receptor, cubilin, cause hereditary
megaloblastic anaemia 1. Nat Genet. 1999;21:
309-313.
3.
Tanner SM, Aminoff M, Wright FA, et al. Amnionless, essential for mouse gastrulation, is mutated
in recessive hereditary megaloblastic anemia.
Nat Genet. 2003;33:426-429.
Regulatory T cells in
Hodgkin lymphoma
The presence of an extensive lymphoid infiltrate distinguishes Hodgkin lymphoma
(HL) from most other lymphomas. With
respect to this infiltrate, a number of questions can be asked. The first one is what
causes such an extensive infiltrate in the
presence of only a few tumor cells. The explanation may well be that Reed-Sternberg
cells produce and secrete high amounts of
chemokines, in particular thymus and activation-regulated chemokine (TARC) and
macrophage-derived chemokine (MDC), that
attract cells expressing the CCR4 receptor,
such as the T-helper 2 (Th2) lymphocyte.1
Another important question is why there is
no effective immune response against the
tumor cells.
Marshall and colleagues (page 1755)
confirm previous findings that the HLinfiltrating lymphocytes are anergic to stimulation with some mitogens and primary as
well as recall antigens but also demonstrate
that these cells suppress peripheral blood
mononuclear cell (PBMC) responses. They
identify the presence of interleukin-10
(IL-10)–secreting cells and CD4⫹CD25⫹
regulatory T cells. The immunosuppressive
effect of the HL-infiltrating cells could be
neutralized with anti–IL-10, by preventing cell-to-cell contact, and by anti–
cytotoxic T-lymphocyte–associated antigen 4
(anti–CTLA-4).
Marshall and colleagues also conclude
that the lymphocytes in HL do not produce
1565
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cytokines, such as IL-2, IL-4, and interferon-␥
(IFN-␥), with primary (keyhole limpet
hemocyanin [KLH]) and recall (purified
protein derivative [PPD]) antigens and the
mitogen concanavalin A (ConA). However,
in previous studies the lymphocytes were
found to produce these cytokines when
stimulated with phytohemagglutinin (PHA)
or with phorbolester (PMA)–ionomycin.
Specifically, when the CD26⫺ CD4 cells
immediately surrounding the Reed-Sternberg
cells were purified and stimulated with
PMA ionomycin, they produced IL-4 and
IFN-␥. The potential to produce IL-4 was
the reason why these cells were previously
considered Th2-like.2 The absence of IL-2
production upon stimulation is also associated with anergy. The exact nomenclature of
these cells is thus a matter of semantics. In
addition to the IL-10–producing cells (Tr1),
there are also transforming growth factor ␤
(TGF-␤)–producing cells present in the infiltrate, and these have been termed Th3.
The findings by Marshall and colleagues
indicate that there are variations in the populations involved in different cases. It can
be concluded that, as an overall population,
the infiltrating lymphocytes do not have Th1
type functions and are probably attracted
into the tissues by chemokines TARC and
MDC as CCR4-expressing Th2 cells. Although these cells do not spontaneously
produce IL-2 or IL-4, they produce IL-10,
despite not being fully activated, and therefore function as Tr1 cells.
The major remaining question is what
causes the predominance of T cells with
suppressor activity in Hodgkin lymphoma.
It appears that Reed-Sternberg cells, although they have the genotype of B cells,
execute a functional program that is similar
to antigen-presenting cells but results in tolerance. Mechanisms include the production
of immunosuppressive cytokines like IL-10,
especially in Epstein-Barr virus–positive
cases, and TGF-␤, especially in nodular
sclerosis cases, as well as the expression of
FAS ligand that induces cell death in FASexpressing activated T cells, while the ReedSternberg cells themselves are protected by
overexpression of cFLIP (Fas-associating
1566
protein with death domain–like IL-1␤–
converting enzyme [FLICE]–inhibitory
protein) or infrequently by FAS mutation.3
The relevance of these findings is that
they may allow a better design of new treatment modalities. There are indications that
the infiltrating cells in fact support the growth
and survival of the Reed-Sternberg cells,
and therefore blocking chemokines to
prevent the influx of T cells may be effective. On the other hand, blocking of the
immunosuppressive signals, such as IL-10
and TGF-␤ cytokines, or the removal of the
suppressor regulatory T cells may enhance
the effect of adoptive transfer of cytotoxic
T cells.4
—Sibrand Poppema
University Medical Center Groningen
1.
Van den Berg A, Visser L, Poppema S. High expression of CC chemokine TARC in Reed-Sternberg cells: a possible explanation for the characteristic lymphocytic infiltrate in Hodgkin’s disease.
Am J Pathol. 1999;154:1685-1691.
2.
Poppema S, van den Berg A. Interaction between
host T-cells and Reed-Sternberg cells in Hodgkin
Lymphomas. Semin Cancer Biol. 2000;10:345-350.
3.
Maggio E, van den Berg A, de Jong D, Diepstra
A, Poppema S. Low frequency of FAS mutations
in Reed-Sternberg cells of Hodgkin lymphoma.
Am J Pathol. 2003;162:29-35.
4.
Bollard CM, Rossig C, Calonge MJ, et al. Adapting a transforming growth factor beta-related tumor protection strategy to enhance antitumor immunity. Blood. 2002;99:3179-3187.
More complexity in
MLL-associated
leukemias
The mixed lineage leukemia gene (MLL,
also known as ALL-1, and HRX) has rightly
attracted much interest as a major player in
leukemia.1 MLL’s central role is clear by its
involvement with over 30 different partner
genes in recurrent translocations. As if this
were not enough, MLL is also implicated in
leukemia by overexpression in the absence
of overt mutations or by acquisition of
partial tandem MLL duplications. What,
then, accounts for the leukemogenicity of
MLL? Might there be some common functional thread tying together many of the
fusion genes and MLL overexpression? At
least one strong clue has emerged from the
recognition that a major function of MLL,
like its Drosophila homolog Trithorax, is to
serve as a maintenance factor for the expression of many members of the Hox
family of transcription factors. Hox genes
are now recognized as major components of
the regulatory machinery of primitive hematopoietic cells. Strikingly, multiple lines of
evidence link Hox genes directly to leukemic transformation.2,3 This evidence includes induction of leukemia in mice following engineered overexpression of certain
Hox genes (eg, HOXA9 and HOXA10) and
the observed overexpression of multiple
Hox genes in human leukemia and, notably,
in MLL-associated leukemias. Perhaps most
convincingly, multiple members of the Hox
family, HOXA9 for one, have been identified as translocation partners in leukemias
with the common partner Nucleoporin 98.4
Thus, a satisfying model for some if not all
MLL-induced leukemias would be through
induced deregulation of key Hox target
genes. Strong support for this has recently
been reported by Cleary and colleagues,
who found that MLL-ENL lost leukemogenicity in bone marrow cells taken from
Hoxa7 or Hoxa9 knockout mice.5 The jump
to a unifying model involving MLL and
HOXA9, however, is not without a tumble
or two as indicated in the article by Kumar
and colleagues (page 1823) in this issue of
Blood. Their studies reveal unabated leukemogenicity by the fusion gene MLL-AF9 in
the absence of Hoxa9. While there were
clear influences of Hoxa9 on the phenotype
of the leukemia, the essential transformation
was not altered by the presence or absence
of Hoxa9. The striking differences between
these 2 recent studies involving related
partner genes may be a consequence of
several experimental and biologic differences. Kumar and colleagues have used a
gene knock-in model of MLL-AF9 fusion
rather than retroviral overexpression; the
fusion genes may indeed have differential
effects on Hox targets, rendering other
members of the cluster more or less important. Indeed, multiple members of the Hox
A cluster were observed to be up-regulated
by MLL-AF9, making it possible that additive levels of Hox gene expression may be
BLOOD, 1 MARCH 2004 䡠 VOLUME 103, NUMBER 5
From www.bloodjournal.org by guest on June 16, 2017. For personal use only.
2004 103: 1565-1566
doi:10.1182/blood-2003-12-4285
Regulatory T cells in Hodgkin lymphoma
Sibrand Poppema
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