Hodgkin’s Disease: Lineage and Clonality

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AMERICAN JOURNAL OF CLINICAL PATHOLOGY
Editorial
Hodgkin's Disease
Lineage and Clonality
WING C. CHAN AND JAN DELABIE
The cell lineage and clonality of the presumed neoplastic
cells of Hodgkin's disease (HD), the diagnostic Reed-
Sternberg cells and their variants (H/RS cells), have not
yet been clarified since the first description of HD by
Thomas Hodgkin more than 150 years ago.1
The demonstration of clonal cytogenetic abnormali-
ties in cell cultures of Hodgkin's tumor has been taken
as evidence for the neoplastic nature of HD. However,
satisfactory karyotyping is only obtainable in a minority
of cases and no specific cytogenetic abnormalities have
been detected after decades of observation.2
With the advent of immunohistochemistry and the
availability of a rapidly increasing number of mono-
clonal antibodies against a variety of leukocyte-associ-
ated antigens, it was anticipated that the question on the
cell lineage and clonality of
the
H/RS cells would be re-
solved. In the nodular type of lymphocytic predomi-
nance HD (N-LPHD), immunophenotyping has yielded
strong evidence that the H/RS cells belong to the B-cell
lineage, but controversy continues for the other subtypes
of
HD.
3
"
5
Although
CD
15,
CD74, CD30, MHC class II
and several other activation-associated molecules can be
demonstrated on the H/RS cells of
the
majority ofnon-
LPHD, more lineage-restricted markers are inconsis-
tently expressed.5-7 Even when such a marker
is
detected
on the H/RS cells of a case, it is frequently present on
only a subpopulation of the
cells.
In assigning
cell
lineage
based on immunohistochemical findings, it is important
to have a proper perspective of the specificity of the "cell
lineage marker" and the possibility of aberrant expres-
sion of the marker.8 In addition, the detection of a mole-
cule on or in a cell does not necessarily imply that the
molecule is synthesized by the cell as adsorption and/
or passive or active uptake from the surrounding may
account for the findings.9
The cloning of the immunoglobulin
(Ig)
and T-cell an-
tigen receptor (TCAR) genes has opened another avenue
to study the lineage and clonality of H/RS cells. South-
ern blot hybridization
has
been employed and in general,
clonal
Ig
gene rearrangement has been detected in some
cases of HD, but clonal TCAR gene rearrangement is
rarely detectable with the exception of Greisser's
study.10" The latter study may be an overestimation be-
cause the interpretation of clonal TCAR-gamma
gene
re-
arrangement by Southern blotting is difficult because of
the presence of pseudoclonal bands derived from normal
T-lymphocytes.12 The limitations of Southern blot anal-
ysis are its low sensitivity and the inability to determine
which cell population gives rise to a clonal band. To
overcome the sensitivity problem, a limited number of
cases
with large numbers of H/RS
cells,
as well
as
in
vitro-
enriched H/RS cell populations have been studied and
the incidence of clonal IgH rearrangement appeared to
be higher.1314 However, the signal strength is often not
commensurate with the proportion of H/RS cells present
in these
cases.
These observations suggest that the H/RS
cells gave rise to the rearranged bands, but the clonal re-
arrangements may not be present in the entire popula-
tion.
The development of the polymerase chain reaction
(PCR) offers a new and powerful approach to the study
of HD and it has been applied by several groups to study
the presence of clonal IgH gene rearrangement in Hodg-
kin's tumors. From these studies, it is quite clear that
cell populations with clonally rearranged IgH gene are
present in a proportion of LPHD as well as in other sub-
types of HD including cases where the H/RS cells
showed the classic immunophenotype, lacking CD20
staining."15"17 However, the percentages of cases where
clonality is demonstrated are quite variable in the differ-
ent reports.'' ' 5~'8 This discrepancy
is
probably the result
of differences in methodology and interpretation, as well
as bias in case selection. The correlation between the
presence of clonal IgH gene rearrangement and the im-
munophenotype of the H/RS cells is complicated be-
cause there is no clear consensus as to what constitute a
B-
or T-cell phenotype in non-LPHD. Aside from the
above considerations, there are a number of interesting
issues raised by these studies:
Why is the incidence of clonal IgH rearrangement in
some reports so much higher in PCR versus Southern
blotting analysis? The PCR assays used in these studies
may not be more sensitive in detecting a clonal IgH re-
arrangement because of the background smear caused
368
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CHAN AND DELABIE 369
Hodgkin
's
Disease:
by the coexisting polyclonal B-cell population. Several
possibilities to explain the increased rate of detection of
clonal populations by PCR were mentioned by Orazi
and colleagues," but the exact reason remains to be de-
termined. Because the size range of the VDJ amplifica-
tion products is rather narrow, apparent clonal bands
may be observed on occasions when the products are an-
alyzed in gels with low resolution. Such false-positive re-
sults may be avoided by performing the analysis in more
discriminating electrophoresis systems.
Is the clonal rearrangement derived from the H/RS
cells or from other cell populations in the tumor? Al-
though H/RS
cells
are the most likely source of the clonal
rearrangements detected, direct evidence is still lacking.
Several approaches to address this issue have been at-
tempted. This includes the study of Ig light chain restric-
tion in H/RS cells of LPHD by immunohistochemistry
and in situ hybridization. Unfortunately, the results are
controversial and the fact that light chain restriction al-
most always involves the kappa light chain remains to be
explained.1920 More convincing evidence that the H/RS
cells are the clonal population has come from the obser-
vation of Epstein-Barr virus (EBV) infection of the H/
RS cells and the simultaneous demonstration of mono-
clonality of the viral population by Southern hybridiza-
tion.21 The other approach to resolve the issue of lineage
and clonality is to study isolated H/RS cells by PCR. To
date,
only a few studies on a limited number of cases
have been performed and the findings have not been
consistent.22"24 One reason for the discrepancies is likely
because of the methodologic differences and/or case se-
lection bias in the various studies. For instance, neigh-
boring H/RS cells may be derived from a common pro-
genitor and identical IgH sequences would be obtained if
they were picked by micromanipulation out of a histo-
logic section,23 whereas the chance of picking two
closeby cells after tissue disaggregation is unlikely.2224
Resolution of the disparate findings will have to await
the careful study of
a
larger number of
cases
taking into
consideration the techniques used and the various char-
acteristics of the cases studied. However, one of the stud-
ies has already raised the intriguing possibility that
LPHD begins as a polyclonal B-cell proliferation24 and
that this may also
be
true in at least some
cases
of nodular
sclerosis type of HD expressing CD20.25 The emerging
evidence that some cases of HD contain polyclonal H/
RS cells, whereas others contain a clonal population
raises the possibility of clonal evolution in HD.
Is
it likely
that many cases of HD initiate as a polyclonal prolifera-
tion of H/RS cells that may develop into a monoclonal
lesion later in the course of
the
disease? Indeed, there is
some evidence that such clonal evolution may occur in
Vol.
I
eage and Clonality
LPHD,26 and it would be highly interesting to study se-
quential biopsies of other types of HD to examine this
possibility.
Finally, what does antigen-receptor gene rearrangement
tell us concerning the lineage of H/RS cells? While
B-
or T
antigen-receptor gene rearrangement is an important step
in the commitment of a cell toward the B or T lineage, it
may not
be
absolute,
especially in an abnormal
cell,
and we
can only regard it as a useful and additional "marker" of
differentiation. This finding has to be interpreted in the
context of other evidence of lineage commitment.27"30
With the currently available techniques, many of the
hypotheses concerning
the
lineage and clonality of HD can
be
vigorously
tested.
It
is
hoped that carefully planned stud-
ies in the future
will
provide new insights that would clarify
the pathobiology of this enigmatic disease.
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