Rituximab (Rituxan) therapy is successfully
used to treat many B-cell malignancies. Absent or diminished CD20
expression on certain B-cell tumors may limit the efficacy of
We attempted to identify agents that could induce CD20 expression on
these B-cell tumors. We first screened RPMI 8226 multiple myeloma
cells with agents known to modulate lymphoid-cell surface receptors,
and identified interferon-gamma (IFN-gamma [Actimmnue]) as an inducer
of CD20 expression.
We subsequently cultured multiple myeloma patient plasma cells (N =
20), multiple myeloma patient B cells (N = 9), chronic lymphocytic
leukemia (CLL) patient B cells (N = 5), non-Hodgkins lymphoma
(NHL) B cells (N = 3), normal donor B cells
(N = 11), normal donor plasma cells (N = 3), and normal donor
CD34-positive hematopoietic progenitor cells (N = 5) without and with
IFN-gamma (1 to 100 U/mL) for 48 hours, and evaluated changes in CD20
expression with multicolor flow cytometry. Rituximab binding to RPMI
8226 multiple myeloma cells and multiple myeloma patient plasma cells
was also determined before and after IFN-gamma treatment.
These studies demonstrated that IFN-gamma induced CD20 expression
(increased mean intensity, and percentage of cells expressing CD20)
on multiple myeloma plasma cells, multiple myeloma B cells, and
normal donor plasma cells. In contrast, CD20 expression on CLL, NHL
(CD20low), and normal donor B cells, as well as on normal donor
progenitor cells, was unaffected by IFN-gamma.
The levels of IFN-gamma receptor expression alone did not explain the
above differences, since a comparable percentage of multiple myeloma
(82% ± 6%) and CLL B cells (88% ± 9%) expressed the
IFN-gamma receptor. However, IFN-gamma receptor expression was lower
on normal donor (26% ± 6%) vs multiple myeloma and CLL B cells
(P < .001). Rituximab binding to RPMI 8226 cells and to multiple
myeloma patient plasma cells increased following culture with
pharmacologically attainable levels of IFN-gamma (1 to 100 U/mL).
Changes in Pu.1 expression, a transactivator of CD20 expression that
is downregulated with plasma cell differentiation, were also examined
to discern the means by which IFN-gamma induces CD20 on plasma cells.
Western blot analysis and im-munofluorescence staining of RPMI 8226
plasma cells treated with IFN-gamma for 0 to 48 hours showed that
Pu.1 was induced at 6 hours, and coincided with CD20 upregulation.
CONCLUSION: IFN-gamma induces CD20 on multiple myeloma plasma cells
and multiple myeloma B cells and augments rituximab binding to
multiple myeloma plasma cells. These findings provide the rationale
for use of rituximab with CD20-directed serotherapies for multiple myeloma.