Page 168 - Vitamin D and Cancer

7 Induction of Differentiation in Cancer Cells by Vitamin D 155

7.3 Leukemias

Hematological malignances are a diverse group of diseases, but can be divided into
two major groups, the lymphocytic and myeloid leukemias. Although normal acti-
vated B and T lymphocytes express VDR, and 1,25D has antiproliferative effects
on these cell types (e.g., [122, 123]), this does not appear to alter their differentia-
tion state, and lymphocytic leukemia cells do not respond to 1,25D. In contrast,
1,25D has been known since 1981 to induce maturation of mouse myeloid leukemia
cells [124], and this can also take place in a wide variety of human myeloid leukemia
cell lines, with the exception of the lines derived from the most immature acute
myeloid leukemia (AML) blast cells (e.g., [125–127]).
Differentiation induced by 1,25D usually results in a monocyte-like phenotype,
but prolonged exposure to 1,25D confers cell surface changes that result in adher-
ence to the substratum, making the differentiated cells macrophage-like [124, 128].
The monocyte characteristics are recognized by changes related to phagocytosis,
such as the ability to break down esters, assayed by the “non-specific esterase”
(NSE) cytochemical reaction, also known as “monocyte-specific esterase” (MSE)
since in the hematopoietic cells this esterase is specific for monocytes and mac-
rophages [129]. Also related to phagocytosis is the ability to generate reactive oxygen
species (ROS) including superoxide, usually recognized by the nitroblue tetrazolium
(NBT) or cytochrome reduction [130, 131]. The availability of Flow Cytometry (FC)
for the recognition of surface proteins has made the study of the differentiating
effects of 1,25D on myeloid leukemia cells quite simple, using CD14, a receptor for
complexes of lipopolysaccharides (LPS) and LPS-binding protein [132], a near-
definitive marker of the monocytic phenotype. This is usually supplemented by the
FC determination of CD11b, or another subunit of the human neutrophil surface
protein that mediates cellular adherence [133]. In contrast to myeloid cells induced
to differentiate by the phorbol ester TPA, in 1,25D-treated cells the ability to adhere
develops more slowly than the ability to phagocytose. Consequently, 1,25D treat-
ment results in an earlier appearance of the CD14 antigen, usually accompanied in
parallel by MSE positivity, than the appearance of CD11b and NBT positivity
[134, 135]. Generally, at least two of the above parameters are measured to demon-
strate monocytic differentiation, and FC methods require the use of paired isotypic
IgG controls for each test sample to avoid obtaining false-positive data. Exposure of
AML cells to 1,25D also results in G1 phase cell cycle arrest, which follows, rather
than precedes, the phenotypic differentiation [134], and is often taken as the confir-
matory evidence that differentiation has taken place. However, in contrast to cells
from most solid tumors, monocytic differentiation of AML cells is accompanied by
increased expression of anti-apoptotic proteins, and consequently 1,25D-treated
myeloid cells have an increased cell survival potential [136–140].
The topic of 1,25D-induced leukemia cell differentiation has been extensively
studied in many laboratories. These include several groups in Japan [141–145], and
a group in Birmingham, England [146, 147], who made many valuable contribu-
tions to the field. Notably, combined basic and clinical studies of 1,25D-induced