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Experimental Therapeutics, Preclinical Pharmacology |
,25-Dihydroxyvitamin D2 as Therapeutic Agents for Prostate Cancer1
Vitamin D, Skin and Bone Research Laboratory and Endocrine Section, Boston University Medical Center, Boston, Massachusetts 02118 [T. C. C., M. F. H.]; Department of Cancer Biology, Comprehensive Cancer Center of Wake Forest University, Winston-Salem, North Carolina 27157 [G. G. S.]; Department of Molecular and Cellular Pharmacology[K. L. B.]; and Department of Urology, University of Miami School of Medicine, Miami, Florida 33101[B. L. L.]
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ABSTRACT |
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 Top ABSTRACT INTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONREFERENCES |
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,25-dihydroxyvitamin D3
(1,25(OH)2D3), which is known to inhibit the
proliferation and invasiveness of these cells. These findings support
the use of 1,25(OH)2D3 for prostate cancer
therapy. However, because 1,25(OH)2D3 can
cause hypercalcemia, analogues of
1,25(OH)2D3 that are less calcemic but that
exhibit potent antiproliferative activity would be attractive as
therapeutic agents. We investigated the effects of two different types
of less calcemic vitamin D compounds, 25-hydroxyvitamin D3
[25(OH)D3] and 19-nor-1,25-dihydroxyvitamin
D2 [19-nor-1,25(OH)2D2],
and compared their activity to 1,25(OH)2D3
on (a) the proliferation of primary cultures and cell
lines of human prostate cancer cells; and (b) the
transactivation of the VDRs in the androgen-insensitive PC-3 cancer
cell line stably transfected with VDR (PC-3/VDR).
19-nor-1,25(OH)2D2, an analogue of
1,25(OH)2D3 that was originally developed
for the treatment of parathyroid disease, has been shown to be less
calcemic than 1,25(OH)2D3 in clinical
trials. Additionally, we recently showed that human prostate cells in
primary culture possess 25(OH)D3-1-hydroxylase, an
enzyme that hydroxylates the inactive prohormone, 25(OH)D3,
to the active hormone, 1,25(OH)2D3,
intracellularly. We reasoned that the hormone that is formed
intracellularly would inhibit prostate cell proliferation in an
autocrine fashion. We found that 1,25(OH)2D3
and 19-nor-1,25(OH)2D2 caused similar
dose-dependent inhibition in the cell lines and primary cultures in the
[3H]thymidine incorporation assay and that both compounds
were significantly more active in the primary cultures than in LNCaP
cells. Likewise, 25(OH)D3 had inhibitory effects comparable
to those of 1,25(OH)2D3 in the primary
cultures. In the chloramphenicol acetyltransferase (CAT)
reporter gene transactivation assay in PC-3/VDR cells,
1,25(OH)2D3 and
19-nor-1,25(OH)2D2 caused similar increases
in CAT activity between 10-11 and 10-9
M. Incubation of PC-3/VDR cells with 5 x
10-8 M 25(OH)D3 induced a 29-fold
increase in CAT activity, similar to that induced by 10-8
M 1,25(OH)2D3. In conclusion,
our data indicate that 25(OH)D3 and
19-nor-1,25(OH)2D2 represent two different
solutions to the problem of hypercalcemia associated with vitamin
D-based therapies: 25(OH)D3 requires the presence of
1-hydroxylase, whereas 19-nor-1,25(OH)2D2
does not. Both drugs are approved for human use and may be good
candidates for human clinical trials in prostate cancer. |
INTRODUCTION |
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TopABSTRACTINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONREFERENCES |
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In addition to androgens, it is now clear that prostatic cells are
responsive to another class of steroid hormones, namely, vitamin D
(3)
. Most human prostate cells contain specific
intracellular receptors (commonly called
VDRs)3
for
1,25(OH)2D, the active hormonal form of
vitamin D (4
, 5)
. Numerous studies have shown that in
response to 1,25(OH)2D3,
prostate cancer cells show an increase in differentiation and a
decrease in proliferation, invasiveness, and metastasis
(6, 7, 8, 9)
. These findings strongly support the use of vitamin
D-based therapies for prostate cancer, e.g., as
differentiation therapy and/or as a second-line therapy once androgen
deprivation has failed. However, the use of
1,25(OH)2D-based therapies for prostate cancer
is limited by the risk of hypercalcemia and hypercalciuria (10
, 11)
. Thus, less calcemic or noncalcemic analogues of
1,25(OH)2D3 with potent
antiproliferative activity would be attractive therapeutic agents.
Recently, our group has shown that human prostate cancer cells in
primary culture and several prostate cancer cell lines possess
1-OHase, the enzyme that converts the major circulating, prohormonal
form of vitamin D, 25(OH)D, to
1,25(OH)2D (12
, 13)
. Because the
conversion from prohormone to active hormone occurs within the cell,
the problem of systemic hypercalcemia should be avoided. 25(OH)D would
be an attractive candidate for human clinical trials in prostate cancer
because this drug has been approved by the FDA for human use
(e.g., for treating vitamin D deficiency due to liver
disease; Ref. 14
).
Similarly,
19-nor-1,25(OH)2D2, a
synthetic analogue of
1,25(OH)2D2, has
recently been approved by the FDA for the treatment of secondary
hyperparathyroidism. Several randomized controlled clinical trials have
shown that
19-nor-1,25(OH)2D2 is
noncalcemic (15
, 16)
. The structural similarity of
19-nor-1,25(OH)2D2 to
1,25(OH)2D3 suggested to
us that the behavior of
19-nor-1,25(OH)2D2 in
prostatic cells might be similar to that of
1,25(OH)2D3.
In this report, we investigated the effect of
25(OH)D3,
1,25(OH)2D3, and
19-nor-1,25(OH)2D2 on
the proliferation of primary cultures and cell lines of human prostate
cancer. In addition, we evaluated the abilities of these three vitamin
D compounds to transactivate the VDR in a prostate cancer cell line,
PC-3, that was stably transfected with VDR.
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MATERIALS AND METHODS |
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TopABSTRACTINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONREFERENCES |
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,25(OH)2D3 were a
generous gift from Dr. M. Uskokovic (Hoffman-La Roche, Nutley, NJ).
19-nor-1,25(OH)2D2 was a
gift from Tetrionics (Madison, WI).
Cell Cultures.
Prostate cancer cell lines, LNCaP and PC-3 cells, were obtained from
the American Type Culture Collection (Rockville, MD) and were grown on
24-well culture dishes with DMEM (Life Technologies, Inc.) supplemented
with 5% FBS (Life Technologies, Inc.). Cells were fed three times per
week. Primary cultures of human prostate epithelial cells were prepared
as described previously (17)
. Prostate epithelial cells
were cultured in a serum-free defined-growth medium (Prostate
Epithelial Growth Medium BulletKit, Clonetics, San Diego, CA). Prostate
cells used for this study were at their second passage.
[3H]Thymidine Incorporation.
[3H]thymidine incorporation into DNA was used
as an index of cell proliferation as described previously
(18)
. Briefly, when LNCaP cells or the second passage
primary culture cells reached about 50% confluency, FBS (in the case
of LNCaP) or growth factors (in the case of primary cultures) were
removed from the media, and cells were grown for an additional 24 h in the absence of FBS or growth factors. Cells were then treated with
and without different concentrations of 25(OH)D3,
1,25(OH)2D3, or
19-nor-1,25(OH)2D2 as
indicated in the figure legends. Eighteen h later, the dosing medium
was replaced with 0.5 ml of fresh basal medium containing
[methyl-3H]thymidine (New England Nuclear,
Boston, MA) and incubated for 3 h at 37°C.
[3H]Thymidine incorporation into DNA was
stopped by placing the 24-well plates on ice. Unincorporated
[3H]thymidine was then removed, and the cells
were washed three times with ice-cold PBS. DNA labeled with
[3H]thymidine and other macromolecules were
first precipitated with ice-cold 5% perchloric acid for 20 min and
then extracted with 0.5 ml of 5% perchloric acid at 70°C for 20 min
as described previously (19)
. The radioactivity in the
extracts was determined by a liquid scintillation counter. The results
were expressed as percent of control.
Morphological Studies during Cellular Proliferation.
The second-passage primary culture cells were subcultured in the
complete medium into 35-mm dishes for the morphological studies
(18)
. Two days after the initial plating, triplicate
plates of cells were incubated with complete media without insulin but
containing 25(OH)D3,
1,25(OH)2D3, or vehicle.
Cells were dosed again with 25(OH)D or
1,25(OH)2D3 2 and 4 days
later. Three days after the last dosing, the media were removed from
cultures. The attached cells were then trypsinized for 30 min with
0.1% EDTA and 0.1% trypsin at 37°C and then neutralized with basal
medium. The detached cells were spun down and resuspended in a known
volume of basal medium. Triplicate aliquots were applied to a
hemocytometer for cell counting.
Recombinant Plasmids, Transfection, and CAT Assay.
The reporter plasmid MOPVDREtkCAT was constructed as described
previously (19)
and consisted of two copies of the
VDRE of the MOP gene linked 5' to the tk promoter and
CAT gene of the vector pBLCAT2 (20)
.
MOPVDREtkCAT was transfected into a PC-3/VDR clone (clone 3B2) using
the calcium phosphate method. CMV-ß-gal, which encodes the
ß-gal gene driven by the CMV promoter, was included in all
of the transfections to normalize for differences in transfection
efficiency. Clone 3B2 (PC-3/VDR) was generated by transfecting PC-3
cells with the VDR cDNA expression vector pRc CMV-VDR followed
by the selection and expansion of stable clonal isolates as described
previously (21)
. The PC-3/VDR and LNCaP cell line
expressed comparable levels of VDR (approximately 25 fmol/mg protein).
The PC-3/VDR cells transfected with the reporter gene were then
cultured in the presence or absence of 10-8
M 25(OH)D3,
1,25(OH)2D3, or
19-nor-1,25(OH)2D2 in
RPMI containing 10% FBS. Cells were harvested about 40 h after
transfection, and cell extracts were prepared for analyzing ß-gal and
CAT activity. Cell extracts containing equivalent amounts of ß-gal
activities were used for an analysis of CAT using an adaptation of the
method of Gorman et al. (22)
The percentage of
conversion of [14C]chloramphenicol to
acetylated forms on thin-layer chromatograms was quantified using a
Molecular Dynamics Phosphorimager and image Quant software (Sunnyvale,
CA).
Statistical Analysis.
Comparisons of the antiproliferative and transactivation activities
between controls and drug-treated groups, and between two different
drugs, were performed using one-way ANOVA. Differences between groups
were considered statistically significant when Ps were 
0.05.
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RESULTS |
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,25(OH)2D3 and
19-nor-1,25(OH)2D2 on
the [3H]thymidine incorporation into cultured
LNCaP prostate cells are shown in Fig. 1
.
The figure demonstrates that
1,25(OH)2D3 and
19-nor-1,25(OH)2D2 at
10-7 M inhibited
[3H]thymidine incorporation into DNA 21 ±
4% and 18 ± 5%, respectively, as compared with the controls in
the absence of
1,25(OH)2D3 or
19-nor-1,25(OH)2D2. At
10-6 M,
1,25(OH)2D3 and
19-nor-1,25(OH)2D2
inhibited [3H]thymidine incorporation 63 ± 1% and 60 ± 1%, respectively. No significant inhibition was
detected in the presence of 10-8 M
of 1,25(OH)2D3 or
19-nor-1,25(OH)2D2.
Thus, in LNCaP cells,
19-nor-1,25(OH)2D2 was
as active as 1,25(OH)2D3
in inhibiting [3H]thymidine incorporation at
10-7 and 10-6
M.
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shows a dose-dependent
antiproliferative effect of
1,25(OH)2D3 and
19-nor-1,25(OH)2D2 in
the primary cultures of human prostate cancer cells.
1,25(OH)2D3 caused an
18 ± 3%, 41 ± 1%, 77 ± 1%, and 86 ± 1%
inhibition of [3H]thymidine incorporation into
DNA at 10-9, 10-8,
10-7, and 10-6
M, respectively; and
19-nor-1,25(OH)2D2
caused a 20 ± 5%, 39 ± 2%, 80 ± 1%, and 88 ±
1% inhibition of [3H]thymidine incorporation
into DNA at the same concentrations, respectively. The data show
that the activities of
1,25(OH)2D3 and
19-nor-1,25(OH)2D2 in
the primary prostate cultures are very similar. Both compounds were
more effective in inhibiting [3H]thymidine
incorporation in the primary cultures than in LNCaP cells.
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-OHase mRNA and its
enzyme activity in cultured prostate cancer cell lines and in primary
cultures derived from normal, benign prostatic hyperplasia and prostate
cancer cells (PC-3 and DU145; 12
, 13 ). Unlike the primary
cultures, DU145 and PC-3 cells, very little 1-OHase activity
was detected in LNCaP cells. We reasoned that
25(OH)D3 should be converted to
1,25(OH)2D3 by prostatic
cancer cells that possess 1-OHase and that the
1,25(OH)2D3 would cause
an inhibition in the proliferation of these cells. Fig. 3
demonstrates that when primary cultures
of prostatic cells were treated with 25(OH)D3 or
1,25(OH)2D3 for 7 days,
the two vitamin D3 metabolites caused a similar
inhibition of cell proliferation. At 10-6
M, 25(OH)D3 and
1,25(OH)2D3 inhibited
cell proliferation by decreasing cell number 60 ± 3% and 60 ± 1%, respectively, and at 10-7 M,
25(OH)D3 and
1,25(OH)2D3 inhibited
cell proliferation by decreasing cell number 24 ± 1% and 25 ± 5% in primary cultures of prostate cancer cells, respectively. At
10-8 M, neither compound exhibited
significant activity. Using the [3H]thymidine
incorporation assay for cell proliferation, we also demonstrated that
25(OH)D3 was highly active in inhibiting primary
prostate cell proliferation (Fig. 4)
.
25(OH)D3 caused 33 ± 7%, 46 ± 4%,
and 70 ± 3% inhibition at 10-9,
10-8, and 10-7
M, respectively, as compared with 55 ± 4%, 60 ± 4%, and 73 ± 4% inhibition by
1,25(OH)2D3. Thus, in
these assays, 25(OH)D3 and
1,25(OH)2D3 were
equipotent in inhibiting the proliferation of prostate cancer cells at
10-7 M.
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,25(OH)2D2, and
1,25(OH)2D3 to
transactivate VDR in PC-3 cells stably transfected with VDR. PC-3 cells
are well-characterized models of androgen-independent prostate cancer
and were previously shown to contain 1-OHase activity
(12)
. However, unlike many human prostate cancer cells,
PC-3 cells express extremely low levels of VDR. Thus, we chose a
derivative of these cells that have been stably transfected with a VDR
cDNA (PC-3/VDR). To assess the ability of VDR to be activated by
endogenously synthesized
1,25(OH)2D3, we
performed CAT reporter gene transactivation assays in
PC-3/VDR cells cultured in 25(OH)D3. DPPD was
added to inhibit the auto-oxidation of 25(OH)D3
to 1,25(OH)2D3. The
CAT reporter gene plasmid contains two tandem copies of the
VDRE found in the MOP promoter.
1,25(OH)2D3 caused a 2-,
16-, 37-, and 15-fold increase, and
19-nor-1,25(OH)2D2
caused a 3-, 21-, 40-, and 32-fold increase over the controls in CAT
activity at 10-11, 10-10,
10-9, and 10-8
M, respectively (Fig. 5)
. Incubation of PC-3/VDR cells with
5 x 10-8 M
25(OH)D3 induced a 29-fold increase over the
controls in CAT activity, which was similar to that induced by
10-8 M
1,25(OH)2D3 (Fig. 6)
.
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DISCUSSION |
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TopABSTRACTINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONREFERENCES |
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,25(OH)2D3 have
extended far beyond its classical role on intestine, bone, kidney, and
parathyroid glands to regulate serum calcium levels.
1,25(OH)2D3 has been
shown to have important antiproliferative and prodifferentiating
activities in a variety of tissues or cells that possess VDRs,
including prostatic cells (4, 5, 6, 7, 8, 9)
. The epidemiological
similarities between prostate cancer and vitamin D deficiency
(23, 24, 25)
and the impressive anticancer effects of
1,25(OH)2D3 on prostatic
cells has led to great interest in the use of this hormone as a
therapeutic agent for prostate cancer. However, a major drawback of
using vitamin D-based therapies for prostate cancer is hypercalcemia
(10
, 11) . Many different types of non- or less-calcemic
vitamin D analogues have been investigated for their effects on
prostate cancer cell proliferation in vitro. Potent
inhibitors of prostate cancer cell proliferation include
19-nor-hexafluoride vitamin D3 and
20-cyclopropyl-vitamin D3 analogues (26
, 27)
. Hisatake et al. (28)
recently
reported that 5,6-trans-16-ene-Vitamin
D3 analogues were more potent than
1,25(OH)2D3 in
inhibiting LNCaP cells in vitro and were about 40-fold less
calcemic than
1,25(OH)2D3 in normal
mice in vivo. However, the antiproliferative effects of
these analogues has yet to be demonstrated in vivo.
Recently, it was demonstrated that EB1089, an analogue of
1,25(OH)2D3, was as
effective as 1,25(OH)2D3
in inhibiting metastasis in an in vivo model of
androgen-insensitive prostate cancer, the rat Dunning MAT LyLu prostate
cancer model (9)
. Although EB1089 was significantly less
calcemic than
1,25(OH)2D3, it still
caused an 18% increase in serum calcium level (versus a
34% increase by
1,25(OH)2D3). Thus, less
calcemic or noncalcemic analogues of
1,25(OH)2D3 are still
needed. Llach et al. (15)
reported that
19-nor-1,25(OH)2D2 was
as effective as
1,25(OH)2D3 in
suppressing parathyroid hormone secretion in hemodialysis patients with
secondary hyperparathyroidism without inducing hypercalcemia or
hyperphosphatemia. In the current study, we examined the
antiproliferative activity of
19-nor-1,25(OH)2D2 in
LNCaP cells and in primary cultures of prostate cancer cells. In these
two different cultures,
19-nor-1,25(OH)2D2
showed antiproliferative effects similar to those of
1,25(OH)2D3, as
determined by [3H]thymidine incorporation
(Figs. 1
and 2)
. Both compounds had a greater effect in the primary
cultures of prostate cancer cells than in the LNCaP prostate cancer
cell line, which suggests that primary cultures may be a more sensitive
system to differentiate the effectiveness of different vitamin D
compounds in vitro.
1,25(OH)2D3 was
previously shown (21)
to decrease cyclin-dependent kinase
2 activity, resulting in decreased retinoblastoma protein
phosphorylation and accumulation of LNCaP cells in
G1 phase of the cell cycle. Because a functional
retinoblastoma pathway seems to be required for the maximal
antiproliferative effects of
1,25(OH)2D3, primary
prostatic cultures may exhibit increased growth inhibition by
1,25(OH)2D3,
25(OH)D3, and
19-nor-1,25(OH)2D2
because, compared with the cell lines, these cultures are less likely
to have mutations in this pathway.
Because the clinical use of
1,25(OH)2D3 in cancer
therapy is limited by the risk of hypercalcemia, many investigators
have attempted to duplicate the antiproliferative effects of
1,25(OH)2D3 in vivo
using analogues of
1,25(OH)2D3 that are
less calcemic, such as
19-nor-1,25(OH)2D2,
16-ene-23-yne-1,25(OH)2D3
(29)
, and EB1089 (30)
, or they have used a
combination of
1,25(OH)2D3 or
1,25(OH)2D3 analogues
with other drugs (31
, 32)
. However, our discovery that
prostate cells in primary culture express high 1-OHase activity and
can synthesize
1,25(OH)2D3 from
25(OH)D3 suggests that
25(OH)D3 may offer another potential solution to
the problem of hypercalcemia caused by the systemic administration of
1,25(OH)2D3. This is
because 1,25(OH)2D3
would be synthesized intracellularly, act in an autocrine fashion, and
be degraded, and would not be expected to leak into the systemic
circulation and cause hypercalcemia. Our results confirm that
25(OH)D3 is highly active in inhibiting prostate
cells proliferation in vitro (Figs. 3
and 4)
. Similar
antiproliferative effects are observed when
25(OH)D3 is administered to primary cultures of
prostate cells in clonogenic
assays.4
Because
25(OH)D3 binds to the VDRs with only 0.001 to
0.002 of the binding affinity of
1,25(OH)2D3
(33)
, we consider it unlikely that these results are due
to the direct actions of 25(OH)D3 on the VDRs.
Rather, we suggest that these results reflect the conversion of
25(OH)D3 to
1,25(OH)2D3 by
1-OHase present in prostate cells. This conclusion is further
supported by experiments in which LNCaP cells were transfected with
1-OHase cDNA. These cells, and not untransfected cells, responded to
25(OH)D3 by an inhibition of
[3H]thymidine
incorporation.5
Experiments
to determine whether similar effects can be produced in vivo
using human tumor cells xenografted into athymic mice are presently
underway.
Although kidney cells are the "classic" cells that possesses
1-OHase, 1,25(OH)2D levels produced by the
kidney are very tightly regulated by serum levels of parathyroid
hormone (34)
. Thus, in normal individuals, even large
increases in serum 25(OH)D will not result in increased systemic levels
of 1,25(OH)2D (35
, 36)
. However,
the extrarenal synthesis of 1,25(OH)2D is
generally unregulated (13
, 37)
. This suggests that
increases in systemic levels of 25(OH)D could result in increased local
production of 1,25(OH)2D in some extrarenal
sites (i.e., prostate) without producing hypercalcemia.
Two commonly used assay methods were used to study the
antiproliferative activity of 25(OH)D3 in the
primary cultures of prostate cells:
[3H]thymidine incorporation and cell count. The
former involved a short-term, 18-h incubation with the hormone and
studied the effect of the drugs on DNA synthesis.
[3H]thymidine incorporation is an index of cell
division. In certain cell types, decreases in
[3H]thymidine incorporation may also reflect
increases in cell differentiation (38)
. Conversely, cell
count, which requires longer-term incubation (7 days) with the hormone,
reflects cell growth only. The difference in dose-response curves that
we observed in the two assays may reflect differences in the incubation
time of these assays because: (a) with longer incubation
times, more 25(OH)D3 would be converted to
1,25(OH)2D3
intracellularly, and, thus, the effects of
25(OH)D3 would be similar to the effects of the
direct addition of
1,25(OH)2D3; and
(b) more exogenously added
1,25(OH)2D3 would likely
increase its own degradation by inducing the expression of
24-hydroxylase (34)
.
Most of the antiproliferative effects of
1,25(OH)2D3 and its
analogues are believed to be mediated through the functional expression
of VDR. We, therefore, compared the transactivation activity of
1,25(OH)2D3 and
19-nor-1,25(OH)2D2 in PC-3
cells that were stably transfected with VDR (PC-3/VDR), using the
reporter plasmid MOPVDREtkCAT. This plasmid, which contains two copies
of the VDRE found in the MOP gene, was chosen because
previous studies in other well-characterized prostate cancer cell lines
indicated that VDR transcriptional activity could be detected using
this VDRE-containing reporter, even in cell lines that expressed
extremely low levels of VDR (9
, 21)
. Using this system, we
demonstrated that
1,25(OH)2D3 and
19-nor-1,25(OH)2D2 had
almost identical transactivation activity (Fig. 5)
in agreement with
our [3H]thymidine incorporation data (Figs. 1
and 2)
. 25(OH)D3 at 5 x
10-8 M showed a comparable
transactivation activity as that caused by 10-8
M
1,25(OH)2D3, consistent
with our finding that PC-3 cells have the capacity to convert
25(OH)D3 to
1,25(OH)2D3
(12)
.
In summary, this report demonstrates that, like
1,25(OH)2D3,
19-nor-1,25(OH)2D2, and
25(OH)D3 possess potent antiproliferative effects
on human prostate cancer cell lines and on primary cultures of human
prostate cancer cells. Both
19-nor-1,25(OH)2D2 and
25(OH)D3 are equipotent to the parent hormone in
their ability to transactivate the VDR. Although both of these vitamin
D compounds act ultimately on the VDR, their proximal biological
targets are different: 25(OH)D3 requires the
presence of 1-OHase, whereas
19-nor-1,25(OH)2D2 does
not. Our studies of prostate cancer cell lines have shown a large
variation in the expression of 1-OHase. For example, although LNCaP
cells showed profound growth inhibition by
1,25(OH)2D3, these cells
do not express measurable levels of 1-HOase message and activity
(12
, 13)
and, accordingly, are not growth-inhibited by
25(OH)D3 (39)
. This suggests that
prostatic tumors that do not express 1-OHase should be treated
with 1,25(OH)2D3
analogues, such as
19-nor-1,25(OH)2D2 or
EB1089. Because both 25(OH)D3 and
19-nor-1,25(OH)2D2 are
known to be noncalcemic within a wide dosing range (14)
and both are approved for human use (for other indications), these
vitamin D compounds may be excellent candidates for human clinical
trials in prostate cancer, especially for prostate cancers that have
failed conventional therapies such as androgen deprivation.
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ACKNOWLEDGMENTS |
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,25(OH)2D2. |
FOOTNOTES |
|---|
2 To whom requests for reprints should be
addressed, at Vitamin D, Skin and Bone Research Laboratory, Rm M-1022,
Boston University School of Medicine, 715 Albany Street, Boston, MA
02118. Phone: (617) 638-4543; Fax: (617) 638-8882; E-mail: taichen{at}bu.edu
3 The abbreviations used are: VDR, vitamin
D receptor; 25(OH)D, 25-hydroxyvitamin D; 25(OH)D3,
25-hydroxyvitamin D3; 1,25(OH)2D3,
1,25-dihydroxyvitamin D3;
19-nor-1,25(OH)2D2,
19-nor-1,25-dihydroxyvitamin D2; 1-OHase,
1-hydroxylase; FBS, fetal bovine serum; CAT, chloramphenicol
acetyltransferase; DPPD, 1,2-dianilinoethane; VDRE, VDR element; MOP,
mouse osteopontin; tk, thymidine kinase; CMV, cytomegalovirus; ß-gal,
ß-galactosidase.
4 A. Barreto, G. G. Schwartz, P. Woodruff,
and S. D. Cramer. 25-Hydroxyvitamin D3, the prohormone
of 1,25-dihydroxyvitamin D3, inhibits the proliferation of
primary cultures of prostate epithelial cells, submitted for
publication.
Received 8/20/99; revised 11/29/99; accepted 11/29/99.
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TopABSTRACTINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONREFERENCES |
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,25-dihydroxyvitamin D3. Cancer Res., 52: 515-520, 1992.,25-Dihydroxyvitamin D (calcitriol) inhibits the invasiveness of human prostate cancer cells. Cancer Epidemiol. Biomark. Prev., 6: 727-732, 1997.-hydroxylase in cultured human prostatic cells. Bone, 23(Suppl.): S262 1998.
,25-dihydroxyvitamin D3 in human prostate cancer cell line LNCaP involves reduction of cyclin-dependent kinase 2 activity and persistent G1 accumulation. Endocrinology, 139: 1197-1207, 1998.,25-Dihydroxyvitamin D3 and platinum drugs act synergistically to inhibit the growth of prostate cancer cell lines. Clin. Cancer Res., 5: 695-703, 1999.,25-dihydroxyvitamin D2/D3. J. Clin. Investig., 58: 61-70, 1976.
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