| Abstract |
At present, not more than a few permanent human prostate cancer cell lines
have been described. The only androgen responsive in vitro cell line is
LNCaP, which exhibits an atypical hormone response pattern due to a
mutated androgen receptor. The PC-346 xenograft cell line is the only
human prostate cell line known thus far in which initial regression upon
androgen withdrawal is followed by tumor relapse, suggesting the presence
of hormone-dependent and -independent cell populations, Recently we
established in vitro cultures from the PC-346 xenograft model following
collagenase digestion of PC-346 tumor fragments from the 12th mouse
passage. After plating in Matrigel coated culture flasks with low serum
(2%) culture medium, poorly spread epithelial colonies were obtained that
could be passaged by trypsinization. Contamination by (murine) stromal
cells was no longer detectable after two passages. Currently, this new in
vitro cell line, PC-346C, is in its 20th passage. The cells express
prostate specific markers such as PSA (resulting in levels >2500 ng/ml in
culture medium) and, unlike LNCaP, also prostate-specific TGase. Hormone
responsiveness of PC-346C cells was tested in a modified MTT assay. In the
absence of steroids, little if any growth occurred. Dose-dependent
stimulation of growth was observed after addition of testosterone,
dihydrotestosterone or R1881. Even upon androgenic stimulation, however,
growth was relatively slow (doubling time 4-5 days). In contrast to LNCaP,
PC-346C cells were not stimulated by estradiol or flutamide.
Immunocytochemistry, binding assays and Western blot analysis showed the
presence of androgen receptors. Complete sequencing of exons 2 through 8
of the receptor gene demonstrated that mutations were absent in this part
of the gene. Injection of 10x10^6 PC-346C cells into male nude mice
resulted in rapidly growing tumors (doubling time 5.2 +- 1.1 days) within 30
days in 9 of 10 animals. High levels of PSA (>1000 ng/ml) were measured in
the serum of these animals. In the same experiment, no growth was detected
in females until 95 days post-inoculation. Finally, growth occurred in
2/10 female male. From these tumors, another in vitro line (PC346CF) was
established. PC-346CF cells were not stimulated by androgens in vitro.
Further characterization of this new subline and comparative studies of
PC-346C and PC-346CF cells are currently underway. By virtue of their unique
properties, PC-346C and its hormone-insensitive variant will be valuable
new models to study the mechanisms of androgen-regulated growth and gene
expression.
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