| Abstract |
Gonadotrophin-releasing hormone (GnRH) can inhibit proliferation of
multiple reproductive tissue cancer cell lines through direct interaction
with GnRH receptors (GnRHR) on tumour cells. GnRH analogues may therefore
have a role in treating some cancers. The signalling pathways associated
with these inhibitory effects are poorly defined, and characterising them
may help to understand therapeutic sensitivity. To elucidate these
pathways, transcriptomic and proteomic approaches were used to compare the
effects of the GnRH agonist Triptorelin in responsive GnRHR-transfected
HEK293 cells (SCL60) and unresponsive (HEK293) cells both in vitro for up
to 24h and in vivo for up to 7 days. Gene expression profiling
demonstrated that SCL60 gene expression was temporally regulated with
Triptorelin treatment, with expression of some genes increased at one time
point but decreased at another. Early and mid-phase gene expression
changes comprised mainly transcription factors and late changes included
the hormonal signalling component CGA. Pathway analysis implicated mitogen-
activated protein kinase and cell cycle pathways, supporting the detection
of G2/M arrest. Signalling effects within SCL60 xenografts, 4 and 7 days
following Triptorelin treatment, were investigated using a
phosphoproteomic antibody array. Changes included cell cycle and apoptosis
regulators, as well as cell surface receptors and NFkappaB signalling
pathway members. Reverse-phase protein arrays and western blotting also
showed that pAkt was decreased and pNFkappaB-p65 was increased after
Triptorelin treatment in vitro. An NFkappaB inhibitor enhanced the anti-
proliferative effect of Triptorelin in SCL60 cells in vitro, suggesting
that NFkappaB acts as a survival factor in the response to GnRHR
stimulation. A range of GnRHR expression was observed in breast cancer
tumours by immunohistochemistry, and on average GnRHR expression was
significantly higher in the Triple Negative Phenotype (TNP) subgroup and
in grade 3 tumours. A GnRHR-transfected breast cancer cell line, MCF7-h14,
was developed. Despite this expressing a similar level of GnRHR to
responsive SCL60 cells, MCF7-h14 cells were not inhibited by GnRHR
activation, indicating that a high level of GnRHR is insufficient for the
antiproliferative effects of Triptorelin.
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