IBYME   02675
INSTITUTO DE BIOLOGIA Y MEDICINA EXPERIMENTAL
Unidad Ejecutora - UE
congresos y reuniones científicas
Título:
Progestin regulate proliferation of breast cancer through a PR/STAT3/c-Myc/miR-16 and Cyclin E dependent pathway
Autor/es:
MARTÍN A. RIVAS, YI WEN HUANG, ROXANA SCHILLACI, TIM HUI-MING HUANG, PATRICIA V. ELIZALDE.
Reunión:
Congreso; 102nd Annual Meeting of the American Association for Cancer Research; 2011
Resumen:
Breast cancer is the
commonest type of cancer among women in the US and the second leading
cancer-death cause. Understanding its biology will help to design new clinical
approaches to achieve a better outcome of the disease. microRNA (miRNA) are
short ribonucleic acids with important regulatory functions and with an
increasingly acknowledged role in cancer. The present study was designed to
characterize progestin-induced miRNA regulation in breast cancer. Primary
cultures of the murine breast cancer C4HD were treated or not with 10nM of the
synthetic progestin medroxiprogesterone acetate (MPA) for 24 h and total RNA
extracted for miRNA profiling using Applied Biosystems low density qPCR arrays.
Analysis of data rendered a total of 16 miRNA significantly regulated
(P<0.05): miR-378*, miR-376a, miR-224, miR-190b, miR-16, miR-410 and miR-197
were downregulated and miR-191*, miR-17*, miR-470*, miR-451, miR-702,
miR-434-3p, miR-493, miR-23a* and miR-485* were upregulated under treatment
with MPA. Interestingly, miR-16 has already been shown to be a tumor suppressor
in leukemia and so we focused our further study on it. We validated the
downregulation of miR-16 by qPCR (0.4 fold against control) and observed that
small interference RNA (siRNA) to progesterone receptor (PR) or to signal
transducer and activator of signal 3 (STAT3) overcame miR-16 downregulation by
MPA. The oncogene c-Myc was a reported negative regulator of miR-16, and we
observed that siRNA to PR or STAT3 inhibited MPA-induced c-Myc upregulation. A
query for miR-16 targets on miRecords search engine showed that the cell cycle
promoter gene Cyclin E has conserved putative target sites on 3'UTR mRNA.
Consistently, siRNA to PR and to STAT3 inhibited MPA-induced CCNE mRNA and
Cyclin E protein increase as assessed by qPCR and Western blot. In addition,
transfection with pre-miR-16 abrogated MPA capacity of inducing Cyclin E
upregulation. Most importantly, pre-miR-16 inhibited MPA-induced proliferation,
as observed in [3H]-Thymidine incorporation assays and Trypan-blue cell counts.
C4HD is a progestin-dependent tumor since it is unable to grow in vivo under
exogenous progestin's absence. Therefore, 1 x 106 C4HD cells transfected with a
pre-miR-control (CTRL) or with pre-miR-16 were injected s.c. in 8-week female Balb/c mice which had
been previously injected with a MPA slow-release depot. Tumors became palpable
at day 15 but the ones from pre-miR-16 group grew at a lower rate than the ones
from pre-miR-CTRL group, and the mean tumor volume at day 40 was significantly
smaller (312.4 ± 42.3 mm3 for CTRL group against 22.2 ± 34.9 mm3 for pre-miR-16
group). To sum up, these results suggest that progestin induce breast cancer
growth through a pathway involving PR/STAT3/c-Myc and miR-16 which acts on mRNA
targets such as Cyclin E to control cell cycle progression.