p53-Mdm2 interaction targeted therapy by Nutlin-3 on nasopharyngeal carcinoma cells / Voon Yee Lin
Nutlin-3, a small-molecule inhibitor of p53-Mdm2 interaction, is known to be effective against cancers expressing wild-type (wt) p53. p53 mutations are rare in nasopharyngeal carcinoma (NPC), and hence targeting the disruption of p53-Mdm2 interaction to reactivate p53 may offer a promising therap...
Saved in:
Main Author: | |
---|---|
Format: | Thesis |
Published: |
2016
|
Subjects: | |
Online Access: | http://studentsrepo.um.edu.my/7090/4/yee_lin.pdf http://studentsrepo.um.edu.my/7090/ |
Tags: |
Add Tag
No Tags, Be the first to tag this record!
|
Summary: | Nutlin-3, a small-molecule inhibitor of p53-Mdm2 interaction, is known to be
effective against cancers expressing wild-type (wt) p53. p53 mutations are rare in
nasopharyngeal carcinoma (NPC), and hence targeting the disruption of p53-Mdm2
interaction to reactivate p53 may offer a promising therapeutic strategy for NPC. This
study hypothesized that reactivation of p53 in NPC cells may suppress NPC cell
proliferation, and in addition, Nutlin-3 combined with cisplatin may further suppress the
cancer cell proliferation more effectively. To investigate these possibilities, the effects
of Nutlin-3 alone or in combination with cisplatin were tested on C666-1, an Epstein
Barr virus (EBV)-positive NPC cell line bearing wt p53, in parallel with normal
nasopharyngeal epithelial (NPE) NP69 and NP460 cells. Single drug treatment resulted
in a concentration-dependent inhibitory effect on the cancer cell proliferation. Cisplatin
was more cytotoxic to the NPE cells compared to the NPC cells, while Nutlin-3 was
more effective and selective in inhibiting NPC cells. Cisplatin combined with Nutlin-3
showed stronger anti-proliferative effect against NPC cells and markedly suppressed its
anchorage-independent growth on soft agar, suggesting that combined treatment was
more effective than single drug therapy. Prior verification showed that C666-1, NP69
and NP460 cells retained the wt p53. Treatrnent with Nutlin-3 showed significant
accumulation of p53, p2lWafl/Cipl and Mdm2 proteins in cells expressing wt p53 in
comparison to p53-mutated cells. The effect of Nutlin-3 on the restoration of p53,
p2lWafl/Cip1 and Mdm2 expression was impaired following p53-knocked down in
NPC cells, and likewise the cells with p53 knockdown showed less sensitivity to Nutlin
3. These findings suggest that Nutlin-3 activates the p53 pathway and exerts its
cytotoxicity on NPC cells in a pS3-dependent manner. The accumulation of Annexin
V/Pl-stained cells showed treatment of NPC cells with cisplatin resulted in apoptosis
and Nutlin-3-treated cells showed less percentage of apoptotic cells compared to the
lll
cisplatin-treated cells. Apoptosis, however, increased significantly in the cells teated
with cisplatin and Nutlin-3. Similarly, Nutlin-3 positively upregulated BAX and PUMA
protein expressions in NPC cells. The expression levels of these proteins also increased
significantly in cells treated with cisplatin and Nutlin-3, concomitant with the detection
of cleaved PARP level. Taken together, these findings suggest that Nutlin-3 sensitises
NPC cells to cisplatin-induced apoptosis by modulating pro-apoptotic targets via the
p53 pathway. In addition, an extended treatment period of NPC cells with Nutlin-3 did
not result in the emergence of p53-mutated cells, albeit reduced sensitivity to Nutlin-3
was observed. This stresses on the importance of treabnent duration and clinical doses
optimization to improve the efficacy of Nutlin-3 significantly. Therefore, the overall
findings revealed supportive evidence of the effectiveness of combining cisplatin with
Nutlin-3 as potential therapy against NPC.
|
---|