Cyclin dependent kinases (Cdks) are a family of conserved serine/threonine kinases that regulate cell cycle progression in mammalian cells. Activation of individual Cdks at distinct phases of the cell cycle ensures the proper timing and coordination of cell cycle events. Over the past decade, gene knockout models combined with studies using small molecule kinase inhibitors have shown that the effect of individual Cdk inactivation is highly dependent on cellular and genetic context. Here we examine the effect of Cdk1 inhibition on embryonic stem (ES) cells. Additionally, we develop a chemical genetic approach that allows for the selective inhibition of Cdk2 in multiple cell and cancer types.
Embryonic stem (ES) cells are an attractive source for stem cell therapies due to their rapid proliferation and capacity for differentiation. A limitation in the field of regenerative medicine however, is the propensity for ES cells to form teratomas when transplanted in vivo. Selective depletion of undifferentiated cells during regeneration therapies could reduce the carcinogenic risks of these procedures. We show that inhibiting Cdk1 results in the activation of a DNA damage response, nuclear p53 stabilization, and induction of pro-apoptotic p53 target genes in ES but not differentiated cells. Furthermore we show that clinically relevant Cdk1 inhibitors prevent formation of ES cell-derived tumors and inhibit growth of established ES cell-derived teratomas in vivo. Our data demonstrate that ES cells are uniquely sensitive to Cdk1 inhibition, and identify Cdk1 as a pharmacological target that could increase the safety of regeneration therapies. In an independent project, we use a chemical-genetic approach to achieve selective inhibition of Cdk2 kinase activity using an analog sensitive (AS) allele. We show that inhibition of Cdk2 kinase activity slows proliferation of non-transformed cells, whereas siRNA knockdown of Cdk2 does not, highlighting the differences between these approaches. We also show that Cdk2 inhibition attenuates anchorage-independent growth of transformed cells. Finally, we develop a Cdk2-AS mouse model that will allow for the acute inhibition of Cdk2 in a variety of cell types and cancer models. Together, these results enhance our understanding of the effects of Cdk inhibition in defined cellular and genetic contexts.
|Commitee:||Blelloch, Robert, Goga, Andrei, Morgan, David O., Oakes, Scott A.|
|School:||University of California, San Francisco|
|School Location:||United States -- California|
|Source:||DAI-B 74/08(E), Dissertation Abstracts International|
|Subjects:||Molecular biology, Cellular biology, Oncology|
|Keywords:||Analog sensitivity, Cancer, Cell cycle, Cyclin dependent kinase, Stem cells|
Copyright in each Dissertation and Thesis is retained by the author. All Rights Reserved
The supplemental file or files you are about to download were provided to ProQuest by the author as part of a
dissertation or thesis. The supplemental files are provided "AS IS" without warranty. ProQuest is not responsible for the
content, format or impact on the supplemental file(s) on our system. in some cases, the file type may be unknown or
may be a .exe file. We recommend caution as you open such files.
Copyright of the original materials contained in the supplemental file is retained by the author and your access to the
supplemental files is subject to the ProQuest Terms and Conditions of use.
Depending on the size of the file(s) you are downloading, the system may take some time to download them. Please be