NASA Scientific and Technical Aerospace Reports

We have shown by transient expression of hBok that this member of the Bc1-2 pro-apoptotic family is unique since its
translocation to the nucleus is important for protein to induced apoptosis. Concern were raised since our observation did not
apply to endogenous Bok. We are now in a position to detect endogenous Bok by western Blot analysis and have shown that
endogenous Bok is present in the cytoplasm and nucleus of HeLa cells but only in the nucleus of MDA-MB-231 cells
suggesting that the intracellular localization of Bok is cell type specific. Bax is translocated from the cytoplasm to the
mitochondria upon the induction of apoptosis by the kinase inhibitor staurosporine. We therefore treated HeLa cells with
staurosporine to determine if Bok translocation from the cytoplasm to the nucleus is induced by staurosporine. We show that
unlik, Bax, cytoplasmic Bok is modified by a non- phosphorylation event prior to being translocated to the nucleus. We are
currently in the processes of confirming if this modification is required for the overall mechanism of Bok- induced apoptosis
or is a staurosporin dependent event. In addition, we hope to identify the type of modification observed in Bok and determine
its significance in Bok-induced apoptosis.
DTIC
Apoptosis; Cancer; Mammary Glands
20040111559 Virginia Mason Research Center, Seattle, WA
Development of a Transgenic Mouse Model for Breast Cancer that is Optimized for the Study of T Cell-Based
Therapeutic Strategies
Nelson, Brad H.; Jun. 2004; 11 pp.; In English
Contract(s)/Grant(s): DAMD17-00-1-0486
Report No.(s): AD-A425600; No Copyright; Avail: CASI; A03, Hardcopy
We sought to develop a transgenic mouse model for breast cancer that would allow the in viva activities of tumor- specific
T cell clones to be tracked through all stages of tumorigenesis and after various - immunotherapies. We ‘tagged’ the neu
oncogene with two defined T cell epitopes which conferred recognition by OT-l and OT-II T cell receptor (TOR) transgenic
T cells. When expressed as a transgenic in mammary epithelium, epitope-tagged neu (designated neu(subOTl/OT2) was
expected to induce mammary adenocarcinomas that express the epitope tags and hence are recognizable by OT-l and OT-II
T cells. We generated three neu(subOTl/OT2) transgene-positive founder lines, and expression of neu(subOTl/OT2) in
mammary epithelium was confirmed by Northern blot, western blot, and by immunological responses to the epitope tags. On
its own, neu(subOTl/OT2) induced tumors in a minority of mice and with a long latency (12-16 months). Fortunately, when
neu(subOTl/OT2) mice were crossed to mice expressing a mutant p53 transgene, tumors developed in the majority (718) of
mice with a reasonable latency (7-9.5 months). We are currently using this model to analyze the T cell response to spontaneous
mammary tumors and to develop novel immunotherapies for breast cancer. Thus, we fully accomplished the goals of this
project.
DTIC
Cancer; Cells (Biology); Lymphocytes; Mammary Glands; Mice; Therapy
20040111560 Scripps Research Inst., La Jolla, CA
The Effects of Deregulated Cyclin E Expression in Mitosis: A Role in Breast Tumorigenesis
Keck, Jamie M.; May 2004; 10 pp.; In English
Contract(s)/Grant(s): DAMD17-03-1-0383
Report No.(s): AD-A425602; No Copyright; Avail: CASI; A02, Hardcopy
The objective of this study is to identify the mechanism through which deregulated expression of cyclin E leads to
chromosome instability, first observed by Spruck et al. in rat and human cell lines. Cyclin E functions to regulate the timing
of S phase entry and centrosome duplication in cells. Cyclin E overexpression is an indicator of poor prognosis is cancer
patients; however the selective advantage gained by deregulating cyclin E has not yet been explained. We propose that in
addition to delaying S phase, cyclin E deregulation also causes a delay in mitosis. Therefore, cyclin E may be interfering with
mitotic division leading to chromosome instability and eventual tumorigenesis. In this first year of funding, I have
accomplished my goals to observe and define the proposed mitotic delay using flow cytometry, immunofluorescence, and live
cell microscopy. Cells were found delay in mitosis, specifically in prometaphase. In addition, we are now using biochemical
assays to analyze expression of important mitotic regulators and identify a possible substrate of cyclin E/Cdk2 phosphorylation
in mitosis.
DTIC
Cancer; Mammary Glands; Mitosis
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