GLUTAMINE AND BREAST CANCER PROLIFERATION AND TREATMENT
ABCOUWER STEVEN F
Awardee Organization: MASSACHUSETTS GENERAL HOSPITAL
Glutamine is an essential nutrient for the growth and maintenance of mammalian cells in culture, and tumor cells are avid glutamine consumers. However, little is known about the mechanism by which glutamine supports tumor cell proliferation and viability. This revised application will test the overall hypothesis that glutamine provides tumor cells with reductive equivalents as well as precursors for glutathione synthesis, a molecule which is essential for cellular protection against chemical and oxidative damage. Glutamine utilization may contribute to cellular redox metabolism and the mechanism by which cells respond to oxidative and xenobiotic stress. A series of human breast cell lines exhibiting a wide range of glutamine dependence for growth and viability, as well as a wide range of glutamine utilization rates, have been identified. In addition, the response of two highly glutamine-sensitive breast cell lines to glutamine starvation has been characterized and will be used to directly test a number of hypotheses: 1) Glutamine is-used to provide glutamate and cystine for synthesis of glutathione. 2) Glutamine utilization provides reductive equivalents to tumor cell mitochondria and therefore controls tumor cell growth and viability through mitochondrial redox mechanisms. 3) Targeted inhibition of glutaminase expression, the first enzyme involved in glutaminolysis, represses glutamine utilization and thereby inhibits growth and increases the sensitivity of tumor cells to chemotherapeutic drugs which are detoxified by glutathione. In general, methods to block the use of glutamine by tumor cells will be developed, and the efficacy of this strategy for inhibiting tumor growth or for increasing the sensitivity of tumors to chemotoxic drugs will be examined. This may provide the basis for novel metabolically directed cancer therapies.
Suppression by glutamate of proliferative activity through glutathione depletion mediated by the cystine/glutamate antiporter in mesenchymal C3H10T1/2 stem cells. 2007
Although previous studies including ours have demonstrated the functional expression of different glutamate (Glu) signaling machineries such as Glu receptors (GluRs) and transporters in osteoblasts and chondrocytes, little attention has been paid to the role of Glu in their ancestral mesenchymal stem cells to date. In the present study, we have evaluated the possible functionality of Glu in cultured mouse mesenchymal stem cell line C3H10T1/2 cells endowed to proliferate for the self-renewal and to differentiate toward osteoblast, chondrocyte, adipocyte, and myocyte lineages. Expression of mRNA was for the first time shown with the cystine/Glu antiporter composed of xCT and 4F2hc subunits, in addition to particular excitatory amino acid transporter (EAAT) isoforms and ionotropic GluRs, in undifferentiated C3H10T1/2 cells. Glu significantly suppressed the proliferation activity at a concentration over 500 microM without inducing cell death or differentiation, while the suppression occurred in a manner sensitive to the prevention by cystine and reduced glutathione (GSH), but not by EAAT inhibitors. A significant decrease was seen in intracellular GSH levels in C3H10T1/2 cells cultured with Glu, whereas the cellular proliferation activity was drastically decreased by the addition of the GSH depleter cyclohexene-1-one and the GSH biosynthesis inhibitor L-buthionine-[S,R]-sulfoximine, respectively. Transient overexpression of both xCT and 4F2hc subunits led to an increased basal proliferative activity in C3H10T1/2 cells. These results suggest that Glu could suppress the cellular proliferation toward self-renewal through a mechanism associated with the depletion of intracellular GSH after promoting the retrograde operation of the cystine/Glu antiporter in C3H10T1/2 cells.