The goal of this project is the discovery and development of selective small molecule inhibitors of 3- phosphoglycerate dehydrogenase (PHGDH), the first enzyme of the serine biosynthetic pathway. We have found that this pathway is upregulated in estrogen-receptor negative (ER-) breast cancers, which account for 20-25% of breast cancers but are responsible for 50% of breast cancer-related deaths. Breast cancer cells that overexpress PHGDH have higher flux through the serine biosynthesis pathway and are sensitive to knockdown of this enzyme, indicating that inhibitors of this pathway may be useful candidates for the treatment of ER- breast cancer. At present there are no small molecule inhibitors of PHGDH. We have developed a nicotinamide adenine dinucleotide (NADH)-linked high-throughput assay for PHGDH and carried out a pilot screen of 1400 known bioactive compounds, including FDA-approved pharmaceuticals, at the Laboratory for Drug Discovery in Neurodegeneration (LDDN) at the Brigham and Women's Hospital. This screen had a Z'factor of 0.61 and a coefficient of variation of 2%. The screen identified two compounds that dose-dependently inhibit PHGDH.
Our first aim i s to transfer this assay to an MLPCN center for a high- throughput screen of the MLPCN collection of over 350,000 small molecules.
Our second aim i s validation of these hits using a coupled assay of serine biosynthesis, amino acid and metabolic flux analysis to measure inhibition of serine and ?-ketoglutarate production by the serine biosynthesis pathway in vitro and in cells, and counterscreening of these hits against GAPDH to eliminate non-specific dehydrogenase inhibitors.
Our third aim i s assessment of the biological activity of these hits by determining their selective cytotoxicity towards a tumor cell line that overexpresses PHGDH and has high flux through the serine biosynthesis pathway, while sparing a tumor cell line that has low expression of PHGDH and has low serine biosynthesis pathway flux. Inhibitors of PHGDH will be tested in vivo in xenograft models of mouse tumors that overexpress PHGDH. Identification of specific inhibitors of PHGDH will permit the evaluation of serine biosynthesis inhibition as a novel therapeutic target for ER- breast cancer.
Estrogen receptor-negative (ER-) breast cancer accounts for 20-25% of all breast cancers but up to 50% of the 40,000 deaths each year from breast cancers. We have found that many of these cancers are dependent on serine synthesis for growth and survival. We will develop inhibitors of the first enzyme of serine synthesis in the hope of controlling ER- breast cancer. !
|Bar-Peled, Liron; Sabatini, David M (2014) Regulation of mTORC1 by amino acids. Trends Cell Biol 24:400-6|