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Supplementary Figure S1. Western blot characterization of antibodies against BCKDH-E1 and BCKDH-pSer-E1

Supplementary Figure S2. Brain Metastatic Cells Contain High Levels of Glycogen

Breast cancer brain metastasis is resistant to therapy and a particularly poor prognostic feature in patient survival. Altered metabolism is a common feature of cancer cells, but little is known as to what metabolic changes benefit breast cancer brain metastases. We found that brain metastatic breast cancer cells evolved the ability to survive and proliferate independent of glucose due to enhanced gluconeogenesis and oxidations of glutamine and branched chain amino acids, which together sustain the nonoxidative pentose pathway for purine synthesis. Silencing expression of fructose-1,6-bisphosphatases (FBP) in brain metastatic cells reduced their viability and improved the survival of metastasis-bearing immunocompetent hosts. Clinically, we showed that brain metastases from human breast cancer patients expressed higher levels of FBP and glycogen than the corresponding primary tumors. Together, our findings identify a critical metabolic condition required to sustain brain metastasis and suggest that targeting gluconeogenesis may help eradicate this deadly feature in advanced breast cancer patients. Cancer Res; 75(3); 554–65. ©2014 AACR.

Indoleamine 2,3-dioxygenase 1 (IDO1) is a potent immunosuppressive enzyme that inhibits the antitumor immune response through both tryptophan metabolism and non-enzymatic functions. To date, most IDO1-targeted approaches have focused on inhibiting tryptophan metabolism. However, this class of drugs has failed to improve the overall survival of patients with cancer. Here, we developed and characterized proteolysis targeting chimeras (PROTACs) that degrade the IDO1 protein. IDO1-PROTACs were tested for their effects on IDO1 enzyme and non-enzyme activities. After screening a library of IDO1-PROTAC derivatives, a compound was identified that potently degraded the IDO1 protein through cereblon-mediated proteasomal degradation. The IDO1-PROTAC: (i) inhibited IDO1 enzyme activity and IDO1-mediated NF-κB phosphorylation in cultured human glioblastoma (GBM) cells, (ii) degraded the IDO1 protein within intracranial brain tumors in vivo, and (iii) mediated a survival benefit in mice with well-established brain tumors. This study identified and characterized a new IDO1 protein degrader with therapeutic potential for patients with glioblastoma.

Overexpression of epidermal growth factor receptor (EGFR) is associated with poor prognosis in malignant tumors. Sodium/glucose co-transporter 1 (SGLT1) is an active glucose transporter that is overexpressed in many cancers including prostate cancer. Previously, we found that EGFR interacts with and stabilizes SGLT1 in cancer cells.In this study, we determined the micro-domain of EGFR that is required for its interaction with SGLT1 and the effects of activation/inactivation of EGFR on EGFR-SGLT1 interaction, measured the expression of EGFR and SGLT1 in prostate cancer tissues, and tested the effect of inhibition of SGLT1 on the sensitivity of prostate cancer cells to EGFR tyrosine inhibitors.We found that the autophosphorylation region (978-1210 amino acids) of EGFR was required for its sufficient interaction with SGLT1 and that this interaction was independent of EGFR's tyrosine kinase activity. Most importantly, the EGFR-SGLT1 interaction does not respond to EGFR tyrosine kinase modulators (EGF and tyrosine kinase inhibitors). EGFR and SGLT1 co-localized in prostate cancer tissues, and inhibition of SGLT1 by a SGLT1 inhibitor (Phlorizin) sensitized prostate cancer cells to EGFR inhibitors (Gefitinib and Erlotinib).These data suggest that EGFR in cancer cells can exist as either a tyrosine kinase modulator responsive status or an irresponsive status. SGLT1 is a protein involved in EGFR's functions that are irresponsive to EGFR tyrosine kinase inhibitors and, therefore, the EGFR-SGLT1 interaction might be a novel target for prostate cancer therapy.