Recently two distinct small molecule

Recently, two distinct small-molecule inhibitors of PHGDH were identified using high-throughput screens, both of which inhibit de novo serine biosynthesis and show selective toxicity to cancer Milnacipran HCl with high SSP flux [50,51]. The inhibitor NCT-503, which has an IC50 of 2.5μM, reduces the growth and weight of PHGDH-dependent breast tumor xenografts without affecting PHGDH-independent xenografts [50]. PHGDH inhibition suppresses production of labeled serine/glycine from [U-13C]glucose and incorporation of labeled one-carbon units derived from [U-13C]glucose into nucleotides (Fig. 3b). Total intratumoral serine concentration is unaffected by PHGDH inhibition because of influx of extracellular serine, consistent with the cell culture studies described above. Indeed, when cells are cultured in medium containing [U13C]serine, PHGDH inhibition results in an increased intracellular pool of exogenous (labeled) serine. However, PHGDH inhibition decreases the incorporation of one-carbon units from extracellular serine into nucleotides. Instead, cytosolic SHMT1 becomes activated and consumes 5,10-CH2-THF and glycine to generate serine at the expense of nucleotide synthesis (Fig. 3b). In the presence of the PHGDH inhibitor NCT-503, genetic deletion of SHMT1 permits the incorporation of exogenous serine-derived carbon units (via SHMT2 and the GCS) into nucleotide biosynthesis [50]. Thus, the development of PHGDH inhibitors has provided researchers with new tools to investigate PHGDH function in cancer cells but also represents an important step in the development of drugs that target the SSP.
Other potential targets One of the most overexpressed metabolic genes in human tumors is PYCR1, which encodes pyrroline-5-carboxylate reductase 1, a mitochondrial enzyme in the pathway that synthesizes proline from glutamate (Fig. 2) [46]. PYCR1 was identified as one of 16 essential metabolic genes in a breast cancer xenograft model [49], and its expression is upregulated by the oncogenic transcription factor c-Myc, which simultaneously downregulates enzymes that catalyze proline catabolism [52]. A recent study using ribosome profiling found that proline is the restrictive amino acid in kidney tumors and breast cancer xenografts (ribosomes were ‘stalled’ at proline codons) [53]. Thus, proline itself, rather than an intermediate or byproduct of the biosynthesis pathway, can be limiting for tumor growth. Although CRISPR/Cas9-mediated knockout of PYCR1 has no effect on breast cancer cell proliferation in monolayer culture, it completely abolishes tumor formation in vivo[53]. These data indicate that the proline biosynthesis pathway, and in particular PYCR1, could be targeted for cancer therapy. Finally, a distinct aspect of amino acid metabolism is being investigated as a target for cancer immunotherapy. The mutations that underlie cancer lead to expression of abnormal antigens by cancer cells but tumors acquire the ability to evade host immunosurveillance (tumor-induced tolerance) [54]. One of the mechanisms for tumor-induced tolerance involves indoleamine 2,3-dioxygenase (IDO), which is overexpressed by most human tumors and catalyzes the rate-limiting reaction for tryptophan catabolism via the kynurenine pathway [55]. Both isoforms of IDO (IDO1 and IDO2) are overexpressed in cancer but research to date has focused on IDO1. IDO activity limits the immune response by depleting tryptophan, which is essential for T cell proliferation, from the tumor microenvironment, and also by causing accumulation of the tryptophan metabolite kynurenine and its derivatives, which further inhibit immune cell proliferation [55]. Preclinical studies have shown that targeting IDO1 inhibits tumor growth and elicits an antitumor immune response in rodent models, and small-molecule inhibitors of IDO1 along with an IDO1-targeting vaccine are now being assessed in clinical trials [28,55]. Tryptophan 2,3-dioxygenase (TDO2) also catalyzes tryptophan degradation via the kynurenine pathway. Although TDO2 is normally present only in liver tissue and has a higher KM than IDO1 for tryptophan, it is also an immunosuppressive enzyme and is frequently upregulated in human tumors. Inhibition of TDO2 has shown promise in preclinical models [56] but is not currently being evaluated in clinical trials.