Fundamentally, we examined the power of PHGDH inhibitors for the 4T1 cancers having IDH2-highest accounts
In view of one’s part out-of PHGDH and you can PSAT1 in mediating IDH2-centered metabolic remodeling, we investigated the fresh proteomic negative effects of this type of relationships. Healthy protein employed in kcalorie burning, translation devices, ribosome biogenesis, splicing, and you may telephone migration was basically upregulated because of the IDH2 and you can downregulated which have PHGDH and PSAT1 knockouts (Second Fig. S8A and you may S8B; Secondary Desk Ssix). Biggest metabolic healthy protein included the newest cytochrome family (CYCS, CYC1, CYB5R1), glutamine uptake and you will glutamate k-calorie burning (SLC1A5 and amor en linea dating apps you may GLUD1), solute provider transporters (SLC25A1 – CIC, citrate/malate transporter, SLC25A11 – OGC, alpha-ketoglutarate/malate transporter and you may SLC25A5 – ATP/ADP transporter), lipid kcalorie burning (SOAT1, TSPO, ACAD9), and glycolytic necessary protein (HK1 and you can PKM). I speculated one a decrease in the fresh new metabolic pastime through to PHGDH and you may PSAT1 knockout you’ll subscribe to new redox imbalance and sensitize the latest cells so you’re able to oxidative damage. S8C). Thus, PHGDH and you can PSAT1 enjoy a significant character from inside the providing anabolic present regarding nucleotides, lipids, and you will amino acids for the muscle with a high IDH2, and you may help mobile fret resistance (Secondary Fig. S8D).
In fact, the increased loss of PHGDH and you can PSAT1 created vulnerability in order to oxidative destroy while the phone survival was less than the control tissue (Additional Fig
Aiming to translate the SDL interaction to cancer therapy, we examined the sensitivity of IDH2-high cells to PHGDH inhibitors, in vitro and in vivo. Cells with stable IDH2 overexpression and IDH2 knockout were treated with PHGDH inhibitor (NCT-fifty2) for 48 hours in RPMI medium without serine and glycine. Initial metabolic analysis showed that PHGDH inhibition reduced serine (m3) and glycine (m2) labeling from 13 C6-glucose (Supplementary Fig. S8E-S8H). The dose range of NCT-502 was calibrated for each cell line (HCC38 and HCC1143), due to basal differences in cell line sensitivities. In agreement with the SDL prediction, HCC38 cells with IDH2 overexpression were more sensitive to NCT-502 treatment (IC50: 0.05 ?mol/L) compared with the control cells with low IDH2 expression (IC50: 0.18 ?mol/L; Fig. 7A). Control knockout HCC1143 cells with high basal IDH2 were more sensitive to NCT-502 (IC50: 0.5 ?mol/L) compared with the cells with IDH2 knockout (IC50: 2.2 ?mol/L; Fig. 7B). Next, we examined the efficacy of the PHGDH inhibitor in an in vivo murine model, 4T1 TN breast cancer cells, with high basal IDH2 and PHGDH expression. We knocked down IDH2 using stable shRNA constructs and the knockdown was confirmed by Western blotting (Supplementary Fig. S8I). 4T1 cells exhibited reduced cell proliferation and colony formation upon IDH2 knockdown (Fig. 7C and D). In addition, DMKG supplement to the murine 4T1 cells with IDH2 knockdown rescued the reduced cell proliferation and colony formation (Fig. 7C and D). 4T1 cells with high and low IDH2 expression were injected orthotopically to mammary glands of female mice and treated with the PHGDH inhibitor NCT-503 (Supplementary Fig. S8J), which is reported to have increased solubility in vivo (42). Analysis of tumor growth revealed that 4T1 tumors with high IDH2 showed enhanced tumor growth with larger tumor size and weight compared with the tumors with low IDH2 (Fig. 7E–G; Supplementary Fig. S8K). In addition, only the IDH2-high tumors treated with NCT-503 showed reduced tumor size and weight compared with the IDH2-high tumors treated with vehicle (Fig. 7E–G). IDH2-low tumors treated with either NCT-503 or vehicle were not affected by the treatment. Altogether, pharmacologic inhibition of serine biosynthesis using PHGDH inhibitor affects only the growth of IDH2-high cells. This in vivo validation demonstrated the SDL interaction between PHGDH and IDH2 and strengthened the metabolic alterations and the in vitro protumorigenic phenotypes. Our study emphasizes PHGDH inhibition as a promising therapeutic approach for TN breast tumors with high IDH2.