Figure 3. PHGDH expression overcomes serine limitation in tumors growing in the mammary fat pad.
(A) Breast cancer cells from a breast tumor arising in a BRCAfl/fl; Trp53+/−; MMTV-Cre mouse with either a non-targeting sgRNA (sgctrl) or an sgRNA targeting PHGDH (sgPHGDH-1) were injected into the mammary fat pad of female NSG mice fed a control diet. Tumor size over time is shown. Tumor size at the final time point is significantly different based on an unpaired, two-tailed Welch’s t test (p = 0.0027). n=5 tumors per genotype. (B)-(C) Breast cancer cells from a breast tumor arising in BRCAfl/fl; Trp53+/−; MMTV-Cre mice with either a non-targeting sgRNA (sgctrl) (B) or an sgRNA targeting PHGDH (sgPHGDH-1) (C) were injected into the mammary fat pad of female NSG mice fed the indicated diets. Mice were fed the indicated diets starting the day that the cells were injected, and tumor size over time is shown. n=5 tumors per genotype. In (B), by comparing all three diet conditions, there was no significant correlation between increased dietary serine and increased tumor volume by an ANOVA test for trend (p=0.0770). There was no significant difference in final tumor volume between mice fed –ser/gly, control, or high serine diets based on unpaired, two-tailed Welch’s t tests for –ser/gly versus control (p=0.0986) and for control versus high serine diet (p=0.1217). In (C), by comparing all three diet conditions, there was a significant correlation between increased dietary serine and increased tumor volume by an ANOVA test for trend (p = 0.0022). There was a significant difference in final tumor volume between mice fed –ser/gly and control diets based on an unpaired, two-tailed Welch’s t test (p=0.0559). (D) Serine amount in control tumors (sgctrl) compared to PHGDH knockdown tumors (sgPHGDH-1). sgctrl tumors display a significant increase in serine concentration based on an unpaired, two-tailed Welch’s t test (p=0.032). n=4 tumors per genotype. (E) Glycine amount in control tumors (sgctrl) compared to PHGDH knockdown tumors (sgPHGDH-1). sgctrl tumors display a significant increase in glycine concentration based on an unpaired, two-tailed Welch’s t test (p=0.0332). n=4 tumors per genotype (F) Amount of α-ketoglutarate in control (sgctrl) and PHGDH knockdown (sgPHGDH-1) mammary fat pad orthotopic tumors in mice fed either a –ser/gly diet, a control diet, or a high serine diet as measured by LC/MS. No significant change in α-ketoglutarate levels were detected between sgctrl and sgPHGDH-1 tumors as determined by unpaired, two-tailed Welch’s t tests (p=0.5917, 0.6834, and 0.2908, respectively). (G) Amount of 2-hydroxyglutarate in control (sgctrl) and PHGDH knockdown (sgPHGDH-1) mammary fat pad orthotopic tumors in mice fed either a –ser/gly diet, a control diet, or a high serine diet as measured by LC/MS. No significant change in 2-hydroxyglutarate levels were detected between sgctrl and sgPHGDH-1 tumors as determined by unpaired, two-tailed Welch’s t tests (p=0.8387, 0.4603, and 0.8368, respectively). n=4 tumors per genotype.
Mean +/− SD is shown for all panels.