Article GSTP1 Is a Driver of Triple-Negative Breast Cancer Cell Metabolism and Pathogenicity Graphical Abstract Authors Sharon M. Louie, Elizabeth A. Grossman, Lisa A. Crawford, ..., Andrei Goga, Eranthie Weerapana, Daniel K. Nomura Correspondence [email protected] In Brief Using a reactivity-based chemoproteomic platform, Louie et al. have identified GSTP1 as a triple-negative breast cancer target that, when inhibited, impairs breast cancer pathogenicity through inhibiting GAPDH activity and downstream metabolism and signaling pathways. Highlights d We used chemoproteomics to profile metabolic drivers of breast cancer d GSTP1 is a novel triple-negative breast cancer-specific target d GSTP1 inhibition impairs triple-negative breast cancer pathogenicity d GSTP1 inhibition impairs GAPDH activity to affect metabolism and signaling Louie et al., 2016, Cell Chemical Biology 23, 1–12 May 19, 2016 ª 2016 Elsevier Ltd. http://dx.doi.org/10.1016/j.chembiol.2016.03.017 Please cite this article in press as: Louie et al., GSTP1 Is a Driver of Triple-Negative Breast Cancer Cell Metabolism and Pathogenicity, Cell Chemical Biology (2016), http://dx.doi.org/10.1016/j.chembiol.2016.03.017 Cell Chemical Biology Article GSTP1 Is a Driver of Triple-Negative Breast Cancer Cell Metabolism and Pathogenicity Sharon M. Louie,1 Elizabeth A. Grossman,1 Lisa A. Crawford,2 Lucky Ding,1 Roman Camarda,3 Tucker R. Huffman,1 David K. Miyamoto,1 Andrei Goga,3 Eranthie Weerapana,2 and Daniel K. Nomura1,* 1Departments of Chemistry and Nutritional Sciences and Toxicology, University of California, Berkeley, Berkeley, CA 94720, USA 2Department of Chemistry, Boston College, Chestnut Hill, MA 02467, USA 3Department of Cell and Tissue Biology and Medicine, University of California, San Francisco, San Francisco, CA 94143, USA *Correspondence: [email protected] http://dx.doi.org/10.1016/j.chembiol.2016.03.017 SUMMARY subtypes that are correlated with heightened malignancy and poor prognosis remain poorly understood. Breast cancers possess fundamentally altered meta- Mortality from breast cancer is almost always attributed to bolism that fuels their pathogenicity. While many metastatic spread of the disease to other organs, thus pre- metabolic drivers of breast cancers have been iden- cluding resection as a treatment method. Unfortunately, con- tified, the metabolic pathways that mediate breast ventional chemotherapy fails to eradicate most human cancers, cancer malignancy and poor prognosis are less including aggressive breast cancers. Studies over the past well understood. Here, we used a reactivity-based decade have uncovered certain breast cancer types and cell types that are associated with poor prognosis, such as chemoproteomic platform to profile metabolic en- estrogen/progesterone/HER2 receptor-negative (triple-nega- zymes that are enriched in breast cancer cell types tive) breast cancers (TNBCs) or cancer stem/precursor cells linked to poor prognosis, including triple-negative that possess self-renewing and tumor-initiating capabilities, breast cancer (TNBC) cells and breast cancer cells epithelial-to-mesenchymal transition (EMT), poor prognosis, that have undergone an epithelial-mesenchymal and chemotherapy resistance within breast tumors (Dawson transition-like state of heightened malignancy. We et al., 2009; Dietze et al., 2015; Polyak and Weinberg, 2009). identified glutathione S-transferase Pi 1 (GSTP1) as While eliminating these breast cancer types is critical in combat- a novel TNBC target that controls cancer pathoge- ting breast cancer, there are currently few to no therapies that nicity by regulating glycolytic and lipid metabolism, target this malignant population of breast cancer cells. energetics, and oncogenic signaling pathways In this study, we used a reactivity-based chemoproteomic plat- through a protein interaction that activates glyceral- form to identify metabolic enzymes that are heightened in TNBC cells or upon induction of an EMT-like programming of height- dehyde-3-phosphate dehydrogenase activity. We ened malignancy in breast cancer cells. Through this profiling show that genetic or pharmacological inactivation effort, we identified glutathione S-transferase Pi 1 (GSTP1) as a of GSTP1 impairs cell survival and tumorigenesis critical metabolic driver that is heightened specifically in TNBCs in TNBC cells. We put forth GSTP1 inhibitors as a to control multiple critical nodes in cancer metabolism and novel therapeutic strategy for combatting TNBCs signaling pathways to drive breast cancer pathogenicity. through impairing key cancer metabolism and signaling pathways. RESULTS Profiling Dysregulated Metabolic Enzymes in TNBC INTRODUCTION Cells and CDH1 Knockdown Breast Cancer Cells To identify metabolic drivers of breast cancer pathogenicity in Breast cancers possess fundamentally altered metabolism that aggressive breast cancer cell types associated with malignancy drives their pathogenic features. Since Otto Warburg’s seminal and poor prognosis, we used a reactivity-based chemical prote- discovery in the 1920s that cancer cells have heightened glucose omic strategy to map cysteine and lysine reactivity in TNBC cells uptake and aerobic glycolysis, recent studies have identified and breast cancer cells with EMT-like features (Figure 1; Table many other biochemical alterations in cancer cells, including S1). Both TNBC cells and breast cancer cells that have under- heightened glutamine-dependent anaplerosis and de novo lipid gone EMT have been linked to heightened aggressiveness and biosynthesis, which serve as metabolic platforms for breast poor prognosis. Specifically, we wanted to (1) identify TNBC- cancer cells to generate biomass for cell division and metabo- specific metabolic enzyme targets by comparing a panel of lites that modulate cancer cell signaling, epigenetics, and path- four non-TNBC and five TNBC cell lines, and (2) identify upregu- ogenicity (Benjamin et al., 2012; Cantor and Sabatini, 2012; lated enzyme targets in MCF7 breast cancer cells upon knock- Pavlova and Thompson, 2016). While targeting dysregulated down of CDH1, a critical mediator of EMT and cell-cell adhesion. metabolism is a promising strategy for breast cancer treatment, We knocked down CDH1 in MCF7 cells with short-hairpin (sh) the metabolic pathways that drive pathogenicity in breast cancer oligonucleotides (shCDH1 cells) to induce an EMT-like state. Cell Chemical Biology 23, 1–12, May 19, 2016 ª 2016 Elsevier Ltd. 1 Please cite this article in press as: Louie et al., GSTP1 Is a Driver of Triple-Negative Breast Cancer Cell Metabolism and Pathogenicity, Cell Chemical Biology (2016), http://dx.doi.org/10.1016/j.chembiol.2016.03.017 ABClysine reactivity in non-TNBC cysteine reactivity significantly upregulated metabolic enzyme and TNBC cells in MCF7 shCDH1 cells targets in TNBC cells 100 metabolic enzymes metabolic enzymes * s non-TNBC 80 non-TNBC TNBC TNBC 60 al count r 40 * ct pe shControl shCDH1 s ZR751 HCC70 231MFP MCF10A MCF7 T47D MDA-MB-361 HCC1143 HCC38 MDA-MB-468 20 * GSTP1 ALDH1A3 PSAT1 LDHB ASS1 ENO3 AGPS PFKM ACAA2 CTH *** SOD2 ACOT7 AGPS SAMHD1 PYGL ALDH1L2 MTHFD2 DLAT PLOD1 0 GNE NCEH1 ACSL5 PSAT1 CTPS LDLR PNPLA6 3 ACOT9 GLS ACP6 DPYD ALDH3A2 ALDH2 T1 PLOD3 PSPH PC SGPL1 ADH5 CBR1 GCAT GMDS AT2 PGM1 PLD3 ALDH1A2 AK2 ALDH1A1 CPS1 ASS1 DHCR7 TXNRD2 ENPP1 GYS1 PLAA GSTP1 TYMP GSTP1 ACSL PSA AC ABHD12 CBS GSTK1 PLCG1 OAS3 PNPT1 RDH11 ADSL PPAT BCKDK C11orf54 DPYSL2 GSTM1 CTPS LDHAL6B MGST3 PNP SOAT1 ACSS2 AGPAT9 AKR1B1 AKR1B10 ADSL AKR1C3 CA2 CPPED1 DEGS1 PFAS DHPS DHRS7 ETFDH D significantly upregulated metabolic enzyme targets FAR1 ACADSB GALE GCLM GGCX GNPTAB ACLY IDI1 LPCAT2 MCAT SCP2 ME2 MPI MTAP in MCF7 shCDH1 cells NMT1 GSPT1 80 P4HA1 P4HA2 PCYT2 PDPR ESD PIP4K2C PLCG2 UGCG MAOA PRPSAP2 GLS ACAT2 PC ALDH1B1 CKB shControl OAT ACSL4 ts * NAMPT PHGDH IDH3B PHGDH GSTO1 C21orf33 ALDH1B1 PGLS OGT 60 shCDH1 ACADM CBR1 ASNS NSDHL MTHFD1L ADK SORD PYGB ADSS SUCLG2 oun GART ALDH7A1 UGP2 PLOD2 HMGCL ACAA1 OAT ACSL3 PYGM PFKP DLAT ACAT1 ACOT8 SCP2 ECHS1 40 OXCT1 PAFAH1B1 LSS LDHB ral c ILVBL GLRX3 SDHA CMAS t SDHB ACAD9 SHMT2 PPA1 GLUD1 * HPRT1 * ENO2 PDHA1 SHMT2 DHCR24 DCI * FDPS PYCR2 MDH1 HSD17B4 MAT2A PGLS ECH1 20 SUCLG1 ENO1 NNT PCMT1 spec NDUFA9 * AGPAT1 * FLAD1 * * TALDO1 GOT2 OGDH HADHA * * * GPD2 * ACO2 PDHB * PGK1 ** DLD * * * MTHFD1 USP9X GFPT1 * * * * HK2 * GANAB ACADVL IMPDH2 LDHA ACLY 0 PDHA1 TKT MTHFD1L ALG2 GLO1 GOT2 IMPDH2 ACAT2 ACACA 2 B ALDH18A1 ACAT1 M L2HGDH CMAS PKLR PDXK CS ALDH18A1 K HK1 DH PTGES2 PCK2 MCCC1 GSTM3 GLS MDH2 C OAT GLUD2 PFKL PKM2 ALDH9A1 HIBADH DLAT HSDL2 LDHB PPATCTPS ADSLPFAS IDH3A PFKAGPS ASNS PGD PAPSS1 SORD HMGCS1 CCBL2 PSAT1 C12orf5 MDH1 ALDH GSTP1PLCG1 SOAT1 ACAT1 ESD GPX1 PHG GSTZ1 NDUFV1 ACAA2 LDH1A2 PAFAH1B3 UGGT1 PDHB ACADSB HADHB GMPS AHCY A ALDH1B1 ACSL1 ALDH7A1 GPI CKMT1A IDH1 GMPS NDUFS1 HMOX2 FH MVK ALDOA DHTKD1 MTHFD1 HADH PDXDC1 ACO1 GAPDH SDHA PGAM1 ACSF2 SACM1L GOT1 MDH2 GSTP1 lysine reactivity in TNBC cells MAT2B E NDUFS2 PFKL FAH GSTO1 LTA4H PFKM HSD17B10 OXCT1 SHMT1 ACOX1 250 ISYNA1 ts 100 IVD TKT * MPST s PCYOX1 CKMT1A ALDOC ASMTL ALDH2 NANS COMT ALDH5A1 HADHA BCKDHA NME1 IDH2 HSDL2 200 FDXR 80 FASN NQO1 SORD ACO2 ALDH9A1 G6PD ALDH16A1 coun GRHPR HSD17B10 SPTLC2 CKB DECR1 count CPT1A PRDX1 AKR1A1 150 PTPLAD1 l 60 ALDH6A1 ALDH5A1 PAPSS2 GSR UGDH GSTM3 ral PTPLAD1 a LPCAT1 COASY t BLVRA r MCCC2 TXNDC5 FBP1 t ABAT c ACOT2 100 ACOX3 GFPT1 40 AKR1C1 AKR7A2 c ALDH4A1 ALOX15B PTGES2 APRT BCKDHB BDH1 GAPDH pe BLMH CMBL COMTD1 DBT ECHS1 pe DCXR s 50 DERA DHRS2 20 DLST PGD DUT s EPHX1 FAHD1 FAHD2A ALDH7A1