Aberrant FGFR Tyrosine Kinase Signaling Enhances the Warburg … · Metabolism and Chemical Biology...

Metabolism and Chemical Biology

Aberrant FGFR Tyrosine Kinase SignalingEnhances the Warburg Effect by ReprogrammingLDH Isoform Expression and Activity inProstate CancerJunchen Liu1,2, Guo Chen1,2, Zezhen Liu1,2, Shaoyou Liu1,2, Zhiduan Cai1,2,Pan You3, Yuepeng Ke2, Li Lai2, Yun Huang2, Hongchang Gao4,Liangcai Zhao4, Helene Pelicano5, Peng Huang5,Wallace L. McKeehan2,Chin-Lee Wu6, Cong Wang4,Weide Zhong1,7, and Fen Wang2

Abstract

The acquisition of ectopic fibroblast growthfactor receptor 1 (FGFR1) expression is well docu-mented in prostate cancer progression. How it con-tributes to prostate cancer progression is not fullyunderstood, although it is known to confer a growthadvantage andpromote cell survival.Here,we reportthat FGFR1 tyrosine kinase reprograms the energymetabolismof prostate cancer cells by regulating theexpression of lactate dehydrogenase (LDH) iso-zymes. FGFR1 increased LDHA stability throughtyrosine phosphorylation and reduced LDHBexpression by promoting its promoter methylation,thereby shifting cell metabolism from oxidativephosphorylation to aerobic glycolysis. LDHAdeple-tion compromised, whereas LDHB depletionenhanced the tumorigenicity of prostate cancercells. Furthermore, FGFR1 overexpression andaberrant LDH isozyme expression were associatedwith short overall survival and biochemical recur-rence times in patients with prostate cancer.Our results indicate that ectopic FGFR1 expressionreprograms the energy metabolism of prostatecancer cells, representing a hallmark change inprostate cancer progression.

Significance: FGF signaling drives theWarburg effect through differential regulation of LDHA and LDHB, thereby promotingthe progression of prostate cancer.

Graphical Abstract: http://cancerres.aacrjournals.org/content/canres/78/16/4459/F1.large.jpg. Cancer Res; 78(16); 4459–70.�2018 AACR.

© 2018 American Association for Cancer Research

suppressing LDHB expression in prostate cancer.

– FGFR1 signaling

HSP

G

Glucose

O2

CO2

Pyruvate

LDHBLDHA

Lactate

+ FGFR1 signaling

HSP

G

Glucose

O2

CO2

Pyruvate

LDHBLDHA

Lactate

P P

P P

1Department of Urology, Guangdong Key Laboratory of Clinical MolecularMedicine and Diagnostics, the Second Affiliated Hospital of South China Uni-versity of Technology, Guangzhou, China. 2Institute of Biosciences and Tech-nology, College of Medicine, Texas A&M University, Houston, Texas. 3XianyueHospital, Xiamen, China. 4Wenzhou Medical University, Wenzhou, China.5Departments of Translational Molecular Pathology, MD Anderson CancerCenter, Houston, Texas. 6Departments of PathologyandUrology,MassachusettsGeneral Hospital and Harvard Medical School, Boston, Massachusetts. 7Depart-ment of Urology, Guangzhou Medical University, Guangzhou, China.

Note: Supplementary data for this article are available at Cancer ResearchOnline (http://cancerres.aacrjournals.org/).

J. Liu, G. Chen, and Z. Liu contributed equally to this article.

Corresponding Authors: Fen Wang, Institute of Biosciences and Technol-ogy, Texas A&M Health Science Center, Houston, TX 77030-3303. Phone:713-677-7522; Fax: 713-677-7512; E-mail: [email protected]; WeideZhong, Department of Urology, Guangdong Key Laboratory of ClinicalMolecular Medicine and Diagnostics, Guangzhou First People's Hospital,the Second Affiliated Hospital of South China University of Technology,Guangzhou 510180, China. E-mail: [email protected]; and Cong Wang,Wenzhou Medical University, Wenzhou, Zhejiang, China, E-mail:[email protected]

doi: 10.1158/0008-5472.CAN-17-3226

�2018 American Association for Cancer Research.

CancerResearch

www.aacrjournals.org 4459

on June 22, 2021. © 2018 American Association for Cancer Research. cancerres.aacrjournals.org Downloaded from

Published OnlineFirst June 11, 2018; DOI: 10.1158/0008-5472.CAN-17-3226

http://crossmark.crossref.org/dialog/?doi=10.1158/0008-5472.CAN-17-3226&domain=pdf&date_stamp=2018-7-28http://cancerres.aacrjournals.org/content/canres/78/16/4459/F1.large.jpghttp://cancerres.aacrjournals.org/

IntroductionMetabolic reprograming from oxidative phosphorylation to

aerobic glycolysis is a common event in cancer progression.Although glycolysis is less efficient than oxidative phosphoryla-tion for providing energy with respect to the number of ATP perglucose, it meets the demand of rapidly growing cancer cells forbuilding blocks. In addition, increased glycolysis results in glu-cose deprivation and lactate accumulation in the tumor micro-environment, which suppresses lymphocyte infiltration and com-promises anti-immunotherapies (1, 2). In line with those effects,the glycolytic phenotype is associated with prostate cancer pro-gression and aggressiveness (3–10). An understanding of howtumor cells reprogram their metabolism from oxidative phos-phorylation to aerobic glycolysismay provide a novel approach toselectively inhibit aerobic glycolysis in tumor cells.

The prostate consists of epithelial and stromal compartments,which maintain active two-way communication through para-crine mechanisms, including fibroblast growth factor (FGF) sig-naling in which FGF and FGF receptor (FGFR) are partitionedbetween the two compartments (11). That precisely balancedcommunication is critical for preserving the tissue homeostasisand function of the prostate. The FGF family consists of 18 ligandsthat exert their regulatory signals by activating the FGFR tyrosinekinases encoded by 4 homologous genes. FGF and FGFR areexpressed throughout the body in a pattern that is spatiotempo-rally and cell-type specific, controlling embryonic developmentand maintaining adult tissue homeostasis and function.

There is extensive evidence that ectopic activation of theFGF/FGFR signaling axis is associated with prostate cancer devel-opment and progression (11). The acquisition of ectopic FGFR1expression stands out as the most remarkable change among theFGFR isotypes in prostate cancer. The forced expressions of FGFand FGFR have been shown to induce prostate lesions in mousemodels. On the other hand, the ablation of Fgfr1 or Frs2a, whichencodes FGFR-substrate 2a (FRS2a), an adaptor protein neededfor FGFR kinases to activate ERK and PI3K/AKT pathways, signif-icantly reduced prostate cancer development and progression inmice (12, 13).

FGF signaling promotes aerobic glycolysis by increasing hexo-kinase 2 (HK2) expression and the tyrosine phosphorylation ofmultiple enzymes involved in aerobic glycolysis (14). Whetherand how ectopic FGF signaling contributes to prostate cancerprogression by promoting aerobic glycolysis, remains to be deter-mined. The last step of glycolysis is the reduction of pyruvate tolactate, a reversible conversion catalyzedby lactate dehydrogenase(LDH). LDH is a tetrameric enzyme composed of two types ofsubunits, LDHA and LDHB. The combination of the two subunitsyields five isozymes, which catalyze the conversion betweenpyruvate and lactate. LDH1 is composed of four LDHB subunitsand favors the conversion from lactate to pyruvate, allowingoxidation along the pathway of the tricarboxylic acid cycle. LDH5is composed of four LDHA subunits and favors the conversionfrom pyruvate to lactate, allowing the glycolytic pathway to becompleted at the formation of lactate (15). Interestingly, hypoxiainduces LDHA, while inhibits LDHB, expression (16, 17). Emerg-ing evidence shows that LDHA is required for the survival andproliferation of cancer cells. Although still a matter of debate,current data seem to indicate an association between reducedlevels of LDHB and increased malignancy in prostate cancer andother cancers (18–27).

LDHA is tyrosine phosphorylated in cancer cells (28). FGFR1has been reported to phosphorylate LDHA at multiple tyrosineresidues, which enhances tetramer formation andNADHbindingand thus the enzymatic activity of LDHA (29). In this study, wedemonstrate that FGFR1 signaling promotes aerobic glycolysis bystabilizing LDHA through tyrosine phosphorylation and down-regulating LDHB expression through the induction of hyper-methylation in the Ldhb promoter. We found that LDHA ablationcompromised, whereas LDHB ablation enhanced, the tumorige-nicity ofDU145 cells. Furthermore, high levels of phosphorylatedLDHA and low levels of LDHB in human prostate cancer tissueswere associated with short biochemical recurrence and survivaltimes in patients with prostate cancer. Together, our resultssuggest that ectopically expressed FGFR1 in prostate cancerinduces metabolic changes by increasing LDHA levels and low-ering LDHB levels, which promotes prostate cancer growth andprogression.

Materials and MethodsAnimals

Mice were housed under the Program of Animal Resources ofthe Institute of Biosciences and Technology in accordance withthe principles and procedures of the Guide for the Care andUse of Laboratory Animals. All experimental procedures wereapproved by the Institutional Animal Care and Use Committee.Mice were bred and genotyped as described previously(12, 30, 31). For xenograft studies, 2 � 106 DU145 cells weremixed with Matrigel (BD Biosciences) at a 1:1 ratio and subcu-taneously injected into the flanks of nude mice (6-week old).The size of xenograft was measured with a caliper and calculatedasV¼ 0.52� length�width2. The xenografts were harvested afterthe animals were euthanized via CO2 suffocation.

HistologyProstate tissues and xenografts were fixed, embedded, sec-

tioned as described (32). For immunostaining, the antigens wereretrieved by boiling in citrate buffer (10 mmol/L, pH 8.0) for 20minutes. Rabbit antiphosphorylated LDHA (pLDHA, 1:200)and anti-LDHA (1:200) were purchased from Cell SignalingTechnology. Mouse anti-LDHB (1:200), rabbit anti-CD31(1:200) and anti-Ki67 (1:200), and rat anti-F4/80 (1:200) werepurchased from Abcam. A Fluoremetric TUNEL Assay Kit waspurchased from Premega Co. The ExtraAvidin Peroxidase System(Sigma Aldrich) and fluorescence-conjugated secondary antibo-dies (Invitrogen) were used to visualize specifically bound anti-bodies. For immunofluorescence staining, the nuclei were coun-terstained with To-Pro 3 before being observed under a confocalmicroscope (Zeiss LSM 510).

The Massachusetts General Hospital (Boston, MA) prostatecancer tissue microarray (TMA) was used to assess the expres-sion of LDHA, LDHB, and Fgfr1 as well as the level of phos-phorylated LDHA in human prostate cancer samples asdescribed previously (33). Immunostaining of prostate cancercells and that of stromal cells was evaluated separately. Thepercentage of positive cells was calculated and categorized asfollows: 0, 0%; 1, 1%–10%; 2, 11%–50%; 3, 50%–75%; and 4,75%–100%. The staining intensity was visually scored anddefined as follows: 0, negative; 1, weak; 2, moderate; and 3,strong. Final immunoreactivity scores (IRS) were calculated foreach case by multiplying the percentage and intensity scores.

Liu et al.

Cancer Res; 78(16) August 15, 2018 Cancer Research4460



http://cancerres.aacrjournals.org/

Western blottingCells or xenografts were homogenized in RIPA buffer as

described previously (32). Samples containing 30 mg proteinwereseparated by SDS-PAGE and blotted onto polyvinylidene difluor-ide membranes. The antiphosphorylated ERK1/2 (1:1,000), anti-phosphorylated AKT (1:1,000), anti-ERK1/2 (1:1,000), anti-AKT(1:1,000), antiphosphorylated FRS2a (1:1,000), and anti-HA(1:1,000) antibodies were purchased from Santa Cruz Biotech-nology. Anti-pLDHA (1:1,000), anti-LDHB (1:1,000), and anti-LDHA 1:1,000) antibodies and the Glycolysis Antibody SamplerKit containing antibodies against HK1, PFKP, PFKP2, PFKP3,aldolase, PGAM, PKM1/2, and pyruvate dehydrogenase werepurchased from Cell Signaling Technology. The antiphosphotyr-osine 4G10 (1:1,000), anti-TET1 (1:1,000), and anti-Flag(1:1,000) antibodies were purchased from Millipore Sigma. Thespecifically bound antibodies were visualized using the ECL-Pluschemiluminescent reagents. The films were scanned with a den-sitometer for quantitation.

RNA expressionTotal RNA was isolated from cells and tissues using the Ribo-

pure RNA isolation reagent (Ambion), reverse transcripted withSuperScript III (Life Technologies) and random primers, andanalyzed with real-time PCR using the Fast SYBR Green MasterMix (Life Technologies) as described. The results were expressed asthe mean� SD as described (32). The primer sequences are listedin Supplementary Table S1.

Gene ablationThe lentivirus-based CRISPR-Cas9 system was used to ablate

the Ldha, Ldhb, and Fgfr1 alleles in DU145 cells. The sequences ofthe sgRNAs are shown in Supplementary Table S1. Two days afterinfection with the lentivirus, the recombinant cells were selectedvia growth in a medium containing 2 mg/mL puromycin.

Site-directed mutagenesisThe QuikChange Lightning Multi Site-Directed Mutagenesis

Kit (Agilent Technologies) was used to generate LDHA-mutantcDNAs. The primer sequences are listed in SupplementaryTable S1.

NMR analysesCells (2 � 107) were suspended in 450 mL methanol/chloro-

form (2:1) and lysed by ultrasound. The lysates were mixed with450 mL chloroform/H2O. The supernatants were lyophilized andthen resuspended in 500-mL D2O containing 0.25 mmol/L sodi-um trimethylsily 1 propionate-d4. All NMR spectra were recordedon a Bruker AVANCE III 600 MHz NMR spectrometer.

Methylated DNA immunoprecipitationCells were isolated with the Qiagen Kit and ultrasound shear-

ed into smaller fragments (200–600 bp). The sheared gDNAwas denatured by incubation in 1 mol/L NaOH/25 mmol/LEDTA at 95�C for 12 minutes, and then immunoprecipitatedwith anti-5-methylated cytosine (5mC) antibodies. Thebound 5mC-containing DNA was then purified and used tomake the sequencing library with the NEB (E6240S) LibraryPrep Kit, which was subsequently sequenced by the AgrelifeGenomics and Bioinformatics Service, Texas A&M University(College Station, TX).

Sodium bisulfite DNA sequenceGenomic DNA was isolated using the Qiangen Genomic DNA

Kit, followed by bisulfite conversion using the EpiJET BisulfiteConversion Kit (Thermo Scientific). The bisulfite-specific primersfor the PCR amplification were listed in Supplementary Table S1.The PCR products were cloned into pDrive Cloning Vectors(Qiangen) and sequenced.

Protein stability assayStable MEF transfectants expressing HA-tagged LDHA were

treated with cycloheximide for the indicated times. The abun-dance of LDHA was examined by Western blot analysis. Therelative level of endogenous or HA-tagged LDHAwas quantitatedusing ImageJ. The GraphPad software was used to compare theslopes of each curve.

Statistical analysisStatistical analysiswas performedusing the two-tailed t test. For

protein stability assay, the GraphPad software was used to com-pare two curves. P < 0.05 is considered statistical significant. Errorbars indicate SDs.

ResultsAblation of the FGF signaling axis reduces glycolysis andincreases oxygen consumption in MEF

We first determined whether FGF signaling regulated cellmetabolism in mouse embryonic fibroblasts (MEF), whichexpressed FGFR1, FGFR2, and FRS2a. We generated MEFs thatwere devoid of Fgfr1, Fgfr2, and Frs2a by infecting MEFs bearingfloxed Fgfr1, Fgfr2, and Frs2a alleles with adenovirus carrying theCre-GFP coding sequence. The cells, designated MEFDF, did notexpress Fgfr1, Fgfr2, and Frs2amRNA (Fig. 1A). Interestingly, real-time RT-PCR also revealed that at the mRNA level, MEFDF

increased Ldhb, but not Ldha expression. However, Western blotanalyses revealed that at the protein level, comparedwith parentalMEFs, the MEFDF cells had reduced LDHA expression andincreased LDHB expression (Fig. 1B). Those results suggest thatFGF signaling promotes LDHA expression at the protein level andsuppresses LDHB expression at the mRNA level. Western blotanalysis also showed that FGF2 failed to induce phosphorylationof FRS2a, ERK, and AKT, indicating successful abrogation of FGFsignaling in the cells.

LDHA and LDHB favor opposite direction of the conversa-tion between pyruvate and lactate (15). To determine whetherabrogation of the FGF signaling axis in MEFs changes the cellmetabolism, we compared the lactate production and oxygenconsumption in MEFs and MEFDF cells (Fig. 1C). The abro-gation of FGF signaling reduced lactate production andincreased the oxygen consumption rate. In contrast, overexpres-sion of LDHA in MEFDF cells significantly increased lactateproduction (Supplementary Fig. S1). In addition, NMR anal-yses revealed that the abrogation of FGF signaling reduced theconcentrations of isoleucine, valine, lactate, and acetateand increased the concentrations of glutamate and succinate(Fig. 1D). The reduced isoleucine and valine levels in MEFDF

cells suggest that the synthesis of branched chain amino acids(BCAA) was compromised. The dysregulation of BCAA synthe-sis in MEFs lacking FGF signaling is in line with evidence thatBCAAs promote glucose uptake (34). Those results demonstratea shift in energy metabolism from aerobic glycolysis to

FGF Signaling in Cell Energy Metabolism

www.aacrjournals.org Cancer Res; 78(16) August 15, 2018 4461




oxidative phosphorylation in MEFDF cells, suggesting that ener-gy metabolism in MEFs is regulated by FGF signaling.

FGFR1 enhances the stability of LDHA via tyrosinephosphorylation

To determine the role of FGFR1 in the regulation of LDHA atthe protein level, we treated the MEFs and MEFDF cells withcycloheximine to block protein synthesis. Western blot analysesrevealed that the abundance of LDHA declined faster in MEFDF

cells than in the parental MEFs (Fig. 2A). Those results indicatethat the half-life of LDHA is shorter in MEFDF cells than in theparental MEFs, suggesting that FGF signaling enhances thestability of LDHA.

LDHA consists of four tyrosine phosphorylation sites(Fig. 2B). In line with a previous report that FGFR1 directlyphosphorylates LDHA (29), Western blot analyses with theantiphosphotyrosine antibody 4G10 showed that LDHA wastyrosine phosphorylated (Fig. 2C). To identify which tyrosinephosphorylation sites were involved in the regulation of LDHAstability, we employed site-directed mutagenesis to generateLDHAmutants with individual, double (2F), or quadruple (4F)mutations. The individual mutations at each of the four tyro-sine-phosphorylation sites did not affect the phosporylationLDHA. Note that weak expression of FGFR1 in the Y83F groupmight account for the relatively low phosphorylation of Y83F.Only the 4F mutant failed to be phosphorylated by FGFR1. Theother mutants displayed only partially reduced phosphoryla-tion, comfirming the previous report that all four tyrosineresidues are phosphorylated by FGFR1.

Western blot analyses revealed that the individual mutationsat each of the four tyrosine phosphorylation sites and the two2F mutants did not affect the stablility of LDHA (P > 0.05).

However, the 4F mutant showed a statistically significantreduction in stabilitity (Fig. 2D). Together, those results indi-cate that phosphorylation of the four tyrosine residues pro-motes the stability of LDHA.

FGFR1 suppresses LDHB transcription by promoting DNAmethylation in the LDHB promoter

The Ldhb promoter is heavily methylated in prostate cancer,which inhibits the transcription of LDHB (22). To determinewhether FGF signaling suppresses LDHB expression via pro-moter methylation, we employed methylated DNA immuno-precipitation to pull down the methylated DNA for high-throughput sequencing (Fig. 3A). The results showed that theCpG islands in the Ldhb promoter were less methylated inMEFDF cells than in the parental MEFs. Further bisulfitesequencing of the Ldhb promoter area confirmed that DNAdemethylation was reduced in the MEFDF cells (Fig. 3B). Con-sistent with the data that ablation of FGF signaling did notaffect LDHA mRNA expression, no differences were observed inldha promoter methylation between MEF and MEFDF cells(Supplementary Fig. S2).

Because DNA demethylation is catalyzed by the three TETenzymes (TET1–3), we compared the expression levels ofTET1–3 in MEFDF cells and parental MEFs. We found that theexpression of Tet1 at both mRNA and protein levels wassignificantly increased in the MEFDF cells (Fig. 3C and D),suggesting that FGF signaling suppressed Tet1 expression. Con-sistantly, Ldhb, but not ldha, expression was higher in MEFsbearing Tet1null alleles than in parental MEFs (Fig. 3E), furtherindicating that the expression of LDHB was reduced by DNAmethylation.

Figure 1.

Ablation of FGF signaling suppresses aerobic glycolysis and promotes oxidative phosphorylation in MEFs.A, Real-time RT-PCR analyses of the indicated mRNAs. B,Western blot analysis of the indicated proteins. C, Relative oxygen consumption and lactate production. D, NMR analyses demonstrated metabolite changesin MEFDF cells. Each column represents an individual sample. WT, wild-type; Ctrl, control; DF, MEFDF. � , P < 0.05.

Liu et al.





Ablation of the FGF signaling axis reprograms cell metabolismin human prostate cancer cells

DU145 cells highly expressed Fgfr1 (Fig. 4A). To investigatewhether ectopic FGF signaling contributes to metabolic repro-

gramming in prostate cancer, we used CRISPR/Cas9 geneediting to ablate the Fgfr1 alleles in DU145 cells that expressedhigh levels FGFR1. Although it did not affect cell growth inthe medium with 10% FBS, ablation of Fgfr1 compromised cell

Figure 2.

Tyrosine-phosphorylation suppresses thedegradation of LDHA. A, The cells were treated withcycloheximide (CHX) for the indicated times.Endogenous LDHA proteins were determined byWestern blot analysis. The intensity of LDHA relativeto that of b-actin is shown on the right. B, Thestructural domains of LDHA. Green dots, tyrosinephosphorylation sites. C, HA-tagged LDHA wasexpressed in 239T cells with FGFR1. PhosphorylatedLDHA was detected with antiphosphotyrosineantibody 4G10. D, The cells were treated withcycloheximide for the indicated times. The levels ofLDHA were determined by Western blot analysis.The intensity of LDHA relative to that of b-actin isshown on the right. T, tetramer formation domain;NADP, NAD/NADP binding domain; C, C-terminaldomain; 2Fa, Y10F/Y83F mutation; 2Fb, Y172/Y239mutation; 4F, Y10F/Y83F/Y172/Y239 mutation; IP,immunoprecipitation; IB, immunoblot. � , P < 0.01.

Figure 3.

Ablation of FGF signaling reduces DNA methylation in the Ldhb promoter. A, Methylated DNA was immunoprecipitated and subjected to high-throughputsequencing. The level of methylation at CpG islands in the Ldhb promoter region is shown. B, Bisulfite DNA sequencing of the Ldhb promoter area inMEFDF cells. C and D, Real-time RT-PCR and Western blot analyses showing increased Tet1 in MEFDF cells at the mRNA and protein levels. E, Real-time RT-PCRanalyses of ldha and Ldhb in Tet1 null MEF (DTet1). � , P < 0.05.






growth in the medium with 1% serum (Supplementary Fig. S3).In line with the data from MEFs, the expression of Ldha mRNAwas not affected in the Fgfr1null DU145 (DU145DR1) cells,whereas that of Ldhb mRNA was increased (Fig. 4B). We thendetermined the expression profiles of the LDH isozymes in theparental and DU145DR1 cells by Western blot analysis (Fig. 4C).The ablation of Fgfr1 reduced LDHA expression and increasedLDHB expression at the protein level. Interestingly, expressionof FGFR1 protein was increased by FGF2 treatment. However,the underlying mechanism remains to be determined. Togetherwith Fig. 4B, the results further indicate that FGFR1 regulatesLDHA at either the translational or posttranslational level andLDHB at the transcriptional level. Consistent with the reportthat FGFR1 phosphorylates LDHA, Western blot analysesrevealed that the phosphorylation of LDHA at the Y10 residuewas reduced in the DU145DR1 cells. Consistently, the ablationof Fgfr1 reduced glucose uptake and lactate production andincreased O2 consumption (Fig. 4D). Similarly, treating DU145cells with FGFR inhibitor AZD4547 also suppressed glycolysis(Supplementary Fig. S4). Together, the data suggest that theablation of Fgfr1 in prostate cancer cells reprograms the cellenergy metabolism from oxidative phosphorylation to aerobicglycolysis.

LDHA ablation reduces, whereas LDHB ablation enhances, thetumorigenesis of DU145 cells

To investigate the functions of the LDH isoforms in the tumor-igenic activity of prostate cancer cells, we employed CRISPR/Cas9to delete the Ldha or Ldhb alleles in DU145 cells, designatedDU145DLdha andDU145DLdhb, respectively. Western blot analysesof the xenografts revealed that the ablation of Ldha increasedPGAM levels, while the ablationof Ldhb increasedHK1, PFK3, andaldolase levels (Fig. 5A). Those results indicate that LDHA deple-

tion compromised, whereas LDHB depletion enhanced, glycoly-sis inDU145 cells. Althoughnodifferencewas observed in growthrates (Fig. 5B), when grafted subcutaneously into the flanks ofnudemice, DU145DLdha cells generated smaller tumors comparedwith parental DU145 cells (Fig. 5C). In contrast, grafts ofDU145DLdhb cells generated larger tumors than those of parentalDU145 cells. Consistantly, similar results were derived from PC3cells (Supplementary Fig. S5).

LDHA deletion and LDHB deletion both stimulated com-pensatory upregulation of glycolytic-related proteins but hadopposite effects on tumor growth, suggesting that there mightbe unidentified proteins that mediate LDH-regulated tumorgrowth. Noticebly, ablation of Ldha increased expression ofLDHB and vice versa. Whether these changes are because offeedback regulation to compensate the loss of LDH remains tobe investigated.

Although there were no significant differences in tissuehistology among the parental DU145, DU145DLdha, andDU145DLdhb xenografts, the DU145DLdha xenografts had less,and the DU145Dldhb xenografts had more, Ki67þ cells than theparental DU145 xenografts, indicating that the ablation of Ldhareduced, and the ablation of Ldhb increased, the number ofproliferating cells in the xenografts (Fig. 6A and B). Further-more, there were more apoptotic cells in the DU145DLdha

xenografts than in the parental DU145 xenografts, whereas theopposite relation was observed between the DU145DLdhb xeno-grafts and the parental DU145 xenografts (Fig. 6C). To inves-tigate the impact of Ldha or Ldhb ablation on the tumormicroenvironment, we stained DU145DLdha, DU145DLdhb, andparental DU145 xenografts with anti-CD31 antibodies andanti-F4/80 antibodies to examine the densities of microvesselsand macrophages, respectively. There were fewer CD31þ andF4/80þ cells in the DU145DLdha xenografts than in the parental

Figure 4.

Ablation of FGF signaling suppresses aerobic glycolysis and promotes oxidative phosphorylation in DU145 cells. A and B, Real-time RT-PCR analyses ofFGFR and LDH isoform expression. C, Western blot analysis of LDHA and LDHB expression. D, Comparison of glucose uptake, oxygen consumption,and lactate and ATP production. Ctrl, control; DR1, DU145DR1; pLDHA, phosphorylated LDHA; pERK, phosphorylated ERK1/2; pAKT, phosphorylated AKT;b-actin was used as a loading control. � , P < 0.05.

Liu et al.





DU145 xenografts. In contrast, there were more CD31þ and F4/80þ cells in the DU145DLdhb xenografts than in the parentalDU145 xenografts. Those results demonstrated that Ldha abla-tion compromised, whereas Ldhb ablation stimulated, angio-genesis and macrophage infiltration and thus the tumorigenicactivity of DU145 cells.

Hyperphosphorylation of LDHA and reduced LDHBexpression levels in human prostate cancer

We performed IHC staining to determine the expression pat-terns of LDHA and LDHB in a human prostate TMA thatcomprised 225 prostate cancer samples and 27 benign prostatesamples (33). The results suggested that LDHA levels were higherin prostate cancer tissues than in adjacent prostate tissues(Fig. 7A). FGFR1 can phosphorylate human LDHA at multipletyrosine residues (29). Compared with noncancerous tissues,prostate cancer tissues had higher levels of Y10-phosphorylatedLDHA (pLDHA) and lower levels of LDHB (Fig. 7B).

The samples in the TMA were annotated with detailedinformation based on a 15-year follow-up of the patients,including PSA recurrence, Gleason Scores, pathologic stages,patients' age, and survival time. Therefore, we separated thesamples into two groups based on the median level. Higherthan the median is defined as High and lower than the medianis defined as Low. As shown in Fig. 7C and D, the clinicaloutcomes of the patients with high pLDHA levels (N ¼ 153)were worse than those of the patients with low pLDHA levels

(N ¼ 48), while those of patients with low LDHB levels(N ¼ 126) were worse than those of patients with high LDHBlevels (N ¼ 96); the patients with high pLDHA levels andlow LDHB levels (N ¼ 13) clearly had shorter biochemicalrecurrence-free times than those with low pLDHA levels andhigh LDHB levels (N ¼ 86). Those results suggest that pLDHAand LDHB expression have the potential to serve as bio-markers for prostate cancer prognosis.

Fgfr1 is overexpressed in about 40% of human prostatecancers (35, 36). To determine whether the expression level ofFgfr1 correlates with the abundance of pLDHA, we used in situhybridization to assess the expression levels of Fgfr1 in the TMA.Fgfr1 expressed was higher in prostate cancer than in benigntissues (Fig. 7D). The expression was associated with shortPSA-free survival time (Fig. 7E). Furthermore, the Fgfr1 expres-sion level was positively associated with the level of pLDHA(Fig. 7F). Together, the data imply that ectopically expressedFgfr1 in prostate cancer deregulates the expression of LDHisozymes and thus changes the glycolysis and metabolism ofthe cells (Fig. 7G).

DiscussionThere is extensive evidence that ectopic FGF/FGFR1 signaling

is a contributing factor in prostate cancer development andprogression (11, 36–41), that FGF-mediated glycolysis playsa pivotal role in development (14, 42), and that FGFR1 directly

Figure 5.

LDHA ablation reduces, and LDHB ablationenhances, the tumorigenicity of DU145 cells. A,Western blot analysis of the expression of enzymesrelated to aerobic glycolysis. B, The indicatedDU145 cells (2 � 104) were plated on 6-cm dishes.Cell numbers were counted every other day.C, Xenografts derived from the indicated controlDU145 cells. Note that the DLdha and DLdhb tumorswere harvested at different days, because theDLdhb tumors reached the limit of tumor burdenearlier than the DLdha tumors. The averagexenograft weight was calculated from all individualxenografts and is presented on the right. DLDHA,DU145DLdha; DLDHB, DU145DLdhb; Ctrl, control DU145cells; HK1, hexokinase 1; PD, pyruvatedehydrogenase. � , P < 0.05.






phosphorylates LDHA and enhances its enzymatic activity(29). Herein, we showed that FGFR1 signaling promotes aer-obic glycolysis by upregulating LDHA at the protein level and

downregulating LDHB at the transcriptional level. The ablationof LDHA compromised, whereas that of LDHB enhanced, thetumorigenic activity of DU145 prostate cancer cells.

Figure 6.

Differential impacts of LDHA and LDHB on cell survival, angiogenesis, and inflammation in prostate cancer xenografts. A and B, Hematoxylin and eosin (H&E)and IHC staining. The numbers of Ki67þ cells per viewing area were calculated from 20 viewing areas per tumor from six pairs of tumors and are presented asthe mean � SD on the right. C, Tissue sections of the xenografts were immunostained with TUNEL, anti-CD31, or anti-F4/80 antibodies as indicated. Thenumbers of positively stained cells per viewing area were calculated from 25 viewing areas from six pairs of tumors and are presented as the mean � SD.

Liu et al.





Furthermore, high levels of phosphorylated LDHA and lowlevels of LDHB in human prostate cancer tissues were associ-ated with short biochemical-recurrence and survival times ofpatients. Our results suggest that ectopic FGFR1 signalingcontributes to prostate cancer progression by reprograming cellenergy metabolism and those high levels of phosphorylatedLDHA and high expression levels of LDHB are potential bio-markers for prostate cancer diagnosis and prognosis.

LDHA has four tyrosine phosphorylation sites. Phosphory-lation at tyrosine 10 enhances the formation of tetramers andincreases enzymatic activity, while phosphorylation at tyrosine83 enhances the binding of NADH (29). In addition, highlevels of FGFR1 expression are associated with high levels ofphosphorylated LDHA (43). Tyrosine 10 only exists in humanLDHA, suggesting that Y10 phosphorylation is not the onlyway in which FGFR1 activates LDHA. We showed that

Figure 7.

High pLDHA and low LDHB expression predicts poor prognosis in patients with prostate cancer. A, Representative images of immunochemicalstaining of pLDHA and LDHB in the MGH prostate cancer TMA. B, Statistical analyses of the expression of pLDHA and LDHB in prostate cancer andbenign prostate. C, PSA failure-free survival time in patients with low versus high phosphorylated LDHA a LDHB expression. D, Statisticalanalyses of FGFR1 in benign and cancer tissues. E, PSA failure-free survival time in patients with low- versus high-FGFR1 expression. F, Pearsoncorrelation of Fgfr1 and pLDHA in the prostate cancer TMA. G, Model of ectopic FGFR1 signaling in reprograming cell metabolism and promotingtumorigenesis.






phosphorylation of the four tyrosine phosphorylation sites byFGFR1 extended the half-life of LDHA. Ablation of the FGFsignaling axis significantly reduced the half-life of LDHA; thesubstitution of phenylalanine at the four tyrosine residues inLDHA had the same effect (Fig. 2). Together, the results suggestthat tyrosine phosphorylation not only increases the enzymaticactivity of LDHA but also enhances the stability of LDHA. Moreresearch is needed to determine how tyrosine phosphorylationaffects the half-life of LDHA.

It has been reported that LDHB expression is silenced byhypermethylation of the LDHB promoter in human prostatecancer (23). We found that Fgfr1 ablation reduced the meth-ylation of CpG islands in the Ldhb promoter (Fig. 3A and B).The ablation of FGF signaling in MEFs increased the expressionof Tet1, which catalyzes the conversion of methylated guani-dine to hydroxyl-methylated guanidine, the first step of DNAdemethylation. Moreover, ablation of Tet1 increased LDHBexpression although ablation of LDHA or LDHB did not affectTET1 expression (Supplementary Fig. S6). Thus, our resultsdemonstrate that FGF signaling suppresses LDHB expressionby promoting Tet1 expression.

Because LDHA promotes aerobic glycolysis while LDHBfacilitates oxidative phosphorylation, LDHA may enhancetumor progression by fueling aerobic glycolysis and LDHBexerts opposite effects in glycolytic prostate cancer cells. Ourresults in Fig. 4 revealed that LDHA and LDHB have oppositeeffects on prostate cancer growth further support this hypoth-esis. Hence, it is essential to develop new strategies to specif-ically inhibit LDHA without compromising LDHB activity.FGFR1 selectively phosphorylates LDHA on multiple tyrosineresidues, which stabilizes LDHA, and concurrently enhancesthe expression of LDHB. Yet, other signaling pathways may alsoregulate cell metabolism, which may explain why ablation ofFGF signaling did not fully convert LDH expression from LDHAto LDHB. It has been reported that LDHA is degraded viachaperone-mediated autophagy (44). However, no differencein LDHA degradation was observed in DU145 and DU145DR1

cells with or without treating with protease inhibitor leupeptinor autophagy inhibitor bafilomycin (Supplementary Fig. S7),suggesting that FGF signaling protected LDHA degradation viaother mechanism. Although both lactate production and glu-cose consumption are reduced in DU145DR1 cells (Fig. 4), thegrowth curves of DU145 and DU145DR1 were not significantlydifferent in glucose-free medium with or without lactate sup-plement, suggesting that deletion of FGFR1 did not confer thecells ability to use lactate as a main energy source. Furtherefforts are warranted to fully understand how cell metabolismis regulated.

Our data are consistent with reports that loss of LDHBcorrelated with malignancy (20, 26, 45). However, it is dif-ferent from the report that LDHB expression is associated withpoor survival in patients with uterine cancer (27, 46). Oneexplanation is that in the cancer, such as uterine cancer, whichlargely rely on the TCA cycle to fuel cellular activities, can beinhibited by LDHB depletion. In other cancers, such as met-astatic prostate cancer, however, rely on aerobic glycolysis, andtherefore, is benefited from LDHB depletion. Therefore, morestudies are needed to understand the role of LDHB in varioustypes of malignancies. Our data show that although thedeletion of FGFRs led to an increase in oxygen consumption,it still reduced ATP production. These results suggest that the

upregulation of oxidative phosphorylation is not sufficientto make up the energy loss because of FGFR ablation, andthat prostate cancer cells derive ATP primarily from aerobicglycolysis.

Although FRS2a is required for the FGF kinase to activatethe ERK and PI3K/AKT pathways, we found that deletion ofFrs2a alleles did not affect LDHA and LDHB expressions aswell as lactate production in MEFs (Supplementary Fig. S8).This is consistent with the fact that FGFR1 directly phos-phorylates LDHA. The mechanism by which FGF sig-naling regulates LDHB promoter methylation remains to bedetermined.

In summary, we demonstrated that FGF signaling promotesaerobic glycolysis and that ectopic FGF signaling in prostatecancer reprograms cell metabolism, suppressing aerobic glycoly-sis and promoting oxidative phosphorylation. LDHA deletionsuppressed, whereas LDHB deletion promoted, the tumorigenicactivity of prostate cancer cells. Furthermore, the LDHA over-expression and LDHB downregulation correlated with short bio-chemical recurrence and survival times in patients with prostatecancer. Our results shed new light on how the manipulation ofectopic FGF signaling may serve as a strategy for prostate cancertreatment.

Disclosure of Potential Conflicts of InterestNo potential conflicts of interest were disclosed.

Authors' ContributionsConception and design: J. Liu, W.L. McKeehan, C. Wang, W. Zhong,F. WangDevelopment of methodology: J. Liu, G. Chen, Z. Liu, S. Liu, Z. Cai, P. You,Y. Ke, Y. Huang, H. Pelicano, C.-L. Wu, C. Wang, W. Zhong, F. WangAcquisition of data (provided animals, acquired and managed patients,provided facilities, etc.): J. Liu, G. Chen, Z. Liu, S. Liu, Z. Cai, P. You, Y. Ke,L. Zhao, H. Pelicano, C.-L. WuAnalysis and interpretation of data (e.g., statistical analysis, biostatistics,computational analysis): J. Liu, G. Chen, Z. Liu, S. Liu, Z. Cai, P. You, Y. Ke,L. Lai, Y. Huang, H. Gao, L. Zhao, H. Pelicano, P. Huang, W.L. McKeehan,C.-L. Wu, C. Wang, W. Zhong, F. WangWriting, review, and/or revision of the manuscript: J. Liu, G. Chen, Z. Liu,S. Liu, Z. Cai, P. You, Y. Ke, Y. Huang, H. Gao, P. Huang, C.-L. Wu, C. Wang,W. Zhong, F. WangAdministrative, technical, or material support (i.e., reporting or organizingdata, constructing databases): J. Liu, C. Wang, W. Zhong, F. WangStudy supervision: W.L. McKeehan, C.-L. Wu, C. Wang, W. Zhong, F. Wang

AcknowledgmentsWe thank Drs. Juha Patanen and David Ornitz for sharing the Fgfr1floxed and

Fgfr2floxed mice, respectively. This work was supported by the NIH CA96824,TAMU1400302, CPRIT 110555 grants, and a gift from Agilent Technology, Inc.(to F. Wang), the National Natural Science Foundation of China 81101712,31371470, and 81270761 grants (to C.Wang); andNational Key Basic ResearchProgram of China (2015CB553706), National Natural Science Foundation ofChina (81571427) and the Fundamental Research Funds for the CentralUniversities 2017PY023 (to W.D. Zhong).

The costs of publication of this article were defrayed in part by thepayment of page charges. This article must therefore be hereby markedadvertisement in accordance with 18 U.S.C. Section 1734 solely to indicatethis fact.

Received October 20, 2017; revised March 29, 2018; accepted June 4, 2018;published first June 11, 2018.

Liu et al.





References1. Fischer K,HoffmannP, Voelkl S,MeidenbauerN, Ammer J, EdingerM, et al.

Inhibitory effect of tumor cell-derived lactic acid on human T cells. Blood2007;109:3812–9.

2. Brand A, Singer K, Koehl GE, Kolitzus M, Schoenhammer G, Thiel A, et al.LDHA-associated lactic acid production blunts tumor immunosurveillanceby T and NK cells. Cell Metab 2016;24:657–71.

3. Ciccarese C, Santoni M, Massari F, Modena A, Piva F, Conti A, et al.Metabolic alterations in renal and prostate cancer. Curr Drug Metab2015;17:150–5.

4. Xian ZY, Liu JM, ChenQK, ChenHZ, Ye CJ, Xue J, et al. Inhibition of LDHAsuppresses tumor progression in prostate cancer. Tumour Biol 2015;36:8093–100.

5. Pertega-Gomes N, Felisbino S, Massie CE, Vizcaino JR, Coelho R, Sandi C,et al. A glycolytic phenotype is associated with prostate cancer progressionand aggressiveness: a role for monocarboxylate transporters as metabolictargets for therapy. J Pathol 2015;236:517–30.

6. Mraz J, Vrubel F,HanselovaM. Carcinoma of the prostate. II. Serum activityof acid phosphatase, prostatic acid phosphatase, LDH and its isoenzymes.Int Urol Nephrol 1979;11:301–9.

7. Vrubel F, Mraz J, Nemecek R, Papousek F, Hanselova M. Carcinoma of theprostate. I. Histochemical examination as an aid in evaluating prostatecarcinoma. Int Urol Nephrol 1979;11:295–9.

8. Oliver JA, el-Hilali MM, Belitsky P, MacKinnon KJ. LDH isoenzymes inbenign and malignant prostate tissue. The LDH V-I ratio as an index ofmalignancy. Cancer 1970;25:863–6.

9. Naruse K, Yamada Y, Aoki S, Taki T, Nakamura K, TobiumeM, et al. Lactatedehydrogenase is a prognostic indicator for prostate cancer patients withbone metastasis. Hinyokika Kiyo 2007;53:287–92.

10. Keshari KR, SriramR, VanCriekingeM,WilsonDM,Wang ZJ, VigneronDB,et al. Metabolic reprogramming and validation of hyperpolarized 13Clactate as a prostate cancer biomarker using a human prostate tissue sliceculture bioreactor. Prostate 2013;73:1171–81.

11. Wang F, Luo Y, McKeehan W. The FGF signaling axis in prostate tumor-igenesis. In: E. G, C. S, F. R, editors. Molecular oncology: causes of cancerand targets for treatment. London,UnitedKingdom:CambridgeUniversityPress; 2013. p. 186–9.

12. Zhang Y, Zhang J, Lin Y, Lan Y, Lin C, Xuan JW, et al. Role of epithelialcell fibroblast growth factor receptor substrate 2{alpha} in prostatedevelopment, regeneration and tumorigenesis. Development 2008;135:775–84.

13. Yang F, ZhangY, Ressler SJ, IttmannMM,AyalaGE,Dang TD, et al. FGFR1 isessential for prostate cancer progression and metastasis. Cancer Res2013;73:3716–24.

14. Yu P, Wilhelm K, Dubrac A, Tung JK, Alves TC, Fang JS, et al. FGF-dependent metabolic control of vascular development. Nature 2017;545:224–8.

15. Fritz PJ. Rabbit muscle lactate dehydrogenase 5; a regulatory enzyme.Science 1965;150:364–6.

16. Semenza GL, Jiang BH, Leung SW, Passantino R, Concordet JP,Maire P, et al. Hypoxia response elements in the aldolase A, enolase1, and lactate dehydrogenase A gene promoters contain essentialbinding sites for hypoxia-inducible factor 1. J Biol Chem 1996;271:32529–37.

17. Liang X, Liu L, Fu T, ZhouQ, ZhouD, Xiao L, et al. Exercise inducible lactatedehydrogenase B regulates mitochondrial function in skeletal muscle.J Biol Chem 2016;291:25306–18.

18. Dennison JB, Molina JR, Mitra S, Gonzalez-Angulo AM, Balko JM, KubaMG, et al. Lactate dehydrogenase B: a metabolic marker of response toneoadjuvant chemotherapy in breast cancer. Clin Cancer Res 2013;19:3703–13.

19. McCleland ML, Adler AS, Deming L, Cosino E, Lee L, BlackwoodEM, et al. Lactate dehydrogenase B is required for the growth ofKRAS-dependent lung adenocarcinomas. Clin Cancer Res 2013;19:773–84.

20. McClelandML, Adler AS, Shang Y, Hunsaker T, Truong T, Peterson D, et al.An integrated genomic screen identifies LDHB as an essential gene fortriple-negative breast cancer. Cancer Res 2012;72:5812–23.

21. Zha X,Wang F,Wang Y,He S, Jing Y,Wu X, et al. Lactate dehydrogenase B iscritical for hyperactive mTOR-mediated tumorigenesis. Cancer Res2011;71:13–8.

22. Maekawa M, Taniguchi T, Ishikawa J, Sugimura H, Sugano K, Kanno T.Promoter hypermethylation in cancer silences LDHB, eliminating lactatedehydrogenase isoenzymes 1–4. Clin Chem 2003;49:1518–20.

23. Leiblich A, Cross SS, Catto JW, Phillips JT, Leung HY, Hamdy FC, et al.Lactate dehydrogenase-B is silenced by promoter hypermethylation inhuman prostate cancer. Oncogene 2006;25:2953–60.

24. Cui J, Quan M, Jiang W, Hu H, Jiao F, Li N, et al. Suppressed expression ofLDHB promotes pancreatic cancer progression via inducing glycolyticphenotype. Med Oncol 2015;32:143.

25. Kim JH, Kim EL, Lee YK, Park CB, Kim BW, Wang HJ, et al. Decreasedlactate dehydrogenase B expression enhances claudin 1-mediatedhepatoma cell invasiveness via mitochondrial defects. Exp Cell Res2011;317:1108–18.

26. Chen R, Zhou X, Yu Z, Liu J, Huang G. Low expression of LDHB correlateswith unfavorable survival in hepatocellular carcinoma: strobe-compliantarticle. Medicine 2015;94:e1583.

27. Brisson L, Banski P, Sboarina M, Dethier C, Danhier P, Fontenille MJ, et al.Lactate dehydrogenase B controls lysosome activity and autophagy incancer. Cancer Cell 2016;30:418–31.

28. Li SS, Pan YE, Sharief FS, Evans MJ, Lin MF, Clinton GM, et al. Cancer-associated lactate dehydrogenase is a tyrosylphosphorylated formof human LDH-A, skeletal muscle isoenzyme. Cancer Invest 1988;6:93–101.

29. Fan J, Hitosugi T, Chung TW, Xie J, Ge Q, Gu TL, et al. Tyrosine phos-phorylation of lactate dehydrogenase A is important for NADH/NAD(þ)redox homeostasis in cancer cells. Mol Cell Biol 2011;31:4938–50.

30. Lin Y, Liu G, Zhang Y, Hu YP, Yu K, Lin C, et al. Fibroblast growth factorreceptor 2 tyrosine kinase is required for prostatic morphogenesis and theacquisition of strict androgen dependency for adult tissue homeostasis.Development 2007;134:723–34.

31. Wang C, Chang JY, Yang C, Huang Y, Liu J, You P, et al. Type 1 fibroblastgrowth factor receptor in cranial neural crest cells-derived mesenchyme isrequired for palatogenesis. J Biol Chem 2013;288:22174–83.

32. Huang Y, Jin C, Hamana T, Liu J, Wang C, An L, et al. Overexpressionof FGF9 in prostate epithelial cells augments reactive stroma forma-tion and promotes prostate cancer progression. Int J Biol Sci 2015;11:948–60.

33. Zhong WD, Liang YX, Lin SX, Li L, He HC, Bi XC, et al. Expression ofCD147 is associated with prostate cancer progression. Int J Cancer2012;130:300–8.

34. Nishitani S, Takehana K, Fujitani S, Sonaka I. Branched-chain amino acidsimprove glucose metabolism in rats with liver cirrhosis. Am J PhysiolGastrointest Liver Physiol 2005;288:G1292–300.

35. Acevedo VD, Gangula RD, Freeman KW, Li R, Zhang Y, Wang F, et al.Inducible FGFR-1 activation leads to irreversible prostate adenocarci-noma and an epithelial-to-mesenchymal transition. Cancer Cell 2007;12:559–71.

36. Ozen M, Giri D, Ropiquet F, Mansukhani A, Ittmann M. Role of fibroblastgrowth factor receptor signaling in prostate cancer cell survival. J NatlCancer Inst 2001;93:1783–90.

37. Abate-Shen C, Shen MM. FGF signaling in prostate tumorigenesis–newinsights into epithelial–stromal interactions. Cancer Cell 2007;12:495–7.

38. Taylor BS, Schultz N, Hieronymus H, Gopalan A, Xiao Y, Carver BS, et al.Integrative genomic profiling of human prostate cancer. Cancer Cell2010;18:11–22.

39. Giri D, Ropiquet F, IttmannM. Alterations in expression of basic fibroblastgrowth factor (FGF) 2 and its receptor FGFR-1 in human prostate cancer.Clin Cancer Res 1999;5:1063–71.

40. Devilard E, Bladou F, RamuzO, KarsentyG,Dales JP, Gravis G, et al. FGFR1and WT1 are markers of human prostate cancer progression. BMC Cancer2006;6:272.

41. Wang J, Stockton DW, Ittmann M. The fibroblast growth factor receptor-4Arg388 allele is associated with prostate cancer initiation and progression.Clin Cancer Res 2004;10:6169–78.

42. Sugiura K, Su YQ, Diaz FJ, Pangas SA, Sharma S, Wigglesworth K, et al.Oocyte-derived BMP15 and FGFs cooperate to promote glycolysis incumulus cells. Development 2007;134:2593–603.

43. Jin L, Chun J, Pan C, Alesi GN, Li D, Magliocca KR, et al. Phosphorylation-mediated activation of LDHA promotes cancer cell invasion and tumourmetastasis. Oncogene 2017;36:3797–806.






44. Zhao D, Zou SW, Liu Y, Zhou X, Mo Y, Wang P, et al. Lysine-5 acetylationnegatively regulates lactate dehydrogenase A and is decreased in pancreaticcancer. Cancer Cell 2013;23:464–76.

45. Koh YW, Lee SJ, Park SY. Prognostic significance of lactate dehydroge-nase B according to histologic type of non-small-cell lung cancer and its

association with serum lactate dehydrogenase. Pathol Res Pract 2017;213:1134–8.

46. Li C, Chen Y, Bai P, Wang J, Liu Z, Wang T, et al. LDHBmay be a significantpredictor of poor prognosis in osteosarcoma. Am J Transl Res 2016;8:4831–43.


Liu et al.




2018;78:4459-4470. Published OnlineFirst June 11, 2018.Cancer Res Junchen Liu, Guo Chen, Zezhen Liu, et al. Prostate CancerEffect by Reprogramming LDH Isoform Expression and Activity in Aberrant FGFR Tyrosine Kinase Signaling Enhances the Warburg

Updated version

10.1158/0008-5472.CAN-17-3226doi:

Access the most recent version of this article at:

Material

Supplementary

http://cancerres.aacrjournals.org/content/suppl/2018/10/18/0008-5472.CAN-17-3226.DC1

Access the most recent supplemental material at:

Overview

Visual

http://cancerres.aacrjournals.org/content/78/16/4459/F1.large.jpgA diagrammatic summary of the major findings and biological implications:

Cited articles

http://cancerres.aacrjournals.org/content/78/16/4459.full#ref-list-1

This article cites 45 articles, 17 of which you can access for free at:

E-mail alerts related to this article or journal.Sign up to receive free email-alerts

Subscriptions

Reprints and

[email protected]

To order reprints of this article or to subscribe to the journal, contact the AACR Publications Department at

Permissions

Rightslink site. Click on "Request Permissions" which will take you to the Copyright Clearance Center's (CCC)

.http://cancerres.aacrjournals.org/content/78/16/4459To request permission to re-use all or part of this article, use this link



http://cancerres.aacrjournals.org/lookup/doi/10.1158/0008-5472.CAN-17-3226http://cancerres.aacrjournals.org/content/suppl/2018/10/18/0008-5472.CAN-17-3226.DC1http://cancerres.aacrjournals.org/content/78/16/4459/F1.large.jpghttp://cancerres.aacrjournals.org/content/78/16/4459.full#ref-list-1http://cancerres.aacrjournals.org/cgi/alertsmailto:[email protected]://cancerres.aacrjournals.org/content/78/16/4459http://cancerres.aacrjournals.org/

/ColorImageDict > /JPEG2000ColorACSImageDict > /JPEG2000ColorImageDict > /AntiAliasGrayImages false /CropGrayImages false /GrayImageMinResolution 200 /GrayImageMinResolutionPolicy /Warning /DownsampleGrayImages true /GrayImageDownsampleType /Bicubic /GrayImageResolution 300 /GrayImageDepth -1 /GrayImageMinDownsampleDepth 2 /GrayImageDownsampleThreshold 1.50000 /EncodeGrayImages true /GrayImageFilter /DCTEncode /AutoFilterGrayImages true /GrayImageAutoFilterStrategy /JPEG /GrayACSImageDict > /GrayImageDict > /JPEG2000GrayACSImageDict > /JPEG2000GrayImageDict > /AntiAliasMonoImages false /CropMonoImages false /MonoImageMinResolution 600 /MonoImageMinResolutionPolicy /Warning /DownsampleMonoImages true /MonoImageDownsampleType /Bicubic /MonoImageResolution 900 /MonoImageDepth -1 /MonoImageDownsampleThreshold 1.50000 /EncodeMonoImages true /MonoImageFilter /CCITTFaxEncode /MonoImageDict > /AllowPSXObjects false /CheckCompliance [ /None ] /PDFX1aCheck false /PDFX3Check false /PDFXCompliantPDFOnly false /PDFXNoTrimBoxError true /PDFXTrimBoxToMediaBoxOffset [ 0.00000 0.00000 0.00000 0.00000 ] /PDFXSetBleedBoxToMediaBox true /PDFXBleedBoxToTrimBoxOffset [ 0.00000 0.00000 0.00000 0.00000 ] /PDFXOutputIntentProfile (None) /PDFXOutputConditionIdentifier () /PDFXOutputCondition () /PDFXRegistryName () /PDFXTrapped /False

/CreateJDFFile false /Description > /Namespace [ (Adobe) (Common) (1.0) ] /OtherNamespaces [ > /FormElements false /GenerateStructure false /IncludeBookmarks false /IncludeHyperlinks false /IncludeInteractive false /IncludeLayers false /IncludeProfiles false /MarksOffset 18 /MarksWeight 0.250000 /MultimediaHandling /UseObjectSettings /Namespace [ (Adobe) (CreativeSuite) (2.0) ] /PDFXOutputIntentProfileSelector /NA /PageMarksFile /RomanDefault /PreserveEditing true /UntaggedCMYKHandling /LeaveUntagged /UntaggedRGBHandling /LeaveUntagged /UseDocumentBleed false >> > ]>> setdistillerparams> setpagedevice

Aberrant FGFR Tyrosine Kinase Signaling Enhances the Warburg … · Metabolism and Chemical Biology...

Documents

Transcript of Aberrant FGFR Tyrosine Kinase Signaling Enhances the Warburg … · Metabolism and Chemical Biology...