Targeting metabolic remodeling represents a potentially promising strategy for hepatocellular carcinoma (HCC) therapy. In-depth understanding on the regulation of the glutamine transporter alanine-serine-cysteine transporter 2 (ASCT2) contributes to the development of novel promising therapeutics. As a developmentally regulated RNA binding protein, RBM45 is capable to shuttle between nucleus and cytoplasm, and directly interacts with proteins. By bioinformatics analysis, we screened out that RBM45 was elevated in the HCC patient specimens and positively correlated with poor prognosis. RBM45 promoted cell proliferation, boosted xenograft tumorigenicity and accelerated HCC progression. Using untargeted metabolomics, it was found that RBM45 interfered with glutamine metabolism. Further results demonstrated that RBM45 positively associated with ASCT2 in human and mouse specimens. Moreover, RBM45 enhanced ASCT2 protein stability by counteracting autophagy-independent lysosomal degradation. Significantly, wild-type ASCT2, instead of phospho-defective mutants, rescued siRBM45-suppressed HCC cell proliferation. Using molecular docking approaches, we found AG-221, a mutant isocitrate dehydrogenase 2 (mIDH2) inhibitor for acute myeloid leukemia therapy, pharmacologically perturbed RBM45-ASCT2 interaction, decreased ASCT2 stability and suppressed HCC progression. These findings provide evidence that RBM45 plays a crucial role in HCC progression via interacting with and counteracting the degradation of ASCT2. Our findings suggest a novel alternative structural sites for the design of ASCT2 inhibitors and the agents interfering with RBM45-ASCT2 interaction may be a potential direction for HCC drug development.
This is a preview of subscription content, access via your institution
Subscribe to this journal
Receive 50 print issues and online access
$259.00 per year
only $5.18 per issue
Rent or buy this article
Prices vary by article type
Prices may be subject to local taxes which are calculated during checkout
Source data and reagents are available from the corresponding author upon reasonable request.
Gao P, Tchernyshyov I, Chang TC, Lee YS, Kita K, Ochi T, et al. c-Myc suppression of miR-23a/b enhances mitochondrial glutaminase expression and glutamine metabolism. Nature. 2009;458:762–5.
Son J, Lyssiotis CA, Ying HQ, Wang XX, Hua SJ, Ligorio M, et al. Glutamine supports pancreatic cancer growth through a KRAS-regulated metabolic pathway. Nature. 2013;499:101–5.
Wise DR, DeBerardinis RJ, Mancuso A, Sayed N, Zhang XY, Pfeiffer HK, et al. Myc regulates a transcriptional program that stimulates mitochondrial glutaminolysis and leads to glutamine addiction. Proc Natl Acad Sci USA. 2008;105:18782–7.
Liu W, Le A, Hancock C, Lane AN, Dang CV, Fan TW, et al. Reprogramming of proline and glutamine metabolism contributes to the proliferative and metabolic responses regulated by oncogenic transcription factor c-MYC. Proc Natl Acad Sci USA. 2012;109:8983–8.
Jin H, Wang S, Zaal EA, Wang C, Wu H, Bosma A, et al. A powerful drug combination strategy targeting glutamine addiction for the treatment of human liver cancer. Elife. 2020;9:e56749.
Cormerais Y, Massard PA, Vucetic M, Giuliano S, Tambutte E, Durivault J, et al. The glutamine transporter ASCT2 (SLC1A5) promotes tumor growth independently of the amino acid transporter LAT1 (SLC7A5). J Biol Chem. 2018;293:2877–87.
Sun HW, Yu XJ, Wu WC, Chen J, Shi M, Zheng LM, et al. GLUT1 and ASCT2 as predictors for prognosis of hepatocellular carcinoma. PLoS ONE. 2016;11:e0168907.
Hensley CT, Wasti AT, DeBerardinis RJ. Glutamine and cancer: cell biology, physiology, and clinical opportunities. J Clin Invest. 2013;123:3678–84.
Willems L, Jacque N, Jacquel A, Neveux N, Maciel TT, Lambert M, et al. Inhibiting glutamine uptake represents an attractive new strategy for treating acute myeloid leukemia. Blood. 2013;122:3521–32.
van Geldermalsen M, Wang Q, Nagarajah R, Marshall AD, Thoeng A, Gao D, et al. ASCT2/SLC1A5 controls glutamine uptake and tumour growth in triple-negative basal-like breast cancer. Oncogene. 2016;35:3201–8.
Schulte ML, Fu A, Zhao P, Li J, Geng L, Smith ST, et al. Pharmacological blockade of ASCT2-dependent glutamine transport leads to antitumor efficacy in preclinical models. Nat Med. 2018;24:194–202.
Colas C, Grewer C, Otte NJ, Gameiro A, Albers T, Singh K, et al. Ligand discovery for the alanine-serine-cysteine transporter (ASCT2, SLC1A5) from homology modeling and virtual screening. PLoS Comput Biol. 2015;11:e1004477.
Schifferli KP, Monks NR, Tammali R, Borrok MJ, Flynn MG, Hurt EM, et al. MEDI7247: a first in class antibody drug conjugate targeting ASCT2 in a range of solid tumors. Cancer Res. 2018;78:LB-298.
Broer A, Fairweather S, Broer S. Disruption of amino acid homeostasis by novel ASCT2 inhibitors involves multiple targets. Front Pharm. 2018;9:785.
Pore N, Schifferli KP, Monks NR, Tammali R, Borrok M, Hurt E, et al. Discovery and development of MEDI7247, a novel pyrrolobenzodiazepine (PBD)-based antibody drug conjugate targeting ASCT2, for treating hematological cancers. Blood. 2018;132:4071.
Reynolds MR, Lane AN, Robertson B, Kemp S, Liu Y, Hill BG, et al. Control of glutamine metabolism by the tumor suppressor Rb. Oncogene. 2014;33:556–66.
Jeon YJ, Khelifa S, Ratnikov B, Scott DA, Feng Y, Parisi F, et al. Regulation of glutamine carrier proteins by RNF5 determines breast cancer response to ER stress-inducing chemotherapies. Cancer Cell. 2015;27:354–69.
Lee DE, Yoo JE, Kim J, Kim S, Kim S, Lee H, et al. NEDD4L downregulates autophagy and cell growth by modulating ULK1 and a glutamine transporter. Cell Death Dis. 2020;11:38.
Console LSM, Tarmakova Z, Coe IR, Indiveri C. N-linked glycosylation of human SLC1A5 (ASCT2) transporter is critical for trafficking to membrane. Biochim Biophys Acta. 2015;1853:1636–45.
Cooper-Knock J, Robins H, Niedermoser I, Wyles M, Heath PR, Higginbottom A, et al. Targeted genetic screen in amyotrophic lateral sclerosis reveals novel genetic variants with synergistic effect on clinical phenotype. Front Mol Neurosci. 2017;10:370.
Tamada H, Sakashita E, Shimazaki K, Ueno E, Hamamoto T, Kagawa Y, et al. cDNA cloning and characterization of Drb1, a new member of RRM-type neural RNA-binding protein. Biochem Biophys Res Commun. 2002;297:96–104.
Choi SH, Flamand MN, Liu B, Zhu H, Hu M, Wang M, et al. RBM45 is an m(6)A-binding protein that affects neuronal differentiation and the splicing of a subset of mRNAs. Cell Rep. 2022;40:111293.
Mashiko T, Sakashita E, Kasashima K, Tominaga K, Kuroiwa K, Nozaki Y, et al. Developmentally regulated RNA-binding protein 1 (Drb1)/RNA-binding motif protein 45 (RBM45), a nuclear-cytoplasmic trafficking protein, forms TAR DNA-binding protein 43 (TDP-43)-mediated cytoplasmic aggregates. J Biol Chem. 2016;291:14996–5007.
Chen X, Yang Z, Wang W, Qian K, Liu M, Wang J, et al. Structural basis for RNA recognition by the N-terminal tandem RRM domains of human RBM45. Nucleic Acids Res. 2021;49:2946–58.
Gong J, Huang M, Wang F, Ma X, Liu H, Tu Y, et al. RBM45 competes with HDAC1 for binding to FUS in response to DNA damage. Nucleic Acids Res. 2017;45:12862–76.
Bakkar N, Kousari A, Kovalik T, Li Y, Bowser R. RBM45 modulates the antioxidant response in amyotrophic lateral sclerosis through interactions with KEAP1. Mol Cell Biol. 2015;35:2385–99.
del Rio-Moreno M, Alors-Perez E, Gonzalez-Rubio S, Ferrin G, Reyes O, Rodriguez-Peralvarez M, et al. Dysregulation of the splicing machinery is associated to the development of nonalcoholic fatty liver disease. J Clin Endocr Metab. 2019;104:3389–402.
Ma RH, Zhang WG, Tang K, Zhang HF, Zhang Y, Li DP, et al. Switch of glycolysis to gluconeogenesis by dexamethasone for treatment of hepatocarcinoma. Nat Commun. 2013;4:2508.
Liu Q, Li J, Zhang W, Xiao C, Zhang S, Nian C, et al. Glycogen accumulation and phase separation drives liver tumor initiation. Cell. 2021;184:5559–76.e19.
Nicklin P, Bergman P, Zhang B, Triantafellow E, Wang H, Nyfeler B, et al. Bidirectional transport of amino acids regulates mTOR and autophagy. Cell. 2009;136:521–34.
Saito Y, Li L, Coyaud E, Luna A, Sander C, Raught B, et al. LLGL2 rescues nutrient stress by promoting leucine uptake in ER(+) breast cancer. Nature. 2019;569:275–9.
Fuchs BC, Finger RE, Onan MC, Bode BP. ASCT2 silencing regulates mammalian target-of-rapamycin growth and survival signaling in human hepatoma cells. Am J Physiol Cell Physiol. 2007;293:C55–63.
Yoo HC, Park SJ, Nam M, Kang J, Kim K, Yeo JH, et al. A variant of SLC1A5 is a mitochondrial glutamine transporter for metabolic reprogramming in cancer cells. Cell Metab. 2020;31:267–83.e12.
Martinez Molina D, Jafari R, Ignatushchenko M, Seki T, Larsson EA, Dan C, et al. Monitoring drug target engagement in cells and tissues using the cellular thermal shift assay. Science. 2013;341:84–7.
Esslinger CS, Cybulski KA, Rhoderick JF. N-gamma-Aryl glutamine analogues as probes of the ASCT2 neutral amino acid transporter binding site. Bioorg Med Chem. 2005;13:1111–8.
Oppedisano F, Catto M, Koutentis PA, Nicolotti O, Pochini L, Koyioni M, et al. Inactivation of the glutamine/amino acid transporter ASCT2 by 1,2,3-dithiazoles: proteoliposomes as a tool to gain insights in the molecular mechanism of action and of antitumor activity. Toxicol Appl Pharm. 2012;265:93–102.
Hara Y, Minami Y, Yoshimoto S, Hayashi N, Yamasaki A, Ueda S, et al. Anti-tumor effects of an antagonistic mAb against the ASCT2 amino acid transporter on KRAS-mutated human colorectal cancer cells. Cancer Med. 2020;9:302–12.
Garibsingh RAA, Ndaru E, Garaeva AA, Shi YY, Zielewicz L, Zakrepine P, et al. Rational design of ASCT2 inhibitors using an integrated experimental-computational approach. Proc Natl Acad Sci USA. 2021;118:e2104093118.
Dong Y, Wang JL, Eisenberg G, Yu XZ, Schlessinger A, Grewer CT. Conserved allosteric binding pocket in SLC1 glutamate transporter EAAT1 and neutral amino acid transporter ASCT2. Biophys J. 2022;121:250a.
Yu W, Huang JW, Dong QC, Li WT, Jiang L, Zhang Q, et al. Ag120-mediated inhibition of ASCT2-dependent glutamine transport has an anti-tumor effect on colorectal cancer cells. Front Pharm. 2022;13:871392.
Xu D, Hemler ME. Metabolic activation-related CD147-CD98 complex. Mol Cell Proteom. 2005;4:1061–71.
Tao XA, Lu Y, Qiu SB, Wang Y, Qin J, Fan Z. AP1G1 is involved in cetuximab-mediated downregulation of ASCT2-EGFR complex and sensitization of human head and neck squamous cell carcinoma cells to ROS-induced apoptosis. Cancer Lett. 2017;408:33–42.
Yang Z, Follett J, Kerr MC, Clairfeuille T, Chandra M, Collins BM, et al. Sorting nexin 27 (SNX27) regulates the trafficking and activity of the glutamine transporter ASCT2. J Biol Chem. 2018;293:6802–11.
Zhou Q, Lin W, Wang C, Sun F, Ju S, Chen Q, et al. Neddylation inhibition induces glutamine uptake and metabolism by targeting CRL3(SPOP) E3 ligase in cancer cells. Nat Commun. 2022;13:3034.
Clairfeuille T, Mas C, Chan AS, Yang Z, Tello-Lafoz M, Chandra M, et al. A molecular code for endosomal recycling of phosphorylated cargos by the SNX27-retromer complex. Nat Struct Mol Biol. 2016;23:921–32.
Pan YL, Han MZ, Zhang XC, He Y, Yuan CY, Xiong YX, et al. Discoidin domain receptor 1 promotes hepatocellular carcinoma progression through modulation of SLC1A5 and the mTORC1 signaling pathway. Cell Oncol. 2022;45:163–78.
Avissar NE, Sax HC, Toia L. In human entrocytes, GLN transport and ASCT2 surface expression induced by short-term EGF are MAPK, PI3K, and Rho-dependent. Dig Dis Sci. 2008;53:2113–25.
Li Y, Collins M, Geiser R, Bakkar N, Riascos D, Bowser R. RBM45 homo-oligomerization mediates association with ALS-linked proteins and stress granules. Sci Rep. 2015;5:14262.
Huang MS, Chang JH, Lin WC, Cheng YH, Li FA, Suen CS, et al. SLC38A2 overexpression induces a cancer-like metabolic profile and cooperates with SLC1A5 in Pan-cancer prognosis. Chem Asian J 2020;15:3861–72.
Yen K, Travins J, Wang F, David MD, Artin E, Straley K, et al. AG-221, a first-in-class therapy targeting acute myeloid leukemia harboring oncogenic IDH2 mutations. Cancer Discov. 2017;7:478–93.
Wu Y, Du H, Zhan M, Wang H, Chen P, Du D, et al. Sepiapterin reductase promotes hepatocellular carcinoma progression via FoxO3a/Bim signaling in a nonenzymatic manner. Cell Death Dis. 2020;11:248.
Tward AD, Jones KD, Yant S, Cheung ST, Fan ST, Chen X, et al. Distinct pathways of genomic progression to benign and malignant tumors of the liver. Proc Natl Acad Sci USA. 2007;104:14771–6.
Chen X, Calvisi DF. Hydrodynamic transfection for generation of novel mouse models for liver cancer research. Am J Pathol. 2014;184:912–23.
Du H, Chen Y, Hou X, Huang Y, Wei X, Yu X, et al. PLOD2 regulated by transcription factor FOXA1 promotes metastasis in NSCLC. Cell Death Dis. 2017;8:e3143.
Wei X, Li S, He J, Du H, Liu Y, Yu W, et al. Tumor-secreted PAI-1 promotes breast cancer metastasis via the induction of adipocyte-derived collagen remodeling. Cell Commun Signal. 2019;17:58.
Fu T, Zhang M, Zhou Z, Wu P, Peng C, Wang Y, et al. Structural and biochemical advances on the recruitment of the autophagy-initiating ULK and TBK1 complexes by autophagy receptor NDP52. Sci Adv. 2021;7:eabi6582.
Li H, Sun L, Li H, Lv X, Semukunzi H, Li R, et al. DT-13 synergistically enhanced vinorelbine-mediated mitotic arrest through inhibition of FOXM1-BICD2 axis in non-small-cell lung cancer cells. Cell Death Dis. 2017;8:e2810.
Wang Q, Bailey CG, Ng C, Tiffen J, Thoeng A, Minhas V, et al. Androgen receptor and nutrient signaling pathways coordinate the demand for increased amino acid transport during prostate cancer progression. Cancer Res. 2011;71:7525–36.
Wang Z, Zhang Z, Li Y, Sun L, Peng D, Du D, et al. Preclinical efficacy against acute myeloid leukaemia of SH1573, a novel mutant IDH2 inhibitor approved for clinical trials in China. Acta Pharm Sin B. 2021;11:1526–40.
Hashimoto M, Girardi E, Eichner R, Superti-Furga G. Detection of chemical engagement of solute carrier proteins by a cellular thermal shift assay. ACS Chem Biol. 2018;13:1480–6.
Tang Z, Xie H, Heier C, Huang J, Zheng Q, Eichmann TO, et al. Enhanced monoacylglycerol lipolysis by ABHD6 promotes NSCLC pathogenesis. EBioMedicine. 2020;53:102696.
Pang ZQ, Zhou GY, Ewald J, Chang L, Hacariz O, Basu N, et al. Using MetaboAnalyst 5.0 for LC-HRMS spectra processing, multi-omics integration and covariate adjustment of global metabolomics data. Nat Protoc. 2022;17:1735–61.
The study was supported by the National Natural Science Foundation of China (Nos. 82070883, 82273982, 81872892), the Natural Science Foundation of Jiangsu Province, China (BK20221525) and Scientific Research Foundation for high-level faculty, China Pharmaceutical University. We thank all the financial support for this project and Dr. Xin Chen (UCSF) for sharing the oncogene plasmids.
The authors declare no competing interests.
Publisher’s note Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.
About this article
Cite this article
Du, D., Qin, M., Shi, L. et al. RNA binding motif protein 45-mediated phosphorylation enhances protein stability of ASCT2 to promote hepatocellular carcinoma progression. Oncogene 42, 3127–3141 (2023). https://doi.org/10.1038/s41388-023-02795-3