It’s Not What You Say But How You Say It: Targeting RNA Methylation in AML
2020; Elsevier BV; Volume: 78; Issue: 6 Linguagem: Inglês
10.1016/j.molcel.2020.05.027
ISSN1097-4164
AutoresSarah Naomi Olsen, Scott A. Armstrong,
Tópico(s)Cancer-related molecular mechanisms research
ResumoIn this month's issue of Cancer Cell, Su et al., 2020Su R. Dong L. Li Y. Gao M. Han L. Wunderlich M. Deng X. Li H. Huang Y. Gao L. et al.Targeting FTO Suppresses Cancer Stem Cell Maintenance and Immune Evasion.Cancer Cell. 2020; 38: 1-18Abstract Full Text Full Text PDF Scopus (251) Google Scholar describe two small-molecule inhibitors of the RNA demethylase FTO that demonstrate significant anti-tumor effects in various models of acute myeloid leukemia. In this month's issue of Cancer Cell, Su et al., 2020Su R. Dong L. Li Y. Gao M. Han L. Wunderlich M. Deng X. Li H. Huang Y. Gao L. et al.Targeting FTO Suppresses Cancer Stem Cell Maintenance and Immune Evasion.Cancer Cell. 2020; 38: 1-18Abstract Full Text Full Text PDF Scopus (251) Google Scholar describe two small-molecule inhibitors of the RNA demethylase FTO that demonstrate significant anti-tumor effects in various models of acute myeloid leukemia. RNA methylation has emerged as a new and important regulatory mechanism in eukaryotic gene expression. Similar to histone and DNA modifications, RNA is methylated and demethylated in a dynamic process, tightly regulated by designated "writers" and "erasers." Methylated RNA is then recognized by a diverse set of "reader" proteins. While more than 150 modifications have been identified on cellular RNA, the N6-methyladenosine (m6A) RNA modification is the most abundant and well studied (Shi et al., 2019Shi H. Wei J. He C. Where, When, and How: Context-Dependent Functions of RNA Methylation Writers, Readers, and Erasers.Mol. Cell. 2019; 74: 640-650Abstract Full Text Full Text PDF PubMed Scopus (684) Google Scholar). The majority of the m6A methylation on mRNA is installed by an enzymatic complex composed of a heterodimer of METTL3 (catalytic core) and METTL14, as well as additional adaptor proteins (Figure 1). While to date only two erasers have been identified that remove the m6A modification, FTO and ALKBH5, there is an expanding list of reader proteins that recognize the m6A modification and lead to various downstream effects, including mRNA decay, mRNA stabilization, and enhanced translation (Figure 1, reviewed in Shi et al., 2019Shi H. Wei J. He C. Where, When, and How: Context-Dependent Functions of RNA Methylation Writers, Readers, and Erasers.Mol. Cell. 2019; 74: 640-650Abstract Full Text Full Text PDF PubMed Scopus (684) Google Scholar). The maintenance of the dynamic balance between the deposition and removal of the m6A modification is essential for normal biology and development (reviewed in Yue et al., 2015Yue Y. Liu J. He C. RNA N6-methyladenosine methylation in post-transcriptional gene expression regulation.Genes Dev. 2015; 29: 1343-1355Crossref PubMed Scopus (562) Google Scholar). Consequently, dysregulation of this process may be associated with disease, and emerging data suggest that expression levels of m6A readers, writers, and erasers are often dysregulated in cancer (reviewed in Huang et al., 2020Huang H. Weng H. Chen J. m6A Modification in Coding and Non-coding RNAs: Roles and Therapeutic Implications in Cancer.Cancer Cell. 2020; 37: 270-288Abstract Full Text Full Text PDF PubMed Scopus (418) Google Scholar). This recognition has prompted the development of small molecules that target members of the m6A machinery, including small-molecule inhibitors of METTL3 (reviewed in Cully, 2019Cully M. Chemical inhibitors make their RNA epigenetic mark.Nat. Rev. Drug Discov. 2019; 18: 892-894Crossref PubMed Scopus (35) Google Scholar) and FTO (Huang et al., 2019Huang Y. Su R. Sheng Y. Dong L. Dong Z. Xu H. Ni T. Zhang Z.S. Zhang T. Li C. et al.Small-Molecule Targeting of Oncogenic FTO Demethylase in Acute Myeloid Leukemia.Cancer Cell. 2019; 35: 677-691.e10Abstract Full Text Full Text PDF PubMed Scopus (356) Google Scholar). While the development of METTL3 inhibitors is relatively advanced, FTO inhibitors remain somewhat behind, and their potential has been limited due to low sensitivity and/or specificity. In this month's issue of Cancer Cell, Su et al., 2020Su R. Dong L. Li Y. Gao M. Han L. Wunderlich M. Deng X. Li H. Huang Y. Gao L. et al.Targeting FTO Suppresses Cancer Stem Cell Maintenance and Immune Evasion.Cancer Cell. 2020; 38: 1-18Abstract Full Text Full Text PDF Scopus (251) Google Scholar identified two new potent small-molecule inhibitors of FTO, which are significantly more effective than previously developed molecules. Moreover, Su et al. demonstrated that targeting FTO could significantly suppress leukemia by affecting leukemia stem cell associated pathways and T cell cytotoxicity. Su and colleagues identified the new FTO inhibitors through a series of screening and validation experiments. The authors conducted a structure-based virtual screen of the NCI DTP library containing 260,000 compounds and they subsequently tested over 200 of the top hits in cell-based viability assays. Further validation experiments in additional cell lines narrowed down the list to two lead compounds, CS1 and CS2, both of which displayed consistent effects on proliferation in AML cell lines and FTO's demethylase activity in cell-free assays. Through a variety of cell-free and cell-based assays, Su et al., 2020Su R. Dong L. Li Y. Gao M. Han L. Wunderlich M. Deng X. Li H. Huang Y. Gao L. et al.Targeting FTO Suppresses Cancer Stem Cell Maintenance and Immune Evasion.Cancer Cell. 2020; 38: 1-18Abstract Full Text Full Text PDF Scopus (251) Google Scholar were able to demonstrate that CS1 and CS2 interact with FTO by binding to its catalytic pocket. Importantly, CS1 and CS2 efficiently suppressed m6A demethylase activity of FTO with IC50 values in the nanomolar range, and CS1 as well as CS2 treatment disrupted the interaction of FTO with well-known target mRNAs (such as MYC, CEBPA, RARA; Li et al., 2017Li Z. Weng H. Su R. Weng X. Zuo Z. Li C. Huang H. Nachtergaele S. Dong L. Hu C. et al.FTO Plays an Oncogenic Role in Acute Myeloid Leukemia as a N6-Methyladenosine RNA Demethylase.Cancer Cell. 2017; 31: 127-141Abstract Full Text Full Text PDF PubMed Scopus (862) Google Scholar). The authors then proceeded to functionally characterize the CS1 and CS2 compounds. In line with previous publications that established inhibition of FTO as a therapeutic vulnerability in AML (Huang et al., 2019Huang Y. Su R. Sheng Y. Dong L. Dong Z. Xu H. Ni T. Zhang Z.S. Zhang T. Li C. et al.Small-Molecule Targeting of Oncogenic FTO Demethylase in Acute Myeloid Leukemia.Cancer Cell. 2019; 35: 677-691.e10Abstract Full Text Full Text PDF PubMed Scopus (356) Google Scholar, Li et al., 2017Li Z. Weng H. Su R. Weng X. Zuo Z. Li C. Huang H. Nachtergaele S. Dong L. Hu C. et al.FTO Plays an Oncogenic Role in Acute Myeloid Leukemia as a N6-Methyladenosine RNA Demethylase.Cancer Cell. 2017; 31: 127-141Abstract Full Text Full Text PDF PubMed Scopus (862) Google Scholar), treatment with CS1 and CS2 led to cell cycle arrest and apoptosis. Moreover, the authors established that FTO is overexpressed in human primary AML patient cells compared to healthy donors and that FTO levels are higher in CD34+ immature AML cells compared to bulk CD34- cells, indicating that FTO might be particularly important in leukemia stem cells (LSC). Indeed, 50nM CS1 could almost completely inhibit the repopulating capacity of AML cells as measured by limiting dilution assays, highlighting the potent effect of FTO inhibition on suppressing self-renewal of LSCs. Su et al., 2020Su R. Dong L. Li Y. Gao M. Han L. Wunderlich M. Deng X. Li H. Huang Y. Gao L. et al.Targeting FTO Suppresses Cancer Stem Cell Maintenance and Immune Evasion.Cancer Cell. 2020; 38: 1-18Abstract Full Text Full Text PDF Scopus (251) Google Scholar also performed extensive additional in vivo studies with CS1 and CS2 using multiple models of AML, including three different patient-derived xenotransplantation (PDX) models as well as well-defined murine models. FTO inhibition significantly delayed AML progression and increased survival in all models. To address the mechanistic basis of their tool compounds, the authors performed RNA sequencing. Cluster analysis revealed that CS1 and CS2 treated samples grouped together with FTO knockdown, supporting the idea that the molecules act through modulation of the FTO signaling pathway. Su et al., 2020Su R. Dong L. Li Y. Gao M. Han L. Wunderlich M. Deng X. Li H. Huang Y. Gao L. et al.Targeting FTO Suppresses Cancer Stem Cell Maintenance and Immune Evasion.Cancer Cell. 2020; 38: 1-18Abstract Full Text Full Text PDF Scopus (251) Google Scholar also confirmed MYC and CEBPA mRNA downregulation upon CS1 as well as CS2 treatment and RARA and ASB2 upregulation, known positive and negative targets of FTO, respectively (Li et al., 2017Li Z. Weng H. Su R. Weng X. Zuo Z. Li C. Huang H. Nachtergaele S. Dong L. Hu C. et al.FTO Plays an Oncogenic Role in Acute Myeloid Leukemia as a N6-Methyladenosine RNA Demethylase.Cancer Cell. 2017; 31: 127-141Abstract Full Text Full Text PDF PubMed Scopus (862) Google Scholar). Additionally, the authors identified LILRB4—a gene that has been shown to suppress T cell activity and promote tumor infiltration into surrounding tissue (Deng et al., 2018Deng M. Gui X. Kim J. Xie L. Chen W. Li Z. He L. Chen Y. Chen H. Luo W. et al.LILRB4 signalling in leukaemia cells mediates T cell suppression and tumour infiltration.Nature. 2018; 562: 605-609Crossref PubMed Scopus (122) Google Scholar)—as an important target of FTO. Treatment with CS1, CS2, or knockdown of FTO reduced LILRB4 expression, while expression of wild-type FTO, but not mutant FTO, increased LILRB4 mRNA. This led the authors to hypothesize that FTO inhibition can modulate immune responses. To test this, Su et al. pretreated AML cells with CS1 or CS2 and then co-cultured the cells with activated T cells. FTO inhibition did indeed sensitize AML cells to T cell cytotoxicity. The authors also confirmed this in vivo and treated mice that were injected with human AML xenografts either with CS1 or CS2 alone or in combination with activated T cells, demonstrating that combination treatment significantly prolonged survival compared to treatment with single agents. Similarly, combination treatment of CS1 or CS2 with the hypomethylating agent DAC also increased survival in mice compared to single-agent treatments, supporting the idea that FTO inhibition has immune modulatory functions. Finally, the authors performed a proof-of-principle experiment that CS1 and CS2 also have anti-proliferative effects in other tumor types, like breast cancer, indicating that inhibition of FTO might have broader anti-tumor effects. In summary, Su et al. developed two new small-molecule inhibitors of FTO and demonstrated their therapeutic effect in vivo. Moreover, the authors showed that FTO inhibition can influence stem cell self-renewal and T cell cytotoxicity. While Su et al. convincingly demonstrate that CS1 and CS2 inhibit FTO and that inhibition of FTO has potent anti-tumor effects, some mechanistic aspects remained to be defined. MYC and LILRB4 may be important downstream effectors of FTO, however, further studies will be required to determine whether other mechanisms of action also contribute to the anti-cancer effects of CS1 and CS2. More generally, it remains puzzling how "reader" and "writer" proteins with opposing functions on the same modification support cancer proliferation in the same context. Specifically, this study and others have shown that FTO is important for proliferation of AML cells (reviewed in Huang et al., 2020Huang H. Weng H. Chen J. m6A Modification in Coding and Non-coding RNAs: Roles and Therapeutic Implications in Cancer.Cancer Cell. 2020; 37: 270-288Abstract Full Text Full Text PDF PubMed Scopus (418) Google Scholar), while the writer protein METTL3 has also been reported to be a dependency in AML (Barbieri et al., 2017Barbieri I. Tzelepis K. Pandolfini L. Shi J. Millán-Zambrano G. Robson S.C. Aspris D. Migliori V. Bannister A.J. Han N. et al.Promoter-bound METTL3 maintains myeloid leukaemia by m6A-dependent translation control.Nature. 2017; 552: 126-131Crossref PubMed Scopus (588) Google Scholar, Vu et al., 2017Vu L.P. Pickering B.F. Cheng Y. Zaccara S. Nguyen D. Minuesa G. Chou T. Chow A. Saletore Y. MacKay M. et al.The N6-methyladenosine (m6A)-forming enzyme METTL3 controls myeloid differentiation of normal hematopoietic and leukemia cells.Nat. Med. 2017; 23: 1369-1376Crossref PubMed Scopus (697) Google Scholar). Additionally, METTL3 inhibitors have also shown promise in preclinical mouse studies in AML (Cully, 2019Cully M. Chemical inhibitors make their RNA epigenetic mark.Nat. Rev. Drug Discov. 2019; 18: 892-894Crossref PubMed Scopus (35) Google Scholar). Interestingly, METTL3 and FTO are both thought to act at least partially through their effects on MYC. On the one hand, METTL3-mediated MYC m6A methylation has been shown to promote MYC stability (Huang et al., 2018Huang H. Weng H. Sun W. Qin X. Shi H. Wu H. Zhao B.S. Mesquita A. Liu C. Yuan C.L. et al.Recognition of RNA N6-methyladenosine by IGF2BP proteins enhances mRNA stability and translation.Nat. Cell Biol. 2018; 20: 285-295Crossref PubMed Scopus (1066) Google Scholar) as well as promote MYC translation (Vu et al., 2017Vu L.P. Pickering B.F. Cheng Y. Zaccara S. Nguyen D. Minuesa G. Chou T. Chow A. Saletore Y. MacKay M. et al.The N6-methyladenosine (m6A)-forming enzyme METTL3 controls myeloid differentiation of normal hematopoietic and leukemia cells.Nat. Med. 2017; 23: 1369-1376Crossref PubMed Scopus (697) Google Scholar). On the other hand, FTO is thought to promote the stability of MYC mRNA through inhibition of YTHDF2-mediated RNA decay due to decreased m6A abundance on the 5' terminal and middle exons of MYC mRNA (Huang et al., 2020Huang H. Weng H. Chen J. m6A Modification in Coding and Non-coding RNAs: Roles and Therapeutic Implications in Cancer.Cancer Cell. 2020; 37: 270-288Abstract Full Text Full Text PDF PubMed Scopus (418) Google Scholar). How the same mark can confer these different outcomes in the same context remains elusive. The development of more selective and specific inhibitors, like described in Su et al.'s work, will be helpful tools to further elucidate the mechanistic role of m6A RNA modifications more broadly. This work is supported by NIH grants CA176745, CA206963, CA204639, and CA066996. S.A.A. has been a consultant and/or shareholder for Epizyme Inc, Imago Biosciences, Vitae/Allergan Pharma, Cyteir Therapeutics, C4 Therapeutics, Syros Pharmaceuticals, OxStem Oncology, Accent Therapeutics, and Mana Therapeutics. S.A.A. has received research support from Janssen, Novartis, and AstraZeneca. Targeting FTO Suppresses Cancer Stem Cell Maintenance and Immune EvasionSu et al.Cancer CellJune 11, 2020In BriefSu et al. develop two potent small-molecule inhibitors against an RNA N6-methyladenosine demethylase called FTO. FTO inhibition shows anti-tumor effects in several types of cancers in mouse models by restricting self-renewal of cancer stem cells and suppressing immune evasion. Full-Text PDF Open Archive
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