Protein Arginine Methyltransferase 5 (PRMT5) as an Anticancer Target

Subscriber access provided by Kaohsiung Medical University

Perspective

Protein Arginine Methyltransferase 5 (PRMT5) as an Anti-Cancer Target and Its Inhibitor Discovery Yuanxiang Wang, Wenhao Hu, and Yanqiu Yuan J. Med. Chem., Just Accepted Manuscript • DOI: 10.1021/acs.jmedchem.8b00598 • Publication Date (Web): 05 Jun 2018 Downloaded from http://pubs.acs.org on June 5, 2018

Just Accepted “Just Accepted” manuscripts have been peer-reviewed and accepted for publication. They are posted online prior to technical editing, formatting for publication and author proofing. The American Chemical Society provides “Just Accepted” as a service to the research community to expedite the dissemination of scientific material as soon as possible after acceptance. “Just Accepted” manuscripts appear in full in PDF format accompanied by an HTML abstract. “Just Accepted” manuscripts have been fully peer reviewed, but should not be considered the official version of record. They are citable by the Digital Object Identifier (DOI®). “Just Accepted” is an optional service offered to authors. Therefore, the “Just Accepted” Web site may not include all articles that will be published in the journal. After a manuscript is technically edited and formatted, it will be removed from the “Just Accepted” Web site and published as an ASAP article. Note that technical editing may introduce minor changes to the manuscript text and/or graphics which could affect content, and all legal disclaimers and ethical guidelines that apply to the journal pertain. ACS cannot be held responsible for errors or consequences arising from the use of information contained in these “Just Accepted” manuscripts.

is published by the American Chemical Society. 1155 Sixteenth Street N.W., Washington, DC 20036 Published by American Chemical Society. Copyright © American Chemical Society. However, no copyright claim is made to original U.S. Government works, or works produced by employees of any Commonwealth realm Crown government in the course of their duties.

Page 1 of 41 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60

Journal of Medicinal Chemistry

Protein Arginine Methyltransferase 5 (PRMT5) as an Anti-Cancer Target and Its Inhibitor Discovery Yuanxiang Wang*, Wenhao Hu, Yanqiu Yuan* School of Pharmaceutical Sciences, Sun Yat-Sen University, Guangzhou 510006, China KEYWORDS. PRMT5, anticancer target, small molecular inhibitor

ABSTRACT. PRMT5 is a major enzyme responsible for symmetric di-methylation of arginine residues on both histone and non-histone proteins, regulating many biological pathways in mammalian cells. PRMT5 has been suggested as a therapeutic target in a variety of diseases including infectious disease, heart disease and cancer. Many PRMT5 inhibitors have been discovered in the past 5 years and one entered clinical trial in 2015 for the treatment of solid tumor and mantle cell lymphoma (MCL). The aim of this review is to summarize the current understanding of the roles of PRMT5 in cancer and the discovery of PRMT5 enzymatic inhibitors. By reviewing the structure-activity-relationship (SAR) of known inhibitors of PRMT5, we hope to provide guidance for future drug designs and inhibitor optimization. Opportunities and limitations of PRMT5 inhibitors for the treatment of cancer are also discussed.

1. PRMT family Arginine methylation is a post-translational modification that widely exists in mammalian cells.1-3 It is as prevalent as phosphorylation and ubiquitination, and regulates a variety of

ACS Paragon Plus Environment

1

Journal of Medicinal Chemistry 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60

Page 2 of 41

biological processes, including transcription, cell signaling, mRNA translation, DNA damage, receptor trafficking, protein stability and pre-mRNA splicing.4-11 In human, nine members of the PRMT family carry out this modification by catalyzing the transfer of a methyl group from Sadenosylmethionine (SAM) to the guanidinium nitrogen atoms of arginine residue, releasing one equivalent of S-adenosyl-L-homocysteine (SAH). Based on the resulting forms of methylated arginines, PRMTs are grouped into three types (Figure 1), type I (PRMT1, PRMT2, PRMT3, PRMT4, PRMT6 and PRMT8) catalyzing the formation of ω-NG-monomethylarginines (MMA) and ω-NG, NG-asymmetric dimethylarginines (aDMA), type II (PRMT5 and PRMT9) catalyzing the formation of ω-NG-MMA and ω-NG, N’G-symmetric dimethylarginines (sDMA), while Type III (PRMT7) only known to catalyze the formation of ω-NG- MMA.12

Figure 1. (A) Arginine methylation reaction catalyzed by three types of PRMTs. (B) Chemical structures of SAM and SAH. The nine members of human PRMT family share a highly homologous SAM-dependent methyltransferase (MTase) domain, which is the catalytic domain, but they have different motif structures outside MTase domain (Figure 2). PRMT1 and PRMT6 contain MTase domain only, PRMT2, PRMT3, PRMT4, PRMT5, PRMT8 and PRMT9 all have N terminal motifs proceeding the catalytic domain. Both PRMT7 and PRMT9 contain a duplicated MTase domain, although


2

Page 3 of 41 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60


the C terminal one in PRMT9 is suggested to be a pseudodomain.13 It was suggested that the additional N terminal motifs in PRMTs might be involved in substrate recognition or complex formation, contributing to their nonredundant roles in different physiological processes.13,14 In PRMT2, an SH3 domain precedes the MTase domain. SH3 domain is known to interact with other proteins via SH3 binding motif.15 In PRMT5, a TIM barrel precedes the MTase domain and interacts with MEP50 to form a stable complex.16 PRMT9 has a triple TPR motif. TPR motif is known to be involved in protein-protein interactions.17 The nonredundant functions of PRMTs have clearly been shown for the two best-characterized family members, PRMT1 and PRMT4 (also known as CARM1). Genetic knockout of either gene results in accumulation of hypomethylated substrates in vivo, demonstrating that these two PRMTs cannot substitute for each other.18,19 It is possible that the various N terminal motifs in PRMTs provide main or additional recognition site for their respective substrates. However, more studies need to be done to support the claim.

Figure 2. Domain structures of human PRMTs.


3


Page 4 of 41

Despite a unique N-terminal region of variable length and distinct domain motifs, each member of the PRMT family contains a conserved SAM-dependent MTase domain, consisting of SAM-

Figure 3. (A) Sequence alignment of the MTase domains in human PRMTs. Conserved motifs, double E loop and THW residues are shown in red. Residues that are separately conserved between type I PRMTs and PRMT5 are highlighted in yellow. (B) Percent identity matrix of


4

Page 5 of 41 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60


MTase domains in human PRMTs. Numbers are colored in green-yellow-red scale with highest homology being green and lowest being red. (C) Phylogenetic tree of human PRMTs based on sequence alignment of their MTase domains. binding sequences in motif I and post-motif I, substrate-binding sequences including the double E loop (E435 and E444 for PRMT5), and the THW loop (Figure 3A).13 While the two glutamate residues are absolutely conserved in all PRMTs and are required for activity,13 conservation in other regions varies among family members. In general, type I PRMTs share higher sequence homology with each other, PRMT9 is closer to Type I in primary sequence, while PRMT5 and PRMT7 are the most distant members of the family, being also quite different from each other (Figure 3B). Careful examination around C. elegans PRMT5 SAH binding site reveals an interesting finding.20 There are four residues conserved among PRMT5 orthologs, which are Phe379, Lys385, Ser503, and Ser669. However, the corresponding residues in type I PRMTs are Met, Arg, Tyr, and His, respectively, and these four residues are conserved within type I PRMTs. Studies found that a F379M mutant of C. elegans PRMT5 could carry out both symmetric and asymmetric dimethylation of H4R3. Human PRMT5 contains the same four residues around SAM binding site (corresponding residues in human PRMT5 are marked in yellow in Figure 3A) as C. elegans PRMT5. The corresponding mutation (F327M) in human PRMT5 also resulted in the gaining of aDMA activity. The finding implies that Phe327 (human PRMT5) occupies a key position for PRMT5’s sDMA activity, which could be an important factor to consider when designing PRMT5 specific inhibitors.20 We will discuss its specific roles in PRMT5 inhibitor binding and design in later sections of this review.


5


Page 6 of 41

2. PRMT5: biochemistry and structure PRMT5 and PRMT9 are the only known mammalian enzymes capable of forming sDMA residues as type II PRMTs.13,21-23 The loss of PRMT5 activity in mouse embryo fibroblasts results in almost complete loss of sDMA, suggesting that PRMT5 is the primary sDMA-forming enzyme in these cells.13 PRMT5 is highly conserved throughout eukaryotes, with high homology in the SAM-dependent MTases domain. It was shown that human PRMT5 forms a stable complex with MEP50, a WD domain containing cytosolic protein.16 The PRMT5:MEP50 complex was able to mono- and symmetric di-methylate peptide derived from histone 4 N terminal in vitro.16 The complex consistently had a significantly higher level of MTase activity compared with PRMT5 alone, owing to higher affinities for both the peptide and SAM. A positive allosteric effect of MEP50 on the cofactor and substrate binding of PRMT5 was proposed.16 In a biochemical assay using purified PRMT5:MEP50 complex, production of the di-methylated species could not be observed until the concentration of the monomethylated peptide exceeded that of the unmethylated substrate and the concentration of monomethylated peptide produced significantly exceeded the enzyme concentration, suggesting a nonprocessive enzymatic mechanism for peptide substrates.16 This is different from partially processive mechanism described for PRMT1.24,25 The nonprocessive mechanism of PRMT5 may reflect the fact that sDMA happens on two ω-NG-arginine, and in order for the correct positioning of the second guanidinium nitrogen atom a release and rebind mechanism may be the most efficient way for catalysis.


6

Page 7 of 41 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60


Figure 4. (A) Overall structure of PRMT5:MEP50 complex (PDB:4GQB). PRMT5 catalytic domain is colored in green, N terminal TIM barrel is colored in purple, MEP50 is colored in magenta. Highlighted in orange is the linker between the N-terminal TIM barrel and C-terminal MTase catalytic domain of PRMT5. The H4 peptide is shown in stick presentation, colored in magenta, blue and red. (B) Zoomed-in view of H4R3 substrate peptide binding groove. From the crystal structure of PRMT5:MEP50 complex, the overall structure of PRMT5 is composed of a TIM-barrel at the N-terminal and a SAM-dependent MTase domain at the C terminal, separated by a short dimerization domain.20 MEP50 adopts a WD40 β-propeller structure with seven blades. PRMT5 makes extensive contacts with top surface of the WD40 domain mainly through loop structures in the TIM-barrel domain (Figure 4A).16 The determined structure and biochemical experiments suggest that human PRMT5 binds MEP50 to form a hetero-octameric complex with molecular weight around 450 kDa. The catalytic domain of PRMT5 adopts the canonical arginine MTase tertiary structure, which is similar to that of type I PRMTs, with a SAM binding domain containing the nucleotide binding Rossmann fold, followed by a β-sandwich structure involved in substrate binding. In the above crystal structure of PRMT5:MEP50 complex, the histone H4-derived substrate peptide binds in a groove on the surface of the β-barrel domain, inserting the arginine side-chain (Arg3 of the peptide) through a narrow tunnel formed by Leu312, Phe327, and Trp579 (Figure 4B).


7


Page 8 of 41

The phenylalanine residue (Phe327) that has been shown in biochemical assays to be critical in specifying sDMA activity of PRMT5 pi-stacks against the guanidinium group of the arginine residue, orienting the substrate arginine to be methylated. The peptide residues that flank the arginine form a sharp β-turn at the neck of the tunnel are stabilized by hydrogen bonding network mainly between the main-chain of the peptide substrate and protein backbone; Two highly conserved glutamate residues Glu435 and Glu444 in the active site form the so-called double-E loop. Each glutamate residue forms a pair of salt bridges with the guanidinium group of the substrate arginine (H4R3) with the ω-NG nitrogen atom poised for methyl transfer. These glutamate residues are likely involved in de-protonating and activating the ω-NG nitrogen atom.

3. PRMT5 biology and its relation to cancer Being the major sDMA MTase, PRMT5 plays indispensible roles in cell development. It was found highly expressed in ES cells and loss of PRMT5 function is early embryonic-lethal in mouse.26 PRMT5 was also reported to play an essential role in steady-state adult hematopoiesis, complete loss of PRMT5 leading to BM aplasia and lethal pancytopenia.27 PRMT5 acts its function by methylation of a myriad of nuclear and cytoplasmic substrates, including histones H3 and H4,28 Sm proteins,29 nucleolin,30 RAF proteins31, EGFR32, and so on. Histone arginine symmetric dimethylation has been implicated to work as a repression marker,28,33 together with lysine methylation marker, to regulate gene transcription.28 There is also evidence for PRMT5 working in concert with ATP-dependent chromatin remodelers28 and transcriptional repressor complexes, including SIN3A/HDAC, MBD2/NURD, and NCoR/SMRT,34 to induce epigenetic silencing. Investigation in mouse early development


8

Page 9 of 41 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60


observed PRMT5 localization in both nucleus and cytoplasm and its localization was dynamically regulated.26 A growing list of non-histone proteins have also been identified as PRMT5 substrates, whose functions vary from cell signaling to differentiation, mRNA splicing, et. al.29-32,35-37 For example, tumor suppressor p53 is methylated by PRMT5 at three distinct arginines (Arg333, Arg335, Arg337) upon DNA damage. The methylation has an important functional consequence on p53 response.36 Androgen receptor (AR) is methylated at Arg761 by PRMT5, which was suggested as a mechanism for how the ERG oncogene may coax AR towards inducing proliferation versus differentiation.37 EGFR Arg1175 is methylated by PRMT5 and Arg1175 methylation positively modulates EGF-induced EGFR trans-autophosphorylation at Tyr1173, which governs ERK activation. So arginine methylation may crosstalk with phosphorylation to co-modulate cell signalling.32 PRMT5 also associates with other cellular proteins to enable methylation of preferred substrates. It is generally accepted that MEP50 is required for protein substrate recruitment to the catalytic domain of PRMT5.38 Loss of PRMT5 triggered the loss of MEP50 protein, without changing MEP50 mRNA levels.16 This PRMT5:MEP50 complex is likely to be the core structural unit that interacts with partner proteins to form the plethora of multi-subunit complexes with discrete specificities and functions.39 pICln was reported to recruit the spliceosomal Sm proteins to the PRMT5 complex for methylation, which allows their subsequent loading onto snRNA to form small nuclear ribonucleoproteins.29,40 RioK1 functions in analogy to pICln as an adapter protein by recruiting the RNA-binding protein nucleolin to the PRMT5 complex for its sDMA.30 The identified substrates of PRMT5 are recently reviewed by A. Richters,35 and is not the focus of this review.


9


Page 10 of 41

In the last decade, PRMT5 has attracted increasing attention as an anti-cancer target. It’s overexpressed in various cancers, including glioblastoma, leukemia/lymphoma, prostate cancer, colorectal carcinoma, and is associated with poor prognosis.39 In this section, we will review its suggested roles in cancer and its potential as a therapeutic target. There is substantial preclinical justification for targeting PRMT5-driven oncogenic pathways in glioblastoma.41 Firstly, PRMT5 is overexpressed in patient-derived primary tumors as well as cell lines and its overexpression is correlated with cell growth rate and inversely with overall patient survival.42 Genetic attenuation of PRMT5 led to cell-cycle arrest, apoptosis, and loss of cell migratory activity. Global gene profiling and chromatin immunoprecipitation identified the tumor suppressor ST7 as a key gene silenced by PRMT5.43 PRMT5 inhibition limited PRMT5 recruitment to the ST7 promoter, led to restored expression of ST7 and cell growth inhibition, and enhanced survival in a preclinical glioblastoma xenograft model.43 PRMT5 was also found to be highly expressed in human lung cancer tissue samples, whereas its expression was not detectable in benign lung tissues. Silencing PRMT5 expression strongly inhibited proliferation of lung adenocarcinoma A549 cells in tissue culture as well as abolished growth of lung A549 xenografts in mice.44 In vitro and in vivo studies showed that the cell growth arrest induced by loss of PRMT5 expression was partially attributable to down-regulation of fibroblast growth factor receptor signaling.44 Treatment of lung cancer cells with PRMT5 selective inhibitors led to inhibition of the symmetrical arginine methylation of SmD3 and histones, as well as cell proliferation. Oral administration of a PRMT5 inhibitor demonstrated antitumor activity in a lung tumor xenograft model.45 Details about the specific inhibitor used in the study will be discussed in a later section.


10

Page 11 of 41 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60


During a screen for protein targets that regulate Leukemia stem cell (LSC), a reciprocal positive regulation between PRMT5 and BCR-ABL was discovered and PRMT5 was found overexpressed in Chronic Myelogenous Leukemia (CML) LSCs as well as primary CD34+ cells from CML patients.46 Furthermore, PRMT5 expression was increased by BCR-ABL during malignant transformation of human normal CD34+ cells and attenuation of PRMT5 function by either PRMT5 specific shRNA or PRMT5 selective inhibitor reduced survival, serially replating capacity, and long-term culture-initiating cells in human CML stem cells. Similarly, inhibition of PRMT5 prolonged the survival of CML mice and reduced the growth of CML LSCs in mice. In addition, pharmacological inhibition of PRMT5 reduced long-term engraftment of human CML CD34+ cells in NSI mice. As emergence of resistance becomes a serious threat to targeted therapies, strategies that combat resistance to existing therapies are the focus of many studies. As killing LSC could be the key to overcome drug tolerance, the association between PRMT5 and BCR-ABL LSC shows an exciting path to novel and better cancer therapy.46 PRMT5 was reported to promote prostate cancer cell growth by epigenetically activating transcription of the AR in prostate cancer cells.47 Knockdown of PRMT5 or inhibition of PRMT5 by a specific inhibitor reduces the expression of AR and suppresses the growth of multiple AR-positive, but not AR-negative, prostate cancer cells. Significantly, knockdown of PRMT5 in AR-positive LNCaP cells completely suppresses the growth of xenograft tumors in mice. Molecular analysis reveals that PRMT5 binds to the proximal promoter region of the AR gene and contributes mainly to the enriched sDMA of H4R3 in the same region. Mechanistically, PRMT5 is recruited to the AR promoter by its interaction with Sp1, the major transcription factor responsible for AR transcription, and forms a complex with Brg1, an ATP-dependent chromatin remodeler, on the proximal promoter region of the AR gene. Interestingly, in an effort to identify


11


Page 12 of 41

genes that selectively facilitate the growth of TMPRSS2:ERG positive PC cells, a pooled shRNA screen in TMPRSS2:ERG and AR-positive VCaP prostate cancer cells together with coimmunoprecipitation with ERG by mass spectrometry identified PRMT5 as ERG-interacting protein. PRMT5 knockdown with shRNA transduced robust growth inhibition in VCaP cells but had no growth inhibitory effects in ERG-negative 22Rv1 cells, and only minor effects in ERG negative LNCaP cells.37 It is unclear what causes the discrepancy between the above two results regarding the growth effect of PRMT5 knockdown in LNCaP cell, however, these results show that targeting PRMT5 may represent a novel approach for prostate cancer treatment by eliminating AR expression. Recently PRMT5 has gained special attention as a cancer therapeutic target. Three separate groups concluded from pooled shRNA screens48-50 followed by association studies that genetic deficiency in methylthioadenosin phosphorylase (MTAP, P = 1.17 x 10–13) and MTAP low expression (P = 5.03 X10–35) are the top features predictive of cancer cell dependence on PRMT5.48 Because of its proximity to the tumor suppressor gene CDKN2A on human chromosome 9p21, MTAP gene is deleted at high frequency in many human tumors, including 53% of glioblastomas, 26% of pancreatic cancers, among the most frequently mutated genes in cancer.48 This new finding significantly expanded the horizon of targeting PRMT5 as anti-cancer therapies. With careful design, PRMT5 inhibitor could become a highly selective therapy for patients with MTAP deletion or low expression. We will further discuss the approaches and potential pitfalls in this regard in the last section of this review.

4. PRMT5 inhibitors


12

Page 13 of 41 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60


Recently, several groups reviewed on reported inhibitors of protein methyltransferases (PMTs),51-55 including protein lysine methyltransferases (PKMTs) and PRMTs. In 2017, Richters35 reviewed the current status and relevance of PRMT5 to the progression of various diseases in detail, and also provided a brief introduction of several PRMT5 inhibitors. In this section, we will review in detail the discovery, optimization, characterization and application of reported PRMT5 inhibitors, with the hope of shedding light on future work toward developing potent PRMT5 specific inhibitors. 4.1 SAM uncompetitive inhibitors Compound 1 (EPZ015666, Figure 5A) is the first reported cell potent and orally bioavailable PRMT5 inhibitor, with an IC50 value of 22 nM in a biochemical assay.56 Profiling against 20 other

Figure 5. (A) Structures of compound 1-3. (B) Co-crystal structure of compound 1 with PRMT5:MEP50 and SAM (PDB:4X61). PRMTs and PKMTs showed that it was inactive at concentrations up to 50 µM, with the activity against PRMT9 unevaluated. In a panel of five MCL cell lines (Z-138, Maver-1, Mino, Granta519 and Jeko-1), compound 1 decreased the level of SmD3me2s in a concentration-dependent


13


Page 14 of 41

manner. It showed potent antiproliferative effects in vitro with IC50 values of 96 nM and 450 nM in Z-138 and Maver-1 cells, respectively. Taking advantage of a cellular thermal shift assay (CETSA), compound 1 was shown to specifically bind to PRMT5 in

the cell. The compound

also displayed favorable pharmacokinetic (PK) profiles in mice, with plasma clearance of 30 mL/min/Kg, volume of distribution of 1.7L/Kg after iv dosing at 2 mg/Kg, and oral bioavailability of 69% following oral administration at 10 mg/Kg. In SCID mice bearing Z-138 and Maver-1 xenografts, Compound 1 induced tumor stasis at the dose of 200 mg/kg, with the tumor growth inhibition (TGI) values of > 93%. Based on the above potency and specificity data, compound 1 is namely the first tool compound to explore biological functions of PRMT5 and test the therapeutic hypotheses of targeting PRMT5 in cancer.56 In order to understand the mechanism of action and guide further structure optimization, a cocrystal structure of compound 1 with PRMT5:MEP50 and SAM (Figure 5B) was obtained.56 It

Figure 6. Structures of compound 4-8. shows that the compound binds in the peptide-binding site, including the pocket occupied by the substrate arginine residue, in the presence of SAM. The finding is consistent with the enzymatic data, which shows that compound 1 is competitive with the peptide substrate (Ki = 5 ± 0.3 nM) and uncompetitive with SAM. Compound 1 interacts with many residues playing essential roles


14

Page 15 of 41 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60


in peptide binding or in the catalysis of methyl transfer. Firstly, a key cation-π interaction was proposed between the phenyl ring of the tetrahydroisoquinoline (THIQ) and the cofactor SAM, which was believed to contribute to the high binding affinity and selectivity of compound 1 for PRMT5. The THIQ group also forms a potential π–π stacking interaction with Phe327, a conserved residue which was proposed to decide the sDMA of PRMT5. Removing the phenyl from the THIQ would lead to loss of the above interactions and thus resulted in a closely related but inactive compound 2 (EPZ019896, Figure 5A).56 Furthermore, the THIQ is situated in a small hydrophobic pocket formed by Leu319, Tyr324, Phe327 and Trp579, in which polar functional groups or bulky moieties are not favored. For example, pyridine replacement analogue 5 and di-methoxyl substituted analogue 6 lose PRMT5 inhibitory activity (IC50 > 50 µM), compared to their parent compound 4 (IC50 = 0.326 µM, Figure 6).57 Glu435, which is the key residue for enzyme catalysis, is possibly involved in a water-mediated interaction with the tertiary

Figure 7. Structures of compound 9 and 10. nitrogen atom of the THIQ ring system of compound 1. Incorrect orientation of this interaction results in reduced potency, such as analogue 7 (IC50 = 7.45 µM, Figure 6) and 8 (IC50 = 26.8 µM), compared with compound 4. Additionally, there is a possible hydrogen bond interaction between the chiral hydroxyl of compound 1 and the side chain of Glu444, explaining the potency differences between stereoisomers 9 (IC50 = 13 nM, Figure 7) and 10 (IC50 = 95 nM). Moreover,


15


Page 16 of 41

the terminal pyrimidine group of compound 1 is poised for π–π stacking interactions with Phe327 and Phe580, likely contributing to its high inhibitory potency against PRMT5. All above interactions prompt compound 1 to adopt a “U” shaped conformation while binding to PRMT5, and ensure its high affinity and selectivity for PRMT5. Notably, the improved compound 3 (EPZ015938, Figure 5A), which was licensed to GSK (GSK3326595), has already entered Phase 1 clinical trial to assess the safety, PK, and pharmacodynamics (PD) in patients with solid tumors and non-Hodgkin lymphoma (NCT02783300).58

Figure 8. Structures of compound 11-13. By screening a library of 350000 compounds using a Transcreener EPIGENTM technology, Falk and coworkers59 also found a series of SAM uncompetitive inhibitors, with a similar core structure to compound 1. Further optimization led to compound 11 (Figure 8). It exhibits potent PRMT5 enzyme inhibitory activity with an IC50 value of 109 nM and is highly selective for PRMT5. Compound 11 effectively blocks the arginine sDMA of both histone and non-histone substrates in TE-1 cells. Decrease of the H4R3me2s mark in bone marrow cells and peripheral white blood cells of C57BL/6 mice was observed after treatment with compound 11 in a dose-


16

Page 17 of 41 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60


dependent manner and no obvious weight loss was observed. In order to develop more potent PRMT5 inhibitors, diversity of the THIQ group was explored. Moving the alcohol scaffolding motif from the N-position of THIQ to the C3 position provided a new series of analogues with one methylene deletion, such as compound 12 (Figure 8). It is highly potent againt PRMT5, with an IC50 of 26 nM.60 Taking the linear amine group out of the THIQ motif produced analogues featuring a dihydro-indene ring. The optimized compound 13 (Figure 8) possesses similar potency (IC50 = 105 nM) to compound 11.61

Figure 9. Structures of compound 14-16. As depicted in the co-crystal structure of 1 with PRMT5:MEP50 and SAM, PRMT5 possesses two ligand binding pockets: a SAM-binding site and a substrate peptide binding site. Since the substrate-binding sites vary among PRMTs, a virtual screening against substrate-binding pocket could lead to discovery of inhibitors with good selectivity. Based on this theory, Zheng and coworkers62 discovered a novel scaffold PRMT5 inhibitor 14 (Figure 9) by molecular docking and confirmed by 3H-labeled methylation assays. Compound 14 displays 69% inhibitory activity against PRMT5 at the concentration of 50 µM. By analyzing its predictive interactions with PRMT5, it was found that the phenyl group may form π–π stacking interaction with Phe327 and Phe580, similarly to the terminal pyrimidine of compound 1. Removal of the phenyl group afforded compound 15 (Figure 9), which lost PRMT5 inhibitory activity (2% inhibition at 50


17


Page 18 of 41

µM), confirming the contribution of the π–π stacking interaction. Compound 14, however, does not make the cation-π interaction with SAM nor the π–π stacking interaction with Phe327, and has only moderate potency against PRMT5 (IC50 = 35.6 µM). Since these two interactions are critical to the outstanding affinity and selectivity of compound 1 for PRMT5, the result for compound 14 could be expected. Introducing a phenyl to the second chain of 14 restored the interactions and produced compound 16 (Figure 9) with 12 times improved activity (IC50 = 2.8 µM), while it is inactive against PRMT1, EZH2 and DNMT3A. Compound 16 displays potent anti-proliferative activities against Z-138, Maver-1, and Jeko-1 cancer cells with EC50 values of 12.7 µM, 12.7 µM, 10.5 µM, respectively.

Figure 10. (A) Structure of compound 17 and 18. (B) Co-crystal structure of compound 18 with PRMT5: MEP50 in complex with H4 peptide (PDB:4GQB). 4.2. SAM competitive inhibitors PRMTs contain highly conserved SAM-binding pockets, it is not surprising that compound 1763 (Sinefungin, a natural product, Figure 10A) and SAH64, as close analogs of cofactor SAM, exhibit excellent PRMT5 inhibitory activity, but lack of selectivity against other protein methyl transferases. In order to explore the binding mode of these inhibitors with PRMT5 and discover


18

Page 19 of 41 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60


more selective inhibitors, the first crystal structure of human PRMT5:MEP50 in complex with H4 peptide and Compound 18 (dehydrosinefungin, Figure 10B) was published by Antonysamy16 in 2012. As shown in Figure 10B, compound 18 could interact with several residues around SAM binding pocket. The backbone NH of Met420 and the side chain of Asp419 may interact with adenosine moiety via hydrogen bonds. The side chains of Glu392 and Tyr324 form two potential hydrogen bonds with two hydroxy groups of 18. In addition, the NH of allylic amine possibly interacts with guanidinium group of arginine residue located on the H4 peptide substrate (H4R3) via a hydrogen bond. Of particular note, the guanidinium group of H4R3 accepting a methyl group from SAM, is amid of a hydrogen bond network with the side-chains of the conserved “PRMT double-E loop” Glu435 and Glu444. At last, the tail acid group of 18 might be involved in hydrogen bond interactions with residue Tyr334. These interactions anchor compound 18 to the SAM binding pocket. As described above, the SAM binding site is conserved in PRMTs, compound 18 maintains the core interactions within the SAM binding site, but lacks unique interactions with PRMT5, so it is still a pan-protein methyl transferases inhibitor and is not selective toward PRMT5.

Figure 11. Structures of compound 19-21. By replacing the amino-acid end of SAH with ethyl urea group to recapitulate hydrogen bond interactions between the guanidinium group of H4R3 residue and E444, Schapira and coworkers65 designed and prepared compound 19 (DS-437, Figure 11). It inhibits PRMT5:MEP50 complex and PRMT7 with an IC50 value of 6 µM, while it is inactive against 29


19


Page 20 of 41

other human protein-, DNA-, and RNA-MTases. It could also reduce the symmetrical arginine dimethylation of SmD1/3 and SmB proteins in breast cancer cells, representing a new scaffold to investigate the therapeutic potential for targeting the PRMT5-PRMT7 pathway. A potent and selective PRMT5 inhibitor, compound 20 (LLY-283, Figure 11),66 was discovered by Eli Lily. Considering the chemical core structure similarity, compound 20 is still an analogue of SAM, with the methionine moiety replaced by a phenyl group. Compound 20 exhibits potent PRMT5 enzyme inhibitory activity with an IC50 value of 20 nM. It also displays more than 100fold selectivity for PRMT5 over other methyl transferases and nonepigenetic targets. Methylation of SmBB’ is inhibited by 20 with an IC50 of 25 nM in MCF7 cells, while MDM4 splicing is inhibited with an IC50 of 40 nM in A375 cells. Recently, the crystal structure of the PRMT5:MEP50 complex with compound 20 was determined.66 Comparing with the crystal structure of compound 18 and PRMT5:MEP50 complex, the adenine and ribose moieties of compound 20 adopt very similar conformations and make similar interactions with Asp419, Met420, Glu392 and Try324. But the phenyl group of compound 20 occupies the position of the side chain of Phe327, a residue specific to PRMT5. This change may contribute to the high selectivity of compound 20 for PRMT5. Aiming at developing SAM analogues as selective PRMT5 inhibitors with improved PK profile and oral bioavailability, researchers in Janssen displaced the tetrahydrofuran ring of SAM with cyclopentane, and developed a new series of PRMT5 inhibitors, compound 21 being one example with IC50 value for PRMT5 at 6.3 nM.67 R N H N

N H

N N

22 (CMP-5)

N H

O

23 (HLCL-61) R = H 24 (HLCL-65) R = Br

N 25 (PJ-68)


20

Page 21 of 41 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60


Figure 12. Structures of compound 22-25. A series of N-alkyl-9H-carbazole analogs as PRMT5 inhibitors were discovered by Baiocchi and coworkers68 by employing a human PRMT5 catalytic site model and virtual screening of a 10000-compound library. Taking a process of structure-based computational analysis and combinatorial lead optimization, the group obtained compound 22 (CMP5, Figure 12), the lead candidate capable of blocking EBV-driven B-lymphocyte transformation and survival, while keeping normal B cells unaffected. Compound 22 selectively inhibits the formation of sDMA on H4R3, but it is inactive against type I (PRMT1, PRMT4) and Type III (PRMT7) enzymes. A model of compound 22 binding to PRMT5 show that the pyridine ring possibly forms π–π stacking interactions with Phe327 residue, while the carbazole ring of CMP5 is orientated toward the adenosine binding site. It is important to point out that PRMT5 enzymatic inhibitory activity of compound 22 was not reported in the original reference. Until recently Zhang and coworkers showed that 22 displayed 7.8% inhibitory activity against PRMT5 at 50 µM using radioactive methylation assay.69 After the second-round optimization based on the core structure of 22, compound 23 (HLCL-61, Figure 12)70 was developed as a selective PRMT5 inhibitor by replacing the pyridine group of 22 with an ortho-methoxyphenyl group. It was proposed that this change not only reserved the π–π stacking interaction with Phe327, but also introduced a new hydrogen bond interaction with the protein residue. Compound 23 maintains its specificity for PRMT5 and effectively inhibits symmetric arginine dimethylation of histones H3 and H4 in AML samples. Treatment of AML cell lines (MV4-11 and THP-1) and primary blasts with 23 showed decreased cell viability with an IC50 of 7.21–21.46 µM for cell lines and 3.98– 8.72µM for patient samples. Additionally, the effect of introducing a halogen atom to the carbazole ring of 23 was explored. A halogen bond can be formed between a halogen atom and a negative site


21


Page 22 of 41

or the π electrons of an unsaturated system (O, N, S, or π) from the surrounding residues,71 analogous to hydrogen bond interactions.72 As expected, introduction of bromine to the carbazole of 23 produced a more selective and potent PRMT5 inhibitor, compound 24 (HLCL-65, Figure 12).73 It selectively inhibits the formation of sDMA mediated by PRMT5 and demonstrates potent antiproliferative effect against Th1 cells (IC50 = 1.1 µM) and Th2 cells (IC50 = 4.0 µM). To study PRMT5 functions in chronic myelogenous leukemia, Pan and coworkers46 screened and discovered a potent small molecule PRMT5 inhibitor, compound 25 (PJ-68, Figure 12), which employed the core structure of N-ethyl-9H-carbazole. Compound 25 inhibits PRMT5 with an IC50 of 517 nM, while having no effect on the activity of type I PRMT family members (PRMT1, 3, 4, 6, and 8), indicating its specificity for PRMT5. It inhibits the formation of sDMA on histone H2A and H4 in K562 cells in a concentration-dependent manner and significantly decreases the level of BCR-ABL in both CD34+CD38- cells and CD34+CD38+ cells. PK study in SD rats showed that plasma concentrations were 3.57 ± 0.08 µM (1,264.4 ± 28.4 ng/ml) and 7.88 ± 0.31 µM (2,788.5 ± 109.7 ng/ml), respectively, after a single oral or intravenous administration, which were supposed to be sufficient to block PRMT5 activity in vitro.

Figure 13. Structures of compounds 26-31. Recently, by taking advantage of structure-based virtual screening, Luo and coworkers74 discovered a new scaffold of PRMT5 inhibitors, as exemplified by compound 26 (Figure 13) with the IC50 of 0.33 µM. Compound 26 displays a broad selectivity against a panel of MTases. It binds directly to PRMT5 as validated by SPR, with an equilibrium


22

Page 23 of 41 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60


dissociation constant (KD) of 0.987 µM. Compound 26 exhibits competitive inhibition with respect to SAM againt PRMT5 and shows noncompetitive inhibition with respect to peptide substrate, suggesting a SAM competitive PRMT5 inhibitor. Compound 26 was optimized from hit compound 27 (IC50 = 8.1 µM, Figure 13) by introducing a methoxy group to the 5-position of benzimidazole. Molecular docking predicted that the methoxy group would form a hydrogen bond with Tyr33469 and improve inhibitory activity from compound 26. It was also predicted that the methoxycarbonyl substituted phenyl was close to a pocket occupied by the adenosine group of SAM. A hydrogen bond interaction was predicted between the carbonyl oxygen of the methoxycarbonyl group and Lys393,69 a critical interaction determining the high affinity of compound 13 to PRMT5. Analogues such as compound 28-30 without this potential interaction are much weaker binders (Figure 13). Unlike the adenosine group of SAM forming two potential hydrogen bonds with Met420 and Asp419, compound 26 does not engage these interactions. It can be expected that restoring these interactions may further improve its inhibitory activity. To mimic the hydrogen bond interactions between the guanidinium group of substrate arginine residue and “the double E loop” (E435 and E444, Figure 1), Yang and coworkers75 developed a series of new PRMT5 inhibitors. They firstly screened guanidinium analogues, such as 1,2diamines, benzimidamide, semicarbazone, and amines. Then by performing molecular dockingbased virtual screening, a highly selective PRMT5 inhibitor 31 (P5i-6, Figure 13) was discovered. It was based on the fragment of hydrazinecarboxamide. This compound could form multiple hydrogen bonds with residues E435, E444 and L437 in its predicted binding site. Compound 31 displays potent inhibitory activity against PRMT5 with an IC50 value of 0.57 µM, and is inactive against PRMT1, 3, 4, 6, 7 and 8. Further study showed that the compound was active against two colorectal cancer cell lines (HT-29 and DLD-1) and one hepatic cancer cell


23


Page 24 of 41

line (HepG2), and that it selectively inhibited the formation of sDMA on H4R3 and H3R8 in DLD-1 cells. However, it was uncertain whether it was a SAM cooperative inhibitor or a SAM competitive one without kinetic experiment data.

5. Perspectives In the last 5 years, a significant number of chemical structures have been discovered to be PRMT5 inhibitors, as we discussed above, and several typical ones were summarized and compared in Table 1. SAM uncompetitive inhibitors inhibit PRMT5 by occupying the peptide binding pocket in the presence of SAM. This binding mode may offer great advantages. Cellular concentration of SAM is reported to be in several hundred µM range. With PRMT5’s Km for SAM Table 1. Comparison of typical PRMT5 inhibitors compd (no)

binding site

selectivity for PRMT5

1

substrate-binding site

high

16

substrate-binding site

18

MOA

biochemical activity (nM)

cellular activity (nM)/cell line

IC50 = 22

good

SAM-uncompetitive inhibition; substrate-competitive inhibition __

SAM-binding site

no

SAM-competitive inhibition

IC50 = 35

20

SAM-binding site

remarkable

IC50 = 20

IC50 = 25/MCF7 IC50 = 40/A375

24

SAM-binding site

good

SAM-noncompetitive inhibition; substrate-noncompetitive inhibition __

IC50 = 517

26

SAM-binding site

broad

SAM-competitive inhibition; substrate-noncompetitive inhibition

IC50 = 330

IC50 = 1100/Th1 IC50 = 4000/Th2 EC50 = 6530/MV4-11

IC50 = 2800

IC50 = 96/Z-138 IC50 = 450/Maver-1 EC50 = 12700/Z-138 EC50 = 12700/Maver-1 EC50 = 10500/Jeko-1 __

in low µM and structural information pointing to sequential bi-bi mechanism with SAM binding first, majority of the enzyme exists in the SAM complexed form. Kinetically it is advantageous to directly inhibit the major enzyme form. Compound 3 with this mode of action has already progressed into clinical trial for the treatment of MCL and solid tumor, based on preclinical


24

Page 25 of 41 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60


activity on several MCL cell lines, good selectivity profile and animal toxicity results. However, studies reported by Novartis and Agios Inc. raised some concerns regarding this mode of inhibition.48, 49 Both groups were investigating the association between cancer cell dependence on PRMT5 function and MTAP genetic status. They indicated that Compound 1 effectively inhibited in vitro growth of many selected cell lines, however, it does so regardless of MTAP status, nor the results correlates with PRMT5 expression level. Since loss of PRMT5 is known to be embryonic lethal, these findings raised the question if lack of selectivity in cancer cell lines would translate to greater toxicity later in clinical trials. It was shown by Sellers et. al. that MTAP deficient cell lines accumulated metabolic molecule methylthioadenosine (MTA), which specifically inhibits PRMT5 in vitro, and it was suggested that PRMT5 in these cells were indeed partially inhibited by MTA, rendering cell lines especially sensitive to further PRMT5 attenuation. If that’s true, then in MTAP deficient cells, PRMT5 is partially complexed with MTA, which is different from PRMT5:SAM complex in normal cells.This offers a unique opportunity to target PRMT5 in these diseased cells. One strategy involves designing or screening for small molecules that could synergistically bind to PRMT5 with MTA and drive the whole PRMT5 population to inactive state, thus inhibiting cell growth.. For the SAM competitive binding mode, several compounds showed selective inhibition of PRMT5, albeit the highly conserved SAM binding site within the PRMT family and other MTase domain containing proteins. So it is possible to achieve high selectivity among MTases with SAM mimetics. However, PK could be a potential problem with SAM mimetics and thus should be taken into consideration when developing this type of inhibitors into therapies. In addition, care must be taken to develop any PRMT5 inhibitors into clinical use. Experiments with conditional knockout of prmt5 in adult mice demonstrated the essential role of PRMT5 in


25


Page 26 of 41

the maintenance of adult hematopoietic cells.27 PRMT5-deficient hematopoietic stem and progenitor cells exhibited severely impaired cytokine signaling as well as upregulation of p53 and expression of its downstream targets, resulting in a rapid and profound effect on blood cell production. Treating cancer patients with PRMT5 inhibitors could potentially result in significant myelosuppressive side effects. Nonetheless, the clinical activity and side effects of chemical inhibitors of PRMT5 will need to be defined in the clinic. In conclusion, targeting PRMT5 offers a great opportunity to combat cancer in an efficient and selective way. However, more biology regarding PRMT5 function need to be revealed for better understanding of its therapeutic potentials and more potent and selective PRMT5 inhibitors need to be developed to validate the PRMT5 as an anti-cancer target.

AUTHOR INFORMATION Corresponding Author *For Y.W.: E-mail, [email protected]. Fax, +86-20-39943004. *For Y.Y.: E-mail, [email protected]. Fax, +86-20-39943004. ORCID Yuanxiang Wang: 0000-0001-5650-4396 Author Contributions The manuscript was written through contributions of all authors. All authors have given approval to the final version of the manuscript. Notes The authors declare no competing financial interest.


26

Page 27 of 41 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60


Biographies Yuanxiang Wang is currently an Associate Professor at School of Pharmaceutical Sciences, Sun Yat-Sen University (SYSU). He received a Ph.D. degree in Medicinal Chemistry from Shanghai Institute of Materia Medica (SIMM), Chinese Academy of Sciences (CAS) in 2012. Dr. Wang completed postdoctoral training at the University of Arizona and the University of Utah from 2012 to 2017. In 2017, Dr. Wang joined SYSU as a faculty member. His research interests include clinical directed anticancer drug design and discovery, and development of new synthetic methodologies for creating peptide therapeutics. Wenhao Hu is currently a Full Professor and Dean of School of Pharmaceutical Sciences at SYSU. He received his Ph.D. degree from Hong Kong Polytechnic University with Professor Albert S. C. Chan in 1998. He then became a postdoctoral fellow at the University of Arizona with Professor Michael P. Doyle. In 2002-2006, he worked as a medicinal chemist at Genesoft in San Francisco and a process chemist at Bristol-Myers Squibb in New Jersey. In 2006 he joined East China Normal University, where he was a professor of Chemistry, and head of Shanghai Engineering Research Center of Molecular Therapeutics and New Drug Development. He moved to SYSU in August 2016. His research interests include synthetic methodology, multicomponent reactions, medicinal chemistry and process chemistry. Yanqiu Yuan is currently An Associate Professor at School of Pharmaceutical Sciences, SYSU. She received her Ph.D. degree at Harvard University in 2008, subsequently worked at Novartis Institute for Biomedical Research at Cambridge as Investigator and University of Copenhagen as Assistant Professor. Her research interests include the biochemical function of PRMT5 and discovery of small molecule inhibitors as anti-cancer therapy. ACKNOWLEDGMENT


27


Page 28 of 41

We acknowledge startup fundings from SYSU (36000-18831104 to Y.W.; 36000-18831102 to Y.Y.) and Guangdong Innovative and Entrepreneurial Research Team Program (No. 2016ZT06Y337 to W.H.) for financial support. ABBREVIATIONS USED PRMT5, Protein Arginine Methyltransferase 5; SAM, S-adenosylmethionine; SAH, S-adenosylL-homocysteine; aDMA, asymmetric dimethylarginine; sDMA, symmetric dimethylarginine; MMA, monomethylarginine; MTase, methyltransferase; AR, Androgen receptor; LSC, Leukemia stem cell; CML, Chronic Myelogenous Leukemia; MTAP, methylthioadenosin phosphorylase;

PMTs,

protein

lysine

methyltransferases;

PKMTs,

protein

lysine

methyltransferases; MCL, Mantel Cell Lymphoma; CETSA, cellular thermal shift assays; TGI, tumor

growth

inhibition;

THIQ,

tetrahydroisoquinoline;

PK,

pharmacokinetic;

PD,

pharmacodynamics; MTA, methylthioadenosine. REFERENCES (1) Peng, C.; Wong, C. C. L. The story of protein arginine methylation: characterization, regulation, and function. Expert Rev. Proteomics 2017, 14, 157-170. (2) Larsen, S. C.; Sylvestersen, K. B.; Mund, A.; Lyon, D.; Mullari, M.; Madsen, M. V.; Daniel, J. A.; Jensen, L. J.; Nielsen, M. L. Proteome-wide analysis of arginine monomethylation reveals widespread occurrence in human cells. Sci. Signal. 2016, 9, rs9. (3) Guo, A. L.; Gu, H. B.; Zhou, J.; Mulhern, D.; Wang, Y.; Lee, K. A.; Yang, V.; Aguiar, M.; Kornhauser, J.; Jia, X. Y.; Ren, J. M.; Beausoleil, S. A.; Silva, J. C.; Vemulapalli, V.; Bedford, M. T.; Comb, M. J. Immunoaffinity enrichment and mass spectrometry analysis of protein methylation. Mol. Cell. Proteomics 2014, 13, 372-387.


28

Page 29 of 41 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60


(4) McBride, A. E.; Silver, P. A. State of the Arg: Protein methylation at arginine comes of age. Cell 2001, 106, 5-8. (5) Auclair, Y.; Richard, S. The role of arginine methylation in the DNA damage response. DNA Repair 2013, 12, 459-465. (6) Blanc, R. S.; Richard, S. Arginine Methylation: The coming of age. Mol. Cell 2017, 65, 824. (7) Wesche, J.; Kuhn, S.; Kessler, B. M.; Salton, M.; Wolf, A. Protein arginine methylation: a prominent modification and its demethylation. Cell. Mol. Life Sci. 2017, 74, 3305-3315. (8) Jahan, S.; Davie, J. R. Protein arginine methyltransferases (PRMTs): role in chromatin organization. Adv. Biol. Regul. 2015, 57, 173-184. (9) Mowen, K. A.; David, M. Unconventional post-translational modifications in immunological signaling, Nat. Immunol. 2014, 15, 512-520. (10) Wei, H.; Mundade, R.; Lange, K. C.; Lu, T. Protein arginine methylation of non-histone proteins and its role in diseases. Cell Cycle 2014, 13, 32-41. (11) Blackwell, E.; Ceman, S. Arginine methylation of RNA-binding proteins regulates cell function and differentiation. Mol. Reprod. Dev. 2012, 79, 163-175. (12) Zurita-Lopez, C. I.; Sandberg, T.; Kelly, R.; Clarke, S. G. Human protein arginine methyltransferase 7 (PRMT7) is a type III enzyme forming omega-NG-monomethylated arginine residues. J. Biol. Chem. 2012, 287, 7859-7870.


29


Page 30 of 41

(13) Hadjikyriacou, A.; Yang, Y. Z.; Espejo, A.; Bedford, M. T.; Clarke, S. G. Unique features of human protein arginine methyltransferase 9 (PRMT9) and its substrate RNA splicing factor SF3B2. J. Biol. Chem. 2015, 290, 16723-16743. (14) Bedford, M. T. Arginine methylation at a glance. J. Cell Sci. 2007, 120, 4243-4246. (15) Kurochkina, N.; Guha, U. SH3 domains: modules of protein-protein interactions. Biophysical reviews 2013, 5, 29-39 (16) Antonysamy, S.; Bonday, Z.; Campbell, R. M.; Doyle, B.; Druzina, Z.; Gheyi, T.; Han, B.; Jungheim, L. N.; Qian, Y. W.; Rauch, C.; Russell, M.; Sauder, J. M.; Wasserman, S. R.; Weichert, K.; Willard, F. S.; Zhang, A. P.; Emtage, S. Crystal structure of the human PRMT5:MEP50 complex. P. Natl. Acad. Sci. U. S. A. 2012, 109, 17960-17965. (17) Zeytuni, N.; Zarivach, R. Structural and functional discussion of the tetra-trico-peptide repeat, a protein interaction module. Structure 2012, 20, 397-405. (18) Kim, J.; Lee, J.; Yadav, N.; Wu, Q.; Carter, C.; Richard, S.; Richie, E.; Bedford, M. T. Loss of CARM1 results in hypomethylation of thymocyte cyclic AMP-regulated phosphoprotein and deregulated early T cell development. J. Biol. Chem. 2004, 279, 25339-25344. (19) Pawlak, M. R.; Banik-Maiti, S.; Pietenpol, J. A.; Ruley, H. E. Protein arginine methyltransferase I: substrate specificity and role in hnRNP assembly. J. Cell. Biochem. 2002, 87, 394-407. (20) Sun, L. T.; Wang, M. Z.; Lv, Z. Y.; Yang, N.; Liu, Y. F.; Bao, S. L.; Gong, W. M.; Xu, R. M. Structural insights into protein arginine symmetric dimethylation by PRMT5. P. Natl. Acad. Sci. U. S. A. 2011, 108, 20538-20543.


30

Page 31 of 41 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60


(21) Branscombe, T. L.; Frankel, A.; Lee, J. H.; Cook, J. R.; Yang, Z. H.; Pestka, S.; Clarke, S. PRMT5 (Janus kinase-binding protein 1) catalyzes the formation of symmetric dimethylarginine residues in proteins. J. Biol. Chem. 2001, 276, 32971-32976. (22) Cook, J. R.; Lee, J. H.; Yang, Z. H.; Krause, C. D.; Herth, N.; Hoffmann, R.; Pestka, S. FBXO11/PRMT9, a new protein arginine methyltransferase, symmetrically dimethylates arginine residues. Biochem. Biophys. Res. Commun. 2006, 342, 472-481. (23) Yang, Y. Z.; Hadjikyriacou, A.; Xia, Z.; Gayatri, S.; Kim, D.; Zurita-Lopez, C.; Kelly, R.; Guo, A. L.; Li, W.; Clarke, S. G.; Bedford, M. T. PRMT9 is a Type II methyltransferase that methylates the splicing factor SAP145. Nat. Commun. 2015, 6, 6428. (24) Obianyo, O.; Osborne, T. C.; Thompson, P. R. Kinetic mechanism of protein arginine methyltransferase 1. Biochemistry 2008, 47, 10420-10427. (25) Osborne, T. C.; Obianyo, O.; Zhang, X.; Cheng, X.; Thompson, P. R. Protein arginine methyltransferase 1: Positively charged residues in substrate peptides distal to the site of methylation are important for substrate binding and catalysis. Biochemistry 2007, 46, 1337013381. (26) Tee, W. W.; Pardo, M.; Theunissen, T. W.; Yu, L.; Choudhary, J. S.; Hajkova, P.; Surani, M. A. Prmt5 is essential for early mouse development and acts in the cytoplasm to maintain ES cell pluripotency. Genes Dev. 2010, 24, 2772-2777. (27) Liu, F.; Cheng, G. Y.; Hamard, P. J.; Greenblatt, S.; Wang, L.; Man, N.; Perna, F.; Xu, H. M.; Tadi, M.; Luciani, L.; Nimer, S. D. Arginine methyltransferase PRMT5 is essential for sustaining normal adult hematopoiesis. J. Clin. Invest. 2015, 125, 3532-3544.


31


Page 32 of 41

(28) Pal, S.; Vishwanath, S. N.; Erdjument-Bromage, H.; Tempst, P.; Sif, S. Human SWI/SNFassociated PRMT5 methylates histone H3 arginine 8 and negatively regulates expression of ST7 and NM23 tumor suppressor genes. Mol. Cell. Biol. 2004, 24, 9630-9645. (29) Pesiridis, G. S.; Diamond, E.; Van Duyne, G. D. Role of pICLn in methylation of Sm proteins by PRMT5. J. Biol. Chem. 2009, 284, 21347-21359. (30) Guderian, G.; Peter, C.; Wiesner, J.; Sickmann, A.; Schulze-Osthoff, K.; Fischer, U.; Grimmler, M. RioK1, a new interactor of protein arginine methyltransferase 5 (PRMT5), competes with pICln for binding and modulates PRMT5 complex composition and substrate specificity. J. Biol. Chem. 2011, 286, 1976-1986. (31) Andreu-Perez, P.; Esteve-Puig, R.; de Torre-Minguela, C.; Lopez-Fauqued, M.; BechSerra, J. J.; Tenbaum, S.; Garcia-Trevijano, E. R.; Canals, F.; Merlino, G.; Avila, M. A.; Recio, J. A. Protein arginine methyltransferase 5 regulates ERK1/2 signal transduction amplitude and cell fate through CRAF. Sci. Signal. 2011, 4, ra58. (32) Hsu, J. M.; Chen, C. T.; Chou, C. K.; Kuo, H. P.; Li, L. Y.; Lin, C. Y.; Lee, H. J.; Wang, Y. N.; Liu, M.; Liao, H. W.; Shi, B.; Lai, C. C.; Bedford, M. T.; Tsai, C. H.; Hung, M. C. Crosstalk between Arg 1175 methylation and Tyr 1173 phosphorylation negatively modulates EGFR-mediated ERK activation. Nat. Cell Biol. 2011, 13, 174-151. (33) Xu, X. J.; Hoang, S.; Mayo, M. W.; Bekiranov, S. Application of machine learning methods to histone methylation ChIP-Seq data reveals H4R3me2 globally represses gene expression. BMC Bioinf. 2010, 11, 396.


32

Page 33 of 41 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60


(34) Karkhanis, V.; Hu, Y. J.; Baiocchi, R. A.; Imbalzano, A. N.; Sif, S. Versatility of PRMT5induced methylation in growth control and development. Trends Biochem. Sci. 2011, 36, 633641. (35) Richters, A. Targeting protein arginine methyltransferase 5 in disease. Future Med. Chem. 2017, 9, 2081-2098. (36) Jansson, M.; Durant, S. T.; Cho, E. C.; Sheahan, S.; Edelmann, M.; Kessler, B.; La Thangue, N. B. Arginine methylation regulates the p53 response. Nat. Cell Biol. 2008, 10, 14314139. (37) Mounir, Z.; Korn, J. M.; Westerling, T.; Lin, F.; Kirby, C. A.; Schirle, M.; McAllister, G.; Hoffman, G.; Ramadan, N.; Hartung, A.; Feng, Y.; Kipp, D. R.; Quinn, C.; Fodor, M.; Baird, J.; Schoumacher, M.; Meyer, R.; Deeds, J.; Buchwalter, G.; Stems, T.; Keen, N.; Sellers, W. R.; Brown, M.; Pagliarini, R. A. ERG signaling in prostate cancer is driven through PRMT5dependent methylation of the Androgen Receptor. Elife 2016, 5, e13964. (38) Burgos, E. S.; Wilczek, C.; Onikubo, T.; Bonanno, J. B.; Jansong, J.; Reimer, U.; Shechter, D. Histone H2A and H4 N-terminal tails are positioned by the MEP50 WD repeat protein for efficient methylation by the PRMT5 arginine methyltransferase. J. Biol. Chem. 2015, 290, 9674-9689. (39) Stopa, N.; Krebs, J. E.; Shechter, D. The PRMT5 arginine methyltransferase: many roles in development, cancer and beyond. Cell. Mol. Life Sci. 2015, 72, 2041-2059.


33


Page 34 of 41

(40) Meister, G.; Eggert, C.; Buhler, D.; Brahms, H.; Kambach, C.; Fischer, U. Methylation of Sm proteins by a complex containing PRMT5 and the putative U snRNP assembly factor pICln. Curr. Biol. 2001, 11, 1990-1994. (41) Banasavadi-Siddegowda, Y. K.; Welker, A. M.; Imitola, J.; Li, C. L.; Zhang, J. Y.; Beattie, C. E.; Baiocchi, R.; Kaur, B. Prmt5 as a druggable target for glioblastoma therapy. Neuro. Oncol. 2017, Doi: 10.1093/neuonc/nox206. (42) Han, X. S.; Li, R.; Zhang, W. B.; Yang, X. H.; Wheeler, C. G.; Friedman, G. K.; Province, P.; Ding, Q.; You, Z. Y.; Fathallah-Shaykh, H. M.; Gillespie, G. Y.; Zhao, X. Y.; King, P. H.; Nabors, L. B. Expression of PRMT5 correlates with malignant grade in gliomas and plays a pivotal role in tumor growth in vitro. J. Neurooncol. 2014, 118, 61-72. (43) Yan, F. T.; Alinari, L.; Lustberg, M. E.; Martin, L. K.; Cordero-Nieves, H. M.; Banasavadi-Siddegowda, Y.; Virk, S.; Barnholtz-Sloan, J.; Bell, E. H.; Wojton, J.; Jacob, N. K.; Chakravarti, A.; Nowicki, M. O.; Wu, X.; Lapalombella, R.; Datta, J.; Yu, B.; Gordon, K.; Haseley, A.; Patton, J. T.; Smith, P. L.; Ryu, J.; Zhang, X. L.; Mo, X. K.; Marcucci, G.; Nuovo, G.; Kwon, C. H.; Byrd, J. C.; Chiocca, E. A.; Li, C. L.; Sif, S.; Jacob, S.; Lawler, S.; Kaur, B.; Baiocchi, R. A. Genetic validation of the protein arginine methyltransferase PRMT5 as a candidate therapeutic target in glioblastoma. Cancer Res. 2014, 74, 1752-1765. (44) Gu, Z. P.; Gao, S.; Zhang, F. H.; Wang, Z. Q.; Ma, W. C.; Davis, R. E.; Wang, Z. X. Protein arginine methyltransferase 5 is essential for growth of lung cancer cells. Biochem. J. 2012, 446, 235-241.


34

Page 35 of 41 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60


(45) Kong, G. M.; Yu, M.; Gu, Z. P.; Chen, Z.; Xu, R. M.; O'Bryant, D.; Wang, Z. X. Selective small-chemical inhibitors of protein arginine methyltransferase 5 with anti-lung cancer activity. PLoS One 2017, 12, e0181601. (46) Jin, Y.; Zhou, J.; Xu, F.; Jin, B.; Cui, L.; Wang, Y.; Du, X.; Li, J.; Li, P.; Ren, R.; Pan, J. Targeting methyltransferase PRMT5 eliminates leukemia stem cells in chronic myelogenous leukemia. J. Clin. Invest. 2016, 126, 3961-3980. (47) Deng, X.; Shao, G.; Zhang, H. T.; Li, C.; Zhang, D.; Cheng, L.; Elzey, B. D.; Pili, R.; Ratliff, T. L.; Huang, J.; Hu, C. D. Protein arginine methyltransferase 5 functions as an epigenetic activator of the androgen receptor to promote prostate cancer cell growth. Oncogene 2017, 36, 1223-1231. (48) Mavrakis, K.; McDonald, E. R.; Schlabach, M. R.; Billy, E.; Hoffman, G. R.; deWeck, A.; Ruddy, D. A.; Venkatesan, K.; McAllister, G.; deBeaumont, R.; Ho, S.; Liu, Y.; Yan-Neale, Y.; Yang, G. Z.; Lin, F.; Yin, H.; Gao, H.; Kipp, D. R.; Zhao, S. P.; McNamara, J. T.; Sprague, E. R.; Cho, Y. S.; Gu, J.; Crawford, K.; Capka, V.; Hurov, K.; Porter, J. A.; Tallarico, J.; Mickanin, C.; Lees, E.; Pagliarini, R.; Keen, N.; Schmelzle, T.; Hofmann, F.; Stegmeier, F.; Sellers, W. R. Disordered methionine metabolism in MTAP/CDKN2A-deleted cancers leads to marked dependence on PRMT5. Science 2016, 351, 1208-1213. (49) Marjon, K.; Cameron, M. J.; Quang, P.; Clasquin, M. F.; Mandley, E.; Kunii, K.; Mcvay, M.; Choe, S.; Kernytsky, A.; Gross, S.; Konteatis, Z.; Murtie, J.; Blake, M. L.; Travins, J.; Dorsch, M.; Biller, S. A.; Marks, K. M. MTAP deletions in cancer create vulnerability to targeting of the MAT2A/PRMT5/RIOK1 axis. Cell Rep. 2016, 15, 574-587.


35


Page 36 of 41

(50) Kryukov, G. V.; Wilson, F. H.; Ruth, J. R.; Paulk, J.; Tsherniak, A.; Marlow, S. E.; Vazquez, F.; Weir, B. A.; Fitzgerald, M. E.; Tanaka, M.; Bielski, C. M.; Scott, J. M.; Dennis, C.; Cowley, G. S.; Boehm, J. S.; Root, D. E.; Golub, T. R.; Clish, C. B.; Bradner, J. E.; Hahn, W. C.; Garraway, L. A. MTAP deletion confers enhanced dependency on the PRMT5 arginine methyltransferase in cancer cells. Science 2016, 351, 1214-1218. (51) Kaniskan, H. Ü.; Konze, K. D.; Jin, J. Selective inhibitors of protein methyltransferases. J. Med. Chem. 2015, 58, 1596−1629. (52) Kaniskan, H. Ü.; Martini, M. L.; Jin, J. Inhibitors of protein methyltransferases and demethylases. Chem. Rev. 2018, 118, 989-1068. (53) Kaniskan, H. Ü.; Jin, J. Recent progress in developing selective inhibitors of protein methyltransferases. Curr. Opin. Chem. Biol. 2017, 39, 100-108. (54) Hu, H.; Qian, K.; Ho, M. C.; Zheng, Y. G. Small molecule inhibitors of protein arginine methyltransferases. Expert Opin. Inv. Drug. 2016, 25, 335-358. (55) Luo, M. K. Inhibitors of protein methyltransferases as chemical tools. Epigenomics 2015, 7, 1327-1338. (56) Chan-Penebre, E.; Kuplast, K. G.; Majer, C. R.; Boriack-Sjodin, P. A.; Wigle, T. J.; Johnston, L. D.; Rioux, N.; Munchhof, M. J.; Jin, L.; Jacques, S. L.; West, K. A.; Lingaraj, T.; Stickland, K.; Ribich, S. A.; Raimondi, A.; Scott, M. P.; Waters, N. J.; Pollock, R. M.; Smith, J. J.; Barbash, O.; Pappalardi, M.; Ho, T. F.; Nurse, K.; Oza, K. P.; Gallagher, K. T.; Kruger, R.; Moyer, M. P.; Copeland, R. A.; Chesworth, R.; Duncan, K. W. A selective inhibitor of PRMT5 with in vivo and in vitro potency in MCL models. Nat. Chem. Biol. 2015, 11, 432−437.


36

Page 37 of 41 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60


(57) Duncan, K. W.; Rioux, N.; Boriack-Sjodin, P. A.; Munchhof, M. J.; Reiter, L. A.; Majer, C. R.; Jin, L.; Johnston, L. D.; Chan-Penebre, E.; Kuplast, K. G.; Porter Scott, M.; Pollock, R. M.; Waters, N. J.; Smith, J. J.; Moyer, M. P.; Copeland, R. A.; Chesworth, R. Structure and property guided design in the identification of PRMT5 tool compound EPZ015666. ACS Med. Chem. Lett. 2015, 7, 162-166. (58) ClinicalTrials.gov, https://clinicaltrials.gov/ct2/show/NCT02783300 (accessed March 24, 2018). (59) Falk, H.; Foitzik, R. C.; Allan, E.; deSilva, M.; Yang, H.; Bozikis, Y. E.; Nikac, M.; Walker, S. R.; Camerino, M. A.; Morrow, B. J.; Stupple, A. E.; Lagiakos, R.; Pinson, J. A.; Lessene, R.; Kersten, W. J. A.; Ganame, D. G.; Holmes, I. P.; Lunniss, G. E.; Chung, M.; Hermans, S. J.; Parker, M. W.; Thistlethwaite, A.; White, K. L.; Charman, S. A.; Pilling, P.; Monahan, B. J.; Grusovin, J.; Peat, T. S.; Sonderegger, S.; Toulmin, E.; Jane, S. M.; Curtis, D. J.; Stupple, P. A.; Street, I. P. PRMT5 inhibitors as novel treatment for cancers. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia (PA): AACR; Cancer Res. 2015, 75(15 Suppl): Abstract nr 5371. doi:10.1158/1538-7445.AM2015-5371. (60) Bergman, Y. E.; Camerino, M. A.; Stupple, P. A. Benzopiperidine Derivatives and tTheir Use in the Treatment of Cancer and Hemoglobinopathies. WO2017153520 A1, 2017. (61) Stupple, P. A. Pyridine Derivatives and Their Use in the Treatment of Cancer and Hemoglobinopathies. WO2017153521 A1, 2017. (62) Ye, Y.; Zhang, B.; Mao, R.; Zhang, C.; Wang, Y.; Xing, J.; Liu, Y. C.; Luo, X.; Ding, H.; Yang, Y.; Zhou, B.; Jiang, H.; Chen, K.; Luo, C.; Zheng, M. Discovery and optimization of


37


Page 38 of 41

selective inhibitors of protein arginine methyltransferase 5 by docking-based virtual screening. Org. Biomol. Chem. 2017, 15, 3648-3661. (63) Couture, J. F.; Hauk, G.; Thompson, M. J.; Blackburn, G. M.; Trievel, R. C. Catalytic roles for carbon–oxygen hydrogen bonding in SET domain lysine methyltransferases. J. Bio. Chem. 2006, 281, 19280–19287. (64) Chin, H. G.; Patnaik, D.; Esteve, P. O.; Jacobsen, S. E.; Pradhan, S. Catalytic properties and kinetic mechanism of human recombinant lys-9 histone H3 methyltransferase SUV39H1: participation of the chromodomain in enzymatic catalysis. Biochemistry 2006, 45, 3272–3284. (65) Smil, D.; Eram, M. S.; Li, F.; Kennedy, S.; Szewczyk, M. M.; Brown, P. J.; BarsyteLovejoy, D.; Arrowsmith, C. H.; Vedadi, M.; Schapira, M. Discovery of a dual PRMT5-PRMT7 inhibitor. ACS Med. Chem. Lett. 2015, 6, 408-412. (66) Bonday, Z. Q.; Cortez, G. S.; Grogan, M. J.; Antonysamy, S.; Weichert, K.; Bocchinfuso, W. P.; Li, F.; Kennedy, S.; Li, B.; Mader, M. M.; Arrowsmith, C. H.; Brown, P. J.; Eram, M. S.; Szewczyk, M. M.; Barsyte-Lovejoy, D.; Vedadi, M.; Guccione, E.; Campbell, R. M. LLY-283, a potent and selective inhibitor of arginine methyltransferase 5, PRMT5, with antitumor activity. ACS Med. Chem. Lett. 2018, DOI: 10.1021/acsmedchemlett.8b00014. (67) Wu, T.; Brehmer, D.; Beke, L.; Boeckx, A.; Diels, G. S. M.; Gilissen, R. A. H. J.; Lawson, E. C.; Pande, V.; Parade, M. C. B. C.; Schepens, W. B. G. Novel 6-6 Bicyclic Aromatic Ring Substituted Nucleoside Analogues for Use as PRMT5 Inhibitors. WO2017032840 A1, 2017.


38

Page 39 of 41 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60


(68) Alinari, L.; Mahasenan, K. V.; Yan, F.; Karkhanis, V.; Chung, J. H.; Smith, E. M.; Quinion, C.; Smith, P. L.; Kim, L.; Patton, J. T.; Lapalombella, R.; Yu, B.; Wu, Y.; Roy, S.; De Leo, A.; Pileri, S.; Agostinelli, C.; Ayers, L.; Bradner, J. E.; Chen-Kiang, S.; Elemento, O.; Motiwala, T.; Majumder, S.; Byrd, J. C.; Jacob, S.; Sif, S.; Li, C.; Baiocchi, R. A. Selective inhibition of protein arginine methyltransferase 5 blocks initiation and maintenance of B-cell transformation. Blood 2015, 125, 2530−2543. (69) Zhu, K.; Jiang, C. S.; Hu, J.; Liu, X.; Yan, X.; Tao, H.; Luo, C.; Zhang, H. Interaction assessments of the first S-adenosylmethionine competitive inhibitor and the essential interacting partner methylosome protein 50 with protein arginine methyltransferase 5 by combined computational methods. Biochem. Biophys. Res. Commun. 2017, 495, 721-727. (70) Tarighat, S. S.; Santhanam, R.; Frankhouser, D.; Radomska, H. S.; Lai, H.; Anghelina, M.; Wang, H.; Huang, X.; Alinari, L.; Walker, A.; Caligiuri, M. A.; Croce, C. M.; Li, L.; Garzon, R.; Li, C.; Baiocchi, R. A.; Marcucci, G. The dual epigenetic role of PRMT5 in acute myeloid leukemia: gene activation and repression via histone arginine methylation. Leukemia 2016, 30, 789-799. (71) Politzer, P.; Murray, J. S.; Clark, T. Halogen Bonding: An Electrostatically-Driven Highly Directional Noncovalent Interaction. Phys. Chem. Chem. Phys. 2010, 12, 7748−7757. (72) Wang, C.; Jiang, H.; Jin, J.; Xie, Y.; Chen, Z.; Zhang, H.; Lian, F.; Liu, Y. C.; Zhang, C.; Ding, H.; Chen, S.; Zhang, N.; Zhang, Y.; Jiang, H.; Chen, K.; Ye, F.; Yao, Z.; Luo, C. Development of potent type I protein arginine methyltransferase (PRMT) inhibitors of leukemia cell proliferation. J. Med. Chem. 2017, 60, 8888-8905.


39


Page 40 of 41

(73) Webb, L. M.; Amici, S. A.; Jablonski, K. A.; Savardekar, H.; Panfil, A. R.; Li, L.; Zhou, W.; Peine, K.; Karkhanis, V.; Bachelder, E. M.; Ainslie, K. M.; Green, P. L.; Li, C.; Baiocchi, R. A.; Guerau-de-Arellano, M. PRMT5-selective inhibitors suppress inﬂammatory T cell responses and experimental autoimmune encephalomyelitis. J. Immunol. 2017, 198, 1439–1451. (74) Mao, R.; Shao, J.; Zhu, K.; Zhang, Y.; Ding, H.; Zhang, C.; Shi, Z.; Jiang, H.; Sun, D.; Duan, W.; Luo, C. Potent, selective, and cell active protein arginine methyltransferase 5 (PRMT5) inhibitor developed by structure-based virtual screening and hit optimization. J. Med. Chem. 2017, 60, 6289-6304. (75) Ji, S.; Ma, S.; Wang, W. J.; Huang, S. Z.; Wang, T. Q.; Xiang, R.; Hu, Y. G.; Chen, Q.; Li, L. L.; Yang, S. Y. Discovery of selective protein arginine methyltransferase 5 inhibitors and biological evaluations. Chem. Biol. Drug. Des. 2017, 89, 585-598.


40

Page 41 of 41 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60


Table of Contents graphic


41

Protein Arginine Methyltransferase 5 (PRMT5) as an Anticancer Target

Recommend Documents