Allosteric Modulation of Protein Arginine Methyltransferase 5 (PRMT5)

Article abstract of DOI:10.1021/acsmedchemlett.9b00525

Protein arginine methyltransferase 5 (PRMT5) belongs to a family of enzymes that regulate the posttranslational modification of histones and other proteins via methylation of arginine. Methylation of histones is linked to an increase in transcription and

Full text of DOI:10.1021/acsmedchemlett.9b00525

pubs.acs.org/acsmedchemlett
Letter
Allosteric Modulation of Protein Arginine Methyltransferase 5
(PRMT5)
Rachel L. Palte,* Sebastian E. Schneider,* Michael D. Altman, Robert P. Hayes, Shuhei Kawamura,
Brian M. Lacey, My Sam Mansueto, Michael Reutershan, Phieng Siliphaivanh, Christopher Sondey,
Haiyan Xu, Zangwei Xu, Yingchun Ye, and Michelle R. Machacek
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ABSTRACT: Protein arginine methyltransferase 5 (PRMT5)
belongs to a family of enzymes that regulate the posttranslational
modification of histones and other proteins via methylation of
arginine. Methylation of histones is linked to an increase in
transcription and regulates a manifold of functions such as signal
transduction and transcriptional regulation. PRMT5 has been
shown to be upregulated in the tumor environment of several
cancer types, and the inhibition of PRMT5 activity was identified
as a potential way to reduce tumor growth. Previously, four
different modes of PRMT5 inhibition were knowncompeting
(covalently or non-covalently) with the essential cofactor S-
adenosyl methionine (SAM), blocking the substrate binding
pocket, or blocking both simultaneously. Herein we describe an
unprecedented conformation of PRMT5 in which the formation of an allosteric binding pocket abrogates the enzyme’s canonical
binding site and present the discovery of potent small molecule allosteric PRMT5 inhibitors.
KEYWORDS: PRMT5, allosteric inhibition, methyltransferase, peptide competitive, SAM competitive, crystal structure
embers of the protein arginine methyl transferase
M(PRMT) family mono- or dimethylate arginine residues
by catalyzing the transfer of a methyl group from the cofactor
S-adenosyl methionine (SAM) to a guanidine nitrogen atom.
They are categorized into three different classes1 with PRMT5
and PRMT9 belonging to the class II PRMTs that are able to
symmetrically dimethylate arginine residues.2 PRMT enzymes
regulate a variety of cellular processes including gene
expression via histone methylation, signal transduction,
mRNA splicing, and DNA repair.3−7
Methylosome protein 50 (MEP50) has been identified as a
crucial binding partner for PRMT5, increasing its enzymatic
activity.8,9 The PRMT5−MEP50 complex specifically methyl-
ates arginine residues on histones H2A, H3, and H45,10 as well
as transcription factors and other proteins.7,11 PRMT5 is
overexpressed in certain cancer types including lung, color-
ectal, and liquid tumors,4 and the inhibition of PRMT5 has
been linked to tumor regression in mouse models.12,13
the more commonly deleted genes resulting in an increased
concentration of MTA and partial inhibition of PRMT5 in a
SAM-competitive manner.15 Examples for designed small
molecule inhibitors include the peptide-competitive inhibitor
EPZ015666,16 which binds in the catalytic substrate binding
site in a SAM-uncompetitive fashion, and LLY-283,17 a SAM-
competitive inhibitor that binds in the SAM binding site
(Figure 1). A dual SAM/substrate PRMT5 inhibitor has been
described (JNJ-64619178)18 that competes with both the
protein substrate as well as SAM by binding in the SAM
binding site and extending into the catalytic site of PRMT5,
thus prohibiting the binding of the substrate. Additionally,
covalent inhibitors have been developed and form a covalent
bond with Cys449 in the SAM binding site.19 Here, we
describe the binding of an allosteric inhibitor which stabilizes
an alternative state of the enzyme in which large portions of
Different modes of PRMT5 inhibition have been reported in
the literature, with small molecule inhibitors competing with
either the protein substrate or the SAM cofactor. A natural
inhibitor of PRMT5 is methylthioadenosine (MTA) which is
converted to adenosine and 5-methylthioribose-1-phosphate
by the enzyme methylthioadenosine phosphorylase
(MTAP).14 In cancer cells, MTAP has been found as one of
Received: November 14, 2019
Accepted: July 30, 2020
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© XXXX American Chemical Society
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this loop. Despite sharing low sequence homology the
backbone atoms of this amino acid loop for PRMT1-PRMT8
are positioned within 1.29 0.57 Å RMSD (Figure S4, Figure
S5, and Table S2). We hypothesize that the slow on rate for
this compound is due to the required large loop movement
needed for formation of the allosteric pocket.
Figure 2B shows the binding mode of this novel allosteric
inhibitor in the context of the other known modes of
inhibition, specifically substrate-competitive and SAM-com-
petitive inhibitors. While significant, the loop movement is
highly localized and no other large-scale changes in protein
conformation are seen between the three structures.
1a was observed to form several key interactions with nearby
protein residues in the allosteric site (Figure 3). The
adamantane moiety of 1a binds in a mostly hydrophobic
pocket and is surrounded by two leucine residues (Leu436 and
Leu437), two phenylalanines (Phe519 and Phe555), and a
tyrosine residue (Tyr468). The methoxyphenyl group of 1a is
buried between several aromatic residues (Phe519, Phe584,
Phe602) and is engaged in an edge to face stacking interaction
with Tyr613. Interestingly, the methoxyphenyl substituent
replaces a phenylalanine (Phe440) of PRMT5 present in the
EPZ015666-bound crystal structure (Figure 2A) while the
center of mass of Phe440 is displaced roughly 22 Å to occupy
the canonical SAM binding site. The amine of the
iminohydantoin core is interacting via a hydrogen bond with
Glu444, one of the key catalytic residues involved in the
methylation of substrates (Figure 3A). Compared to the
EPZ015666-bound crystal structure, the Cα of Glu444 is only
shifted about 2 Å; however, the overall loop movement and
rotation of Glu444 relocates the side chain carboxyl by
approximately 4.5 Å to form the interaction with 1a.
Visual inspection of the binding site revealed additional
space between the adamantyl group and several of the
surrounding residues enabling future inhibitor design (Figure
3). To further test the flexibility of the allosteric pocket,
analogs with substituents at both positions off the iminohy-
dantoin core were prepared (Table 1). Moving the methoxy
group from the para to the meta position of the benzene ring at
R1 is tolerated, though a small loss in biochemical potency is
observed (2). Furthermore, the methoxyphenyl ring could be
replaced by a naphthalene (4) and retain potency as well as the
binding position within the allosteric pocket (data not shown).
Simplifying the R1 substituent to an unsubstituted phenyl
resulted in 3, which retained its biochemical potency.
Introducing additional polar groups on the adamantyl
moiety was not well tolerated; for example, addition of an
alcohol group to the adamantyl moiety resulted in a loss of
potency (Table 1, Compounds 5 and 6). Replacing the
adamantane with other hydrophobic groups was tolerated but
resulted in a loss of potency. Substituting a bicyclo[2.2.2]-
octane (7) resulted in a 16-fold loss of potency in the
biochemical assay compared to 1a. Similar observations were
made when replacing the adamantane with a propylcyclohex-
ane. In the context of the des-methoxy R1 substituent, 8 lost 5-
fold potency in the biochemical assay.
The crystal structure of 1a also revealed the opening of a
small channel from the allosteric binding site toward the
canonical substrate binding site, offering the opportunity to
grow the molecule in this direction (Figure 3B). Cyclization of
the core allowed the introduction of small substitutions along a
trajectory predicted to access this channel, resulting in 9 and
Figure 1. Structures of EPZ015666, LLY-283, and 1a.
the catalytic site are displaced precluding both substrate and
SAM from binding.
Compound 1a (Figure 1) was previously described as a
BACE1 and BACE2 inhibitor20 and was also identified as a
potent PRMT5 inhibitor through a high throughput screening
campaign. 1a showed on-target potency with an EC50 of 16 nM
in a biochemical methylation assay (Table 1; raw data curves in
Figure S1). Cell-based inhibition of 4.6 μM EC50 was
measured in MCF7 cells via quantitation of symmetrically
dimethylated nuclear protein levels using high content imaging
technology (methods in Supporting Information; high-content
cellular images shown in Figure S2). This compound exhibited
slow binding kinetics with an approximate on-rate of ka =
∼1000 M−1 s−1 (Figure S3) which is nearly 4 orders of
magnitude slower than a typical fast binding step (instrumental
upper limit is 5 × 107 M−1 s−1).21 While the (S)-enantiomer 1b
was described as the more potent inhibitor of BACE1 and
BACE2, the (R)-enantiomer (1a) is nearly 200-fold more
potent than the (S)-enantiomer for PRMT5 in the biochemical
assay. 1a and related analogs were prepared as described in the
Supporting Information.
To understand the binding mode of this novel PRMT5
inhibitor PRMT5:MEP50 was cocrystallized with 1a. The
structure was solved at 2.7 Å resolution with one
PRMT5:MEP50 complex in the asymmetric unit, unambig-
uous ligand density, and an overall Rmerge of 0.07 (Table S1).
Additional details of crystallization protocols and structural
statistics can be found in the Supporting Information. Atomic
coordinates and structure factor files have been deposited in
the Protein Data Bank (6UXX).
The cocrystal structure of the PRMT5:MEP50 complex with
1a reveals a unique binding mode and significant structural
changes in the protein backbone (Figure 2A). An 11 amino
acid loop consisting of residues Glu435 through Leu445 is
displaced by up to 16.5 Å opening a new binding pocket. This
movement causes the residues of the loop to both occupy the
SAM binding site and occlude the substrate binding site so that
neither SAM nor peptide substrate would be expected to bind.
This conformational change was unexpected as published
structures of PRMT enzymes both with and without
orthosteric inhibitors bound reveal the relative stability of
B
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Table 1. Allosteric Inhibitor Biochemical Data
10. Both 9 and 10 lost about 5- to 6-fold biochemical potency
compared to parent 3.
binding of SAM. However, alternative configurations in which
both an allosterically binding molecule and SAM are bound
seem possible.
In summary, we report to our knowledge the first allosteric
dual substrate- and SAM-competitive inhibitor of PRMT5.
Compared to EPZ015666 and LLY-283, these allosteric
inhibitors bind to a PRMT5 conformation in which an 11
Interestingly, structural insight into the binding mode of 8
revealed that alternative configurations of the 11 amino acid
long loop can be stabilized by the allosteric inhibitors (Figure
4; PDB ID 6UXY). 8 binds to an intermediate state of the loop
which resides between the substrate-bound conformation and
the conformation that is stabilized by 1a. This indicates that
although this loop appears conformationally inflexible in the
presence of orthosteric inhibitors a configurational landscape
might exist in which different loop conformations can be
stabilized depending on the allosteric inhibitor present. In both
cases for which we have structural data, the loop prohibits the
̈
amino acid loop is displaced by several Ångstroms, blocking
the SAM binding site and abrogating the substrate binding site.
We believe that this rearrangement is both energetically
unfavorable and conformationally rare and may be the
contributing factor to the observed slow on-rate. Additional
design of analogs showed tractable SAR and revealed that
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Figure 2. (A) Superposition of PRMT5:MEP50 cocrystal structures highlights the significant 11 amino acid loop movement. The currently
reported structure with 1a is shown in slate blue overlaid with the EPZ015666-bound structure in maroon. (B) An overlay of the 1a-bound
structure (slate blue), EPZ015666-bound structure (maroon), and LLY-283-bound structure (green) shows the localized nature of the 11-residue
loop movement (shown in brighter colors) as the rest of the PRMT5 protein fold remains unaffected.
Figure 4. Overlay of the structures bound to 1a (cyan/slate blue) and
8 (yellow) revealing a configurational landscape of stable loop
conformations.
protein backbone24 (see Figure S7). Recently, an allosteric
inhibitor of PRMT6 was developed by the Structural
Genomics Consortium (SGC).25 Our results add PRMT5 to
the list of known allosterically inhibitable PRMTs and
combined with substrate-competitive (EPZ015666), SAM-
competitive (LLY-283), dual SAM/substrate inhibitors (JNJ-
64619178), and covalent SAM-competitive inhibitors (Prelude
Therapeutics), this marks the fifth reported mode of inhibition
for PRMT5.
Figure 3. (A) Closeup view of 1a in the allosteric binding pocket of
PRMT5. 1a shown as cyan sticks in the 2fo-fc electron density
contoured to 1σ. (B) Superposition of 1a and EPZ015666-bound
structures shown in cyan/slate blue and maroon, respectively. The
solvent-accessible channel is shown in gray, highlighting the open
channel between 1a and the canonical substrate binding pocket.
ASSOCIATED CONTENT
* Supporting Information
The Supporting Information is available free of charge at
https://pubs.acs.org/doi/10.1021/acsmedchemlett.9b00525.
■
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additional loop conformations exist to which small molecules
can bind. We report here two crystal structures with
resolutions 2.7 Å or better that shed light on the molecular
interactions of the allosteric inhibitors with the receptor.
Previously, selective small molecule allosteric inhibitors for
PRMT3 were identified22 to occupy a pocket aside of the
substrate or SAM binding sites (see Figure S6) and were
shown to be noncompetitive.23 In contrast to 1a, the known
PRMT3 inhibitors do not induce large scale changes of the
Experimental procedures for the biochemical assay, cell-
based assay, SPR studies, chemical syntheses, NMR and
ECD studies for the determination of absolute stereo-
chemistry, protein expression and purifications, and X-
ray crystallography. Also included is the crystallography
data collection and refinement statistics, PRMT loop
comparisons, and SPR raw data. (PDF)
D
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Accession Codes
Compound 1a, 6UXX; compound 8, 6UXY.
ABBREVIATIONS
■
PRMT5, protein arginine methyl transferase; SAM, S-
adenosylmethionine; MEP50, methylosome protein 50;
MTA, methylthioadenosine; MTAP, methylthioadenosine
phosphorylase; BACE, beta-sacretase; RMSD, root-mean-
square deviation
AUTHOR INFORMATION
■
Corresponding Authors
Rachel L. Palte − Computational and Structural Chemistry,
Merck & Co., Inc., Boston, Massachusetts 02115, United States;
Email: rachel.kubiak@merck.com
Sebastian E. Schneider − Computational and Structural
Chemistry, Merck & Co., Inc., Boston, Massachusetts 02115,
United States; orcid.org/0000-0003-1233-8984;
Email: sebastian.schneider@merck.com
REFERENCES
■
(1) Bedford, M. T.; Clarke, S. G. Protein arginine methylation in
mammals: who, what, and why. Mol. Cell 2009, 33, 1−13.
(2) 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.
(3) 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, 1431.
(4) Yang, Y.; Bedford, M. T. Protein arginine methyltransferases and
cancer. Nat. Rev. Cancer 2013, 13, 37−50.
(5) Zhao, Q.; Rank, G.; Tan, Y. T.; Li, H.; Moritz, R. L.; Simpson, R.
J.; Cerruti, L.; Curtis, D. J.; Patel, D. J.; Allis, C. D.; Cunningham, J.
M. PRMT5-mediated methylation of histone H4R3 recruits
DNMT3A, coupling histone and DNA methylation in gene silencing.
Nat. Struct. Mol. Biol. 2009, 16, 304−311.
(6) Auclair, Y.; Richard, S. The role of arginine methylation in the
DNA damage response. DNA Repair 2013, 12, 459−465.
(7) Karkhanis, V.; Hu, Y. J.; Baiocchi, R. A.; Imbalzano, A. N.; Sif, S.
Versatility of PRMT5-induced methylation in growth control and
development. Trends Biochem. Sci. 2011, 36, 633−641.
(8) Ho, M. C.; Wilczek, C.; Bonanno, J. B.; Xing, L.; Seznec, J.;
Matsui, T.; Carter, L. G.; Onikubo, T.; Kumar, P. R.; Chan, M. K.;
Brenowitz, M. Structure of the arginine methyltransferase PRMT5-
MEP50 reveals a mechanism for substrate specificity. PLoS One 2013,
8, No. e57008.
(9) Antonysamy, S.; Bonday, Z.; Campbell, R. M.; Doyle, B.;
Druzina, Z.; Gheyi, T.; Han, B.; Jungheim, L. N.; Qian, Y.; Rauch, C.;
Russell, M. Crystal structure of the human PRMT5: MEP50 complex.
Proc. Natl. Acad. Sci. U. S. A. 2012, 109, 17960−17965.
(10) Ancelin, K.; Lange, U. C.; Hajkova, P.; Schneider, R.; Bannister,
A. J.; Kouzarides, T.; Surani, M. A. Blimp1 associates with Prmt5 and
directs histone arginine methylation in mouse germ cells. Nat. Cell
Biol. 2006, 8, 623−630.
(11) 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.
Authors
Michael D. Altman − Computational and Structural Chemistry,
Merck & Co., Inc., Boston, Massachusetts 02115, United States
Robert P. Hayes − Computational and Structural Chemistry,
West Point, Pennsylvania 19486, United States
Shuhei Kawamura − Discovery Chemistry, Merck & Co., Inc.,
Boston, Massachusetts 02115, United States
Brian M. Lacey − Quantitative Biosciences, Merck & Co., Inc.,
Boston, Massachusetts 02115, United States
My Sam Mansueto − Quantitative Biosciences, Merck & Co.,
Inc., Boston, Massachusetts 02115, United States
Michael Reutershan − Discovery Chemistry, Merck & Co., Inc.,
Boston, Massachusetts 02115, United States
Phieng Siliphaivanh − Discovery Chemistry, Merck & Co., Inc.,
Boston, Massachusetts 02115, United States
Christopher Sondey − Quantitative Biosciences, Merck & Co.,
Inc., Boston, Massachusetts 02115, United States
Haiyan Xu − Quantitative Biosciences, Merck & Co., Inc.,
Boston, Massachusetts 02115, United States
Zangwei Xu − Quantitative Biosciences, Merck & Co., Inc.,
Boston, Massachusetts 02115, United States
Yingchun Ye − Discovery Chemistry, Merck & Co., Inc., Boston,
Massachusetts 02115, United States
Michelle R. Machacek − Discovery Chemistry, Merck & Co.,
Inc., Boston, Massachusetts 02115, United States
Complete contact information is available at:
https://pubs.acs.org/10.1021/acsmedchemlett.9b00525
Author Contributions
(12) Kaushik, S.; Liu, F.; Veazey, K. J.; Gao, G.; Das, P.; Neves, L.
F.; Lin, K.; Zhong, Y.; Lu, Y.; Giuliani, V.; Bedford, M. T. Genetic
deletion or small-molecule inhibition of the arginine methyltransferase
PRMT5 exhibit anti-tumoral activity in mouse models of MLL-
rearranged AML. Leukemia 2018, 32, 499−509.
(13) 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. A selective inhibitor of PRMT5 with in
vivo and in vitro potency in MCL models. Nat. Chem. Biol. 2015, 11,
432−437.
Rachel L. Palte and Sebastian E. Schneider contributed equally
to the manuscript.
Notes
The authors declare the following competing financial
interest(s): All authors are or were employees of Merck
Sharp & Dohme Corp., a subsidiary of Merck & Co., Inc.,
Kenilworth, NJ, USA.
(14) 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. MTAP deletion confers enhanced dependency on
the PRMT5 arginine methyltransferase in cancer cells. Science 2016,
351, 1214−1218.
ACKNOWLEDGMENTS
■
We thank Charles Lesburg for helpful comments on the
manuscript. We thank Donovon Adpressa and Xiao Wang for
their work to determine the absolute stereochemistry of
compounds 1a and 1b. We thank Viva Biotech for help with
crystallography and data collection. We thank Lianyun Zhao
and the WuXi AppTech chemistry team for help with the
compound syntheses discussed in this article. We would also
like to thank the reviewers for their thoughtful comments and
efforts toward improving our manuscript.
(15) Mavrakis, K. J.; McDonald, E. R.; Schlabach, M. R.; Billy, E.;
Hoffman, G. R.; deWeck, A.; Ruddy, D. A.; Venkatesan, K.; Yu, J.;
McAllister, G.; Stump, M. Disordered methionine metabolism in
MTAP/CDKN2A-deleted cancers leads to dependence on PRMT5.
Science 2016, 351, 1208−1213.
(16) 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
https://dx.doi.org/10.1021/acsmedchemlett.9b00525
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Letter
E.; Kuplast, K. G.; Porter Scott, M. Structure and property guided
design in the identification of PRMT5 tool compound EPZ015666.
ACS Med. Chem. Lett. 2016, 7, 162−166.
(17) 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. LLY-283, a potent and selective inhibitor of
arginine methyltransferase 5, PRMT5, with antitumor activity. ACS
Med. Chem. Lett. 2018, 9, 612−617.
(18) Wu, T.; Brehmer, D.; Beke, L.; Boeckx, A.; Diels, G. S.;
Gilissen, R. A.; Lawson, E. C.; Pande, V.; Parade, M. C.; Schepens, W.
B.; Thuring, J. W.; inventors. Novel 6−6 bicyclic aromatic ring
substituted nucleoside analogues for use as prmt5 inhibitors. United
States patent application US 2018, 15/755, 475.
(19) Lin, H.; Wang, M.; Zhang, Y. W.; Tong, S.; Leal, R. A.; Shetty,
R.; Vaddi, K.; Luengo, J. I. Discovery of potent and selective covalent
protein arginine methyl-transferase 5 (PRMT5) inhibitors. ACS Med.
Chem. Lett. 2019, 10, 1033−1038.
(20) Malamas, M. S.; Erdei, J.; Gunawan, I.; Turner, J.; Hu, Y.;
Wagner, E.; Fan, K.; Chopra, R.; Olland, A.; Bard, J.; Jacobsen, S.
Design and synthesis of 5, 5′-disubstituted aminohydantoins as potent
and selective human β-secretase (BACE1) inhibitors. J. Med. Chem.
2010, 53, 1146−1158.
(21) GE Healthcare Life Sciences. Biacore T200. Available at
https://www.gelifesciences.com/en/us/shop/protein-analysis/spr-
label-free-analysis/systems/biacore-t200-p-05644 (accessed 2019-10-
30).
(22) Liu, F.; Li, F.; Ma, A.; Dobrovetsky, E.; Dong, A.; Gao, C.;
Korboukh, I.; Liu, J.; Smil, D.; Brown, P. J.; Frye, S. V. Exploiting an
allosteric binding site of PRMT3 yields potent and selective
inhibitors. J. Med. Chem. 2013, 56, 2110−2124.
̈
(23) Kaniskan, H.U.; Szewczyk, M. M.; Yu, Z.; Eram, M. S.; Yang,
X.; Schmidt, K.; Luo, X.; Dai, M.; He, F.; Zang, I.; Lin, Y. A potent,
selective and cell-active allosteric inhibitor of protein arginine
methyltransferase 3 (PRMT3). Angew. Chem., Int. Ed. 2015, 54,
5166−5170.
(24) Kaniskan, H. U.; Eram, M. S.; Zhao, K.; Szewczyk, M. M.;
Yang, X.; Schmidt, K.; Luo, X.; Xiao, S.; Dai, M.; He, F.; Zang, I.
Discovery of potent and selective allosteric inhibitors of protein
arginine methyltransferase 3 (PRMT3). J. Med. Chem. 2018, 61,
1204−1217.
(25) SGC6870 A chemical probe for PRMT6. Retrieved (2020-01-
07) from https://www.thesgc.org/chemical-probes/SGC6870.
F
https://dx.doi.org/10.1021/acsmedchemlett.9b00525
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