Discovery of Small Molecule Ligands for MALAT1 by Tuning an RNA-Binding Scaffold

Article abstract of DOI:10.1002/anie.201808823

Structural studies of the 3′-end of the oncogenic long non-coding RNA metastasis-associated lung adenocarcinoma transcript 1 (MALAT1) confirmed a unique triple-helix structure. This structure enables accumulation of the transcript, and high levels of MALAT1 are found in several cancers. Here, we synthesize a small molecule library based on an RNA-binding scaffold, diphenylfuran (DPF), screen it against a variety of nucleic acid constructs, and demonstrate for the first time that the MALAT1 triple helix can be selectively targeted with small molecules. Computational analysis revealed a trend between subunit positioning and composition on DPF shape and intramolecular interactions, which in turn generally correlated with selectivity and binding strengths. This work thus provides design strategies toward chemical probe development for the MALAT1 triple helix and suggests that comprehensive analyses of RNA-focused libraries can generate insights into selective RNA recognition.

Full text of DOI:10.1002/anie.201808823

A Journal of the Gesellschaft Deutscher Chemiker
Angewandte
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Accepted Article
Title: Discovery of the First Small Molecule Ligands for MALAT1 by
Tuning an RNA-Binding Scaffold
Authors: Anita Donlic, Brittany S Morgan, Jason L Xu, Anqi Liu, Carlos
Roble, Jr., and Amanda E Hargrove
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To be cited as: Angew. Chem. Int. Ed. 10.1002/anie.201808823
Angew. Chem. 10.1002/ange.201808823
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10.1002/anie.201808823
Angewandte Chemie International Edition
COMMUNICATION
Discovery of the First Small Molecule Ligands for MALAT1 by
Tuning an RNA-Binding Scaffold
Anita Donlic,[a] Brittany S. Morgan,[a] Jason L. Xu,[a] Anqi Liu,[a] Carlos Roble, Jr.,[a] and Amanda E.
Hargrove*[a]
Abstract: Structural studies of the 3’-end of the oncogenic long non-
coding RNA Metastasis-associated Lung Adenocarcinoma
Transcript 1 (MALAT1) revealed a unique triple helix structure. This
structure enables accumulation of the transcript, and high levels of
MALAT1 are found in several cancers. Here, we synthesize a small
molecule library based on an RNA-binding scaffold, diphenylfuran
(DPF), screen it against a variety of nucleic acid constructs, and
demonstrate for the first time that the MALAT1 triple helix can be
selectively targeted with small molecules. Computational analysis
revealed a trend between subunit positioning and composition on
DPF shape and intramolecular interactions, which in turn generally
correlated with selectivity and binding strengths. This work thus
provides design strategies toward chemical probe development for
the MALAT1 triple helix and suggests that comprehensive analyses
of RNA-focused libraries can generate insights into selective RNA
recognition.
prospect of targeting this unique tertiary structure with small
molecules therefore represents an attractive therapeutic strategy
and a model system for exploring the druggability of lncRNA
structural domains.
While several laboratories have recently reported small
molecule probes for select mammalian RNA structures, newly
discovered structural motifs in lncRNAs are emerging targets
and remain largely underexplored with small molecule probes.[3]
Currently, there are no small molecule ligands reported to target
the MALAT1 triple helix. The most closely related research has
explored the targeting of simple T-A-T and U-A-U triplex
structures with both promiscuous ligands (i.e. aminoglycosides[7]
and dyes[8]) and the tunable diphenylfuran (DPF)-based scaffold,
furamidine. This scaffold has been diversified for preferential
binding to a variety of nucleic acid structures, including various
RNA and DNA duplexes,[9] T-A-T DNA triple helices,[10] and
disease-relevant stem-loops.[11] Varied binding modes have
The recently renewed interest in small molecule:RNA
targeting is in part due to the completion of the ENCODE project,
which resulted in the discovery that the majority of the human
transcriptome is non-protein-coding.[1] Subsequent efforts to
functionally characterize these non-coding transcripts resulted in
the identification of novel RNA classes including long non-coding
RNAs (lncRNAs). These molecules were found to have
developmental- and tissue-specific expression and to regulate
many levels of cellular processes, including expression of
oncogenes.[2] As a result, several lncRNAs have been proposed
as therapeutic targets.[3] As structured RNA elements are
continually identified in these transcripts, utilizing small
molecules to probe the specific functions and interactions of
these domains represents an exciting avenue toward
understanding ncRNA biology.
LncRNA Metastasis-Associated Lung Adenocarcinoma
Transcript 1 (MALAT1) has been recognized as a potential
anticancer target due to its overexpression in cancerous tissues
as well as studies demonstrating that its knockdown reduces
tumor growth and metastasis in preclinical models of various
cancers.[4] A study in colorectal cancer cells showed that a
~1,500 n.t. segment at the evolutionarily conserved 3’-end of
MALAT1 was sufficient to increase invasion and proliferation,
implying that this region enables its oncogenic function.[5] The
recent structural characterization of a 74 n.t. region at the 3’-end
of MALAT1 by X-ray diffraction revealed a unique, bipartite triple
helix where the U-rich stem loop sequesters the A-rich tail,
which is proposed to prevent exonucleolytic degradation (Figure
1a).[6] Notably, the deletion of this segment decreased
accumulation of the MALAT1 transcript. A comparable decrease
been reported, including intercalation, groove binding and
11b]
threading intercalation.[9a,
An additional advantage of the
DPF scaffold is its intrinsic fluorescence, which is quenched
upon binding to RNA and thus enables evaluation of binding
affinities without RNA or small molecule labeling.[11b] The
tunability of this scaffold, its intrinsic fluorescent properties, as
well as literature precedence for binding to a triple helix, position
DPF as an excellent starting point for testing the susceptibility of
a lncRNA triple helix toward targeting with a small molecule. In
this study, we design and synthesize a library of DPF-based
small molecules to identify a first-in-class selective ligand for the
MALAT1 triple helix as well as obtain fundamental insights into
the role of small molecule structure and shape in triple helix
binding strength and selectivity.
We reasoned that a small molecule with the ability to
selectively recognize and disrupt the triple helix could either
directly displace the A-rich tail by binding to the stem-loop
structure or stabilize a non-triple helical conformation. We thus
began by evaluating the original DPF scaffold, furamidine, for
binding to the MALAT1 triple helix and the stem loop construct, a
structure without the triplex-forming A-rich tail (Figure 1a,b). The
change in intrinsic fluorescence of the scaffold was measured
upon increasing RNA concentrations, which enabled us to
calculate the EC50 as an indicator of relative binding strengths as
conducted previously.[9b] The comparison of this binding EC50
with the triple helix and stem loop RNA suggests that furamidine
binds to the triple helix with ~3-fold selectivity as compared to
the stem loop (Figure 1c). Encouraged by this result, we
envisioned a strategy of efficiently tuning the RNA-binding
properties of the DPF scaffold by altering the composition and
placement of subunits to achieve orthogonal recognition of each
structure.
in accumulation was also observed upon mutation of
a
Hoogsteen-positioned uridine, proposed to disrupt the triple helix
structure, indicating that subtle alterations in the stability of this
structure can lead to significant changes in transcript level. The
[a]
A. Donlic, Dr. B. S. Morgan, J. L. Xu, A. Liu, C. Roble, Jr., Prof. Dr.
A. E. Hargrove
Department of Chemistry, Duke University
Durham, NC 27708-0346 (USA)
E-mail: amanda.hargrove@duke.edu
Supporting information for this article is given via a link at the end of
the document.
This article is protected by copyright. All rights reserved.

10.1002/anie.201808823
Angewandte Chemie International Edition
COMMUNICATION
b)
a)
c)
G
C
C
U
A
U
U
A
A
Triple Helix EC50 = 560 10 nM
Stem Loop EC50 = > 1,500 nM
U
30
A
U
A
G
A
A
C
U
U
40
A
U
A
U
A
A
U
H2N
O
NH2
NH
A
G
C
U
C
C
G
U
A
100
50
0
C
G
20
A
U
U
HN
G
50
C
GU
U
C
U
U
U
Furamidine
3’
U
G
A
A
A
U
U
U
U
G
C
U
U
U
U
U
U
U
U
U
U
C
U
C
U
U
U
A
U
U
C
G
A
A
10
U
U
U
U
G
G
A
A
G
G
C
C
60
C
U
U
U
U
A
A
U
U
U
U
A
U
G G
U
67
60
A
A
1
G
5’
3’
G
C
C
U
U
G
A
A
A
Stem Loop (67 n.t.)
A
A
-8
10
-7
10
log (RNA) / M
-6
10
-5
10
G
G
U
U
A
1
67
A
U
5’
C
A
U
U
U
C
A
G
Triple Helix (94 n.t.)
Figure 1. a) MALAT1 triple helix (94 nucleotides, left) and non-triplex-forming stem loop (67 nucleotides, right) constructs. b) Structure of furamidine. c)
Normalized fluorescence change of furamidine (1 µM) upon titration of 0.015 to 5 µM triple helix (purple) and stem loop RNA (green). EC50 values were
determined from triplicate data using the change in fluorescence intensity. No saturable binding was observed for values over 1.5 µM, resulting in ambiguous
curve fits. Error bars represent the standard deviation determined from three independent measurements.
First, we identified and optimized a robust one-step
coupling procedure to access all three symmetric regioisomers
(para-, meta-, and ortho-) of the dinitrile DPF scaffold 3 (Scheme
1).[12] To quickly diversify the scaffolds, we utilized eleven
commercially available primary amine subunits enriched in
heteroatoms and aromatic rings, chemotypes that are known to
be important for selective RNA recognition.[13] Subunit addition to
the three dinitrile scaffolds 3 was achieved by optimizing
previously reported copper[14]- or aluminum[15]-assisted coupling
reactions for diamidine formation to afford three sublibraries (p,
m, and o) with eleven identical subunits on each scaffold (DPF-
R). This final one-step coupling, which had not been applied to
the synthesis of DPF-based small molecules, was found to
increase efficiency relative to a reported two-step procedure that
involves an imidate ester intermediate.[16] To the best of our
knowledge, this route thus represents the shortest synthesis of
DPF-based libraries to date. Further, all of the synthesized
library members are novel with the exception of DPFp1 and
DPFm1. These latter ligands were strong binders of several
therapeutically relevant RNAs,[17] therefore representing an
adequate reference point for binding preferences of the newly
synthesized ligands herein.[9a, 11c]
sublibrary contained the only library member (DPFp8) that
demonstrated binding exclusively with the triple helix (Figure 2a,
Table S1). Further, only this sublibrary contained ligands with
EC50 values in the low nanomolar range, which were similar to
the reported binding affinity of the A-rich tail for the stem loop (Kd
~ 20 nM).[18] Thermal melting studies of these DPFs resulted in
stabilizing effects on the triple helix (Figure S1, Table S2),
though we note these experiments require RNA and ligand
concentrations that are well above the EC50 values in order to
measure absorbance. All meta-substituted DPFs displayed EC50
values in the micromolar range and preferential binding for the
triple helix in the fluorescence assays, which is in agreement
with literature for the binding of other meta-substituted DPFs to a
DNA triple helix.[10] Nevertheless, we noted little variability in
binding strengths and selectivities for the triple helix versus the
stem loop across the different members of the meta-DPF
sublibrary (Figure 2a, Table S1). This observation is also
supported by previous studies that reported lower selectivity
between DNA duplexes for meta- as compared to para-
substituted DPFs.[19] Similarly, the EC50 values obtained for the
ortho-DPF sublibrary indicated preferential binding for the triple
helix over the stem loop, albeit with greater variability in
selectivity between the two targets as compared to meta-DPF
sublibrary (Figure 2a, Table S1).
We next evaluated our DPF-based small molecule library
for binding to the MALAT1 triple helix and stem loop RNAs via
the aforementioned fluorescence-based screen. The para-DPF
Scheme 1. Synthesis of the diphenylfuran-based small molecule library.
R-NH2, CuCl
Pd (OAc)2
EtOH, reflux, 48 h
Br
(0.5 - 1.5 mol %)
NH
HN
HN
NH
NC
CN
NC
O
+
O
KOAc, DMAc
130 ºC, 14 h
O
2
OR
R
R
R-NH2, DABAL-Me3
THF, µW, 100 ºC
1.5 - 8 h
1
3
DPF-R
47 % , CN = para
41 % , CN = meta
60 % , CN = ortho
R =
Cl
N
N
N
N
N
S
Cl
N
H
N
p10, 30% a
m10, 8% a
o10, 8% a
p9, 13% a
m9, 19% a
o9, 6% a
p11, 64%
m11, 8%
o11, 28%
p1, 7% a
m1, 9% a
o1, 22% a
p2, 19%
m2, 46%
o2, 61%
p6, 83%
m6, 48%
o6, 30%
p8, 9%
p7, 19%
p3, 11%
m3, 8%
o3, 44%
p4, 78%
p5, 59 %
m5, 39%
o5, 42%
m8, 18%
o8, 39%
m7, 50%
o7, 24%
m4, 68%
o4, 72%
aLower final yield was due to difficulty in purifying highly polar compounds by normal phase column chromatography.
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COMMUNICATION
para
a)
meta
ortho
p11
p10
p9
p8
p7
p6
p5
p4
p3
p2
p1
o11
o10
o9
o8
o7
o6
o5
o4
o3
o2
o1
m11
m10
m9
m8
m7
m6
m5
m4
m3
m2
m1
0
0
g
50
ng
ndi
100
150
500
500
1500
2500
150
2500
4000
No bi
No bindin
EC50 / nM
EC50 / nM
EC50 / nM
b)
p11
p10
p9
Triple Helix
Stem Loop
tRNA
DNA Duplex
RRE IIB
p8
p6
p4
p1
EC50 / nM
Figure 2. a) EC50 values of the para- (left), meta- (center) and ortho-(right) sublibraries towards the MALAT1 triple helix (purple, PDB I.D.: 4PLX) and stem loop
(green) constructs. b) Selectivity screen of select para-sublibrary members against yeast tRNA (76-90 n.t., PDB I.D.:4TNA) (black), DNA duplex (28 n.t.) (gray)
and RRE Stem Loop IIB (34 n.t.) (blue). EC50 values were determined from triplicate data using the change in fluorescence intensity. First dashed line indicates
measurements above 1,500 nM for which no saturable binding events were observed, resulting in ambiguous curve fits. No binding indicates no change in
fluorescence intensity of the DPF (1 µM) upon RNA/DNA titration (up to 5 µM). Error bars represent the standard deviation determined from three independent
measurements.
We distinguished one subunit-based trend that held for all
scaffolds. Namely, ligands with the benzyl-
significantly weaker binding strengths for other constructs
(Figure 2b, Table S7). It is worth noting that these experiments
specifically capture binding events that modulate the
fluorescence of the DPF scaffold and thus binding events that do
not change fluorescence cannot be ruled out.
three
methylpiperidine subunit 8 were among the most selective
molecules for the triple helix over the stem loop (Table S1). The
remaining subunit-based trends were observed only for the
para-DPF sublibrary. For example, we noted that aliphatic
subunits with protonated nitrogens (DPFp1, -p9 and -p10)
generally increase binding strength but do not confer much
selectivity between the two targets. Presence of a chlorine
moiety on the ortho- position of the benzene subunit (DPFp6 vs.
DPFp4) was important for increasing binding strength towards
the stem loop, while placement at the para-position (DPFp6 vs.
DPFp5) reversed the selectivity between the targets (Figure 2a).
Lastly, we noted that the only sulfur-containing moiety, the
thiophene on DPFp11, allowed selective binding towards the
stem loop.
To gain more insights into the possible subunit-based
determinants of selectivity, seven para-DPF sublibrary members
with varying subunit composition (DPFp1, -p4, -p6, -p8, -p9, -
p10, and -p11) were selected for further evaluations against
nucleic acids of varying topologies. We evaluated the select
library members for binding to: i) tRNA, which constitutes ~20%
of RNA in cells; ii) an AT-rich DNA duplex, a known target for
DPF-based small molecules, and iii) the Rev-Response Element
(RRE) stem loop IIB, a shorter stem-loop construct that was also
evaluated for binding to various DPF scaffold-based libraries.[11b,
We next performed computational analyses to further
understand the impact of subunit positioning on DPF scaffold
tunability for differential recognition of MALAT1. We first
assessed the 3D shape coverage of the three sublibraries (ortho,
meta, para), as shape is known to be important for RNA
recognition.[13a, 20] Specifically, we calculated Principal Moments
of Inertia (PMI), a spatial measurement that describes ligands as
rod, disk, sphere or hybrid.[21] First, a low energy ensemble of
each ligand was generated via molecular dynamics simulations
(MD) utilizing a generalized Born solvation model. PMIs were
calculated for each structure in the ensemble, and the resulting
coordinates were averaged for each ligand using the Boltzmann
equation. Several clear trends were revealed in this analysis: i)
the para-DPF sublibrary is exclusively in the rod sub-triangle, ii)
the majority of the meta-DPF sublibrary is within the disk and
hybrid sub-triangles, with the exception of four ligands in the rod
sub-triangle and iii) the ortho-DPF sublibrary is distributed in the
sphere, disk, and hybrid sub-triangles and is thus the least rod-
like (Figure 3a, Figure S2, Table S11).
Previous PMI calculations performed by our laboratory
revealed that bioactive RNA-targeted ligands, which are
assumed to be selective, have more rod-like character as
compared to FDA-approved drugs or in vitro binding RNA
ligands with no reported bioactivity.[13a] It was therefore of
particular interest that the para-DPF sublibrary, which contained
the strongest binders and the only triple helix-selective ligand,
contained exclusively rod-shaped ligands. A closer look at this
sublibrary revealed that most members had a similar shape
except for DPFp7 and DPFp8. Specifically, DPFp7 had more
disc-like character than the bulk of the library, and it is the only
ligand in the para-DPF sublibrary that did not bind the triple helix
or the stem loop. DPFp8 had the most rod-like character in the
11c, 17]
Our results show that the library members with aliphatic,
positively charged nitrogen subunits (DPFp1, -p9 and -p10) bind
to all RNA controls with similar EC50 values, indicating non-
specific, likely electrostatic interactions with the RNA backbone
(Figure 2b, Tables S3, S6, S8 and S9). On the contrary, library
members with aromatic subunits (DPFp4, -p6 and -p11) appear
to confer selectivity for the MALAT1 stem loop as reflected in
statistically significant differences in their binding strength for
this target over all other nucleic acids tested in this screen
(Figure 2b, Tables S5, S6, and S10). Lastly, the only exclusive
triple helix binder from our initial evaluations (DPFp8) displayed
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a)
c)
Rod
Sphere
Para AVG
Ortho AVG
Meta AVG
Para
1.0
0.9
0.8
0.7
0.6
0.5
Ortho
Meta
Disc
0.0
0.2
0.4
0.6
0.8
1.0
I /I
1 3
DPFo3
b)
0º
0º
17º
24º
37º
88º
DPFp3
DPFm3
DPFo3
Figure 3. a) Principal Moments of Inertia (PMI) calculations of the DPF library. AVG – average. b) Minimal free energy (MFE) structures of example ligands:
DPFp3 (left), DPFm3 (center) and DPFo3 (right) with furan-phenyl dihedral angles indicated. Circles (blue) indicate the dihedral angles between the furan oxygen
and the contiguous phenyl ring carbon atoms. Subunits and amidine nitrogens were omitted for clarity. c) MFE of DPFo3 with intramolecular interactions:
hydrogen bonds (green line), edge-to-face π-π (purple line), and face-to-face π-π (purple dot).
entire library and is the only library member that displayed
exclusive selectivity for the triple helix.
including interactions between subunits and the π-systems on
the scaffold (Figure 3c). For example, we observed CH-π bonds
between an aliphatic subunit and the furan, as well as
alternating edge-to-face and face-to-face π-π interactions
between the phenyl groups on the scaffold and aromatic
subunits (Table S12).
Our analysis suggests that these interactions, concomitant
with the increase in scaffold tilt, ultimately enable the more disc-
and/or sphere-like shape of the MFE structures in the ortho- and
meta-DPF sublibraries. While more detailed experimental work
is needed to establish the relevance of these interactions in
aqueous solution, it is possible that these characteristics, which
are observed with almost all subunits, lead to less variability in
RNA-binding preferences among the sublibrary members.
Further, the energetic cost of breaking intramolecular
interactions may be in part responsible for the decreased
binding strengths in these two sublibraries. Ultimately, we
propose that these sublibraries are less likely to achieve
extended, rod-like shapes that are important for selective RNA
recognition.[13a]
In addition, the benzyl-piperidine subunit of DPFp8 is among the
subunits that provides the most rod-like shapes for the ortho-
and meta- sublibraries (Table S11) and is present in the ligands
that conferred the highest selectivity for the triple helix over stem
loop constructs in all three sublibraries (Table S1). Upon
inspecting all conformations in the ensembles of these ligands,
we noted extended shapes that appeared to be locked by the
ring identity and connectivity of this subunit (Figure S3). Indeed,
this subunit has been used in the literature to achieve
conformationally restricted structures.[22] Interestingly, the aryl-
methyl-piperidine motif is found in three approved drugs as well
as two of the most notable examples of RNA-targeted bioactive
ligands, Ribocil B and C (Table S13). This observation indicates
that incorporating conformationally restricted, multi-ring subunits
on para-DPFs may be a promising future direction for the
development of bioactive probes specific for the MALAT1 triple
helix
and
potentially
other
RNA:ligand
systems.
We were also interested in identifying the specific features
that led to the shape diversity observed for the three sublibraries
and thus inspected the MFE conformation from the MD
simulations. First, we noted that the DPF scaffold geometry
varies among the three different sublibraries. The scaffold is
planar or nearly planar (< 6º) for all para-substituted DPFs
(Figure 3b, Table S12). The scaffold is non-planar for 7/11 meta-
substituted DPFs, with a median furan-phenyl dihedral angle of
12º (range 0º - 26º) (Figure 3b, Table S12). None of the ortho-
substituted DPFs had a planar or nearly planar scaffold, and
these ligands had a larger median dihedral angle of 48º (range
6º - 88º) (Figure 3b, Table S12). Second, we noted that the
differences in scaffold dihedral angles across the three
sublibraries were also marked by differences in the number and
type of intramolecular interactions (Table S12). Specifically, no
intramolecular interactions were observed in the para-DPF
sublibrary as well as the four meta-DPFs that featured alkyl
subunits, which were more rod-like and had planar or nearly
planar scaffolds. On the contrary, distinct intramolecular
interactions were observed for the meta-DPF ligands with
aromatic subunits, which occupied disc and hybrid sub-triangles
(Table S12). These included NH-π or hydrogen bonds between
the amidines and subunits as well as π-π or lone pair-π
interactions between the subunits. Lastly, the ortho-DPF
sublibrary members formed more intramolecular interactions,
In summary, we synthesized a DPF scaffold-based small
molecule library diversified in subunit composition and
positioning to explore the recognition of a newly discovered
lncRNA triple helix. Library screening against MALAT1-related
constructs and controls led to the discovery of the first selective
ligand for the MALAT1 triple helix. This work demonstrates that
a
known RNA-binding scaffold can indeed be tuned to
selectively recognize a unique RNA structural element by relying
on subunit identity and placement-based differentiation. PMI
calculations revealed striking sublibrary-based differences in
small molecule shapes,
a diversity deemed important for
screening libraries but often difficult to achieve with scaffold-
based libraries of similar size.[23] Consistent with our previous
observations,[13a] the most promising ligands had rod-like shapes
in this analysis. The more disc and sphere-like DPF ligands
were predicted to have increased intramolecular interactions and
scaffold dihedral angles, which generally correlated with a
decrease in binding strengths for the triple helix. These
characteristics were particularly prevalent in the meta- and
ortho-sublibraries and may be responsible for the decrease in
differential binding among these ligands. These findings will be
utilized to design more extended and rod-like DPFs and
expedite the discovery of chemical probes for the MALAT1 triple
helix.
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Furthermore, as the intramolecular interactions identified in this
study formed through chemotypes that are commonly found in
RNA-binding small molecules, such as aromatic and
heteroatom-containing moieties, we suggest that future efforts to
design RNA-targeted libraries should investigate the possibility
of intramolecular interactions within ligands containing these
chemotypes.[13a] Lastly, we note that these interactions should
be taken into consideration when performing docking studies to
rank RNA:ligand affinities, as the analysis of energetically
inaccessible conformations due to intramolecular interactions
has led to dead-end designs in protein-based experiments.[24]
We hope that these novel insights and rational strategies
contribute to understanding of general principles for small
molecule:RNA recognition, and we anticipate that this work
encourages further small molecule-based exploration of the
MALAT1 triple helix and other lncRNA structural elements with
emerging roles in disease biology.
[13]
a) B. S. Morgan, J. E. Forte, R. N. Culver, Y. Zhang, A. E.
Hargrove, Angew. Chem., Int. Ed. Engl. 2017, 56, 13498-13502; b)
D. Vourloumis, M. Takahashi, K. B. Simonsen, B. K. Ayida, S.
Barluenga, G. C. Winters, T. Hermann, Tetrahedron Lett. 2003, 44,
2807-2811; c) S. P. Velagapudi, A. Pushechnikov, L. P. Labuda, J.
M. French, M. D. Disney, ACS Chem. Biol. 2012, 7, 1902-1909.
G. Rousselet, P. Capdevielle, M. Maumy, Tetrahedron Lett. 1993,
34, 6395-6398.
Acknowledgements
[14]
[15]
We would like to thank all current and past members of the
Hargrove lab for stimulating input and discussion. Further, we
thank Dr. George Dubay for assistance with mass spectrometry
instrumentation, Dr. David Gooden for assistance with the
selection and optimization of synthetic methodologies, Drs.
Richard Brennan and Todd Woerner for their permission to use
UV-Vis instrumentation in their laboratories, and Duke Data and
Visualization Services for help with figure preparation. A.D. is
grateful to all members of the Al-Hashimi lab, specifically Ms.
Laura Ganser, Dr. Dawn K. Merriman, Dr. Mary Clay, Mr. Atul
Kaushik Rangadurai, and Mr. Bei Liu, as well as members of the
Weeks lab, specifically Dr. Natalie McDonald, for help and
advice regarding RNA preparation and handling. A.E.H. wishes
to acknowledge Duke University, the Prostate Cancer
Foundation Young Investigator Award, and the National Institute
of Health (NIH) Maximizing Investigator’s Research Award
(MIRA), Grant/Award Number: R35GM124785 for financial
support. B.S.M. was supported by the Duke University Katherine
Goodman Stern Fellowship, and J.L.X. was supported through
the Duke University Undergraduate Research Support grant as
well as the Dean’s Summer Research Fellowship.
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This article is protected by copyright. All rights reserved.

10.1002/anie.201808823
Angewandte Chemie International Edition
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DPF shapes up as MALAT1 triplex
threat: The synthesis and evaluation
of a scaffold-based small molecule
library led to the discovery of first
ligands for the MALAT1 triple helix.
Computational analysis revealed the
role of ligand shape for selective
recognition, thereby providing design
strategies toward future probe
Anita Donlic, Brittany S. Morgan, Jason
L. Xu, Anqi Liu, Carlos Roble, Jr., and
Amanda E. Hargrove
Page No. – Page No.
Discovery of the First Small
Molecule Ligands for MALAT1 by
Tuning an RNA-Binding Scaffold
development for this unique target.
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Author(s), Corresponding Author(s)*
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Title
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This article is protected by copyright. All rights reserved.