C O M M U N I C A T I O N S
Table 1. Dissociation Constants (Kd) Measured by SPR and
G-Quadruplex Stabilization (∆Tm) Potential by FRET-Melting
pronounced than in HGC-27 cells, which may be due to the different
basal transcriptional levels of c-kit of these two cell lines. It was
interesting to note that the inhibitory effects were observed at
concentrations near the ligand Kd, which may reflect favorable
cellular uptake of such compounds.
In summary, 3,8,10-trisubstituted isoalloxazines are a promising
class of G-quadruplex binding ligands that show selective binding
to a c-kit promoter quadruplex and have provided proof of concept
for the inhibition of c-kit expression. Detailed investigations on
the chemical biology of 3,8,10-trisubstituted isoalloxazines and
quadruplexes are now underway.
DNA
1a
1b
1c
1d
1e
1f
Kd [µM]
c-kit2
3 ( 0.5 5.8 ( 2.3 9.2 ( 1.5 9.7 ( 0.8 2.8 ( 0.5 7.7 ( 0.2
c-kit1
htelo
ds
6.4 ( 0.1 9.1 ( 2.3 21 ( 2
31 ( 1
b
4.9 ( 0.3 13.7 ( 1
43 ( 3
8.6 ( 2 69 ( 7
21 ( 1
63 ( 2
nd17
nd17
nd17
nd17
nd17
nd17
∆Tm at 1 µM (°C)
c-kit2
c-kit1
htelo
ds
18
17.5
24.6
19.3
0
5.4
11
7
nda
17.2
27.1
15.5
0
14
25.3
19.1
0
nda
nda
nda
13.7
15.4
0
0
G-Quadruplex Binding Discrimination (Fold)
1/2/14 1/2/2 1/2/8 1/3 1/2/7
Acknowledgment. We thank Cancer Research U.K. for pro-
gramme and project funding, the BBSRC for funding, and COM-
SATS University faculty development programme funding for a
studentship to R.S. S.B. is a BBSRC Career Development Research
Fellow. We thank Ashok Venkitaraman for valuable discussion.
c-kit2/
c-kit1/htelo
1/2/9
a Not determined (nd) due to intrinsic fluorescence of ligand. b No
detectable binding was observed; SPR standard deviations are given for
two independent experiments; ∆Tm ( 1 °C.
Supporting Information Available: Experimental procedures for
the synthesis of isoalloxazines 1, SPR binding curves, experimental
details, curves of FRET-melting and experimental procedures of cell
based experiments. This material is available free of charge via the
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