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Supplementary Information

Fluorogenic Pt complexes Distinguish the quantity and Folding

Behavior of RNA G-quadruplexes between Live Cancerous and

Healthy Cells

Lei He,b,c Zhenyu Meng,c Qianqian Guo,a Xiangyang Wu,c Marie-Paule Teulade-Fichou,c,d Edwin Kok Lee Yeow,c and Fangwei Shao*a

a Zhejiang University-University of Illinois at Urbana-Champaign Institute, Zhejiang University, Haining 314400, China

b State Key Laboratory of Chemical Resource Engineering, Beijing University of Chemical Technology, Beijing 100029, China

c School of Physical and Mathematical Sciences, Nanyang Technological University, Singapore 637371d CNRS UMR 9187, INSERM U1196, Institut Curie, PSL Research University and Université Paris Sud, Université Paris Saclay, 91405, Orsay, France

* To whom correspondence should be addressed. Email: fwshao@zju.edu.cn

Electronic Supplementary Material (ESI) for ChemComm.This journal is © The Royal Society of Chemistry 2020

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Methods

Materials. All the chemicals, G-quadruplex folding RNA sequences, ssRNA (5’-

gggaaggccagggaaucuuccc-3’) and tRNA (transferred from bovine liver) were purchased from

Sigma-Aldrich. Human HeLa cell total RNA was purchased from Takara Bio USA. Because all the

RNA GQs fold into parallel topologies regardless of the sequences, two well-characterized GQ-

folding sequences, TERRA (5’-uaggguuagggu-3’) and EBNA1 (5’-ggggcaggagcaggagga-3’), are

selected as representative examples of RNA GQ in this study. ds26 (5’-

CAATCGGATCGAATTCGATCCGATTG-3’) and F-c-myc-T (5’-FAM-

TGGGGAGGGTGGGGAGGGTGGGGAAGG-TAMRA-3') were purchased from Sangon

(Shanghai, China). BG4 antibody was purchased from Absolute Antibody Ltd. PhenDC3 was

provided by Professor Marie-Paule from Institut Curie, France. Milli-Q water was used in all

physical measurement experiments. G-quadruplex RNA structures were annealed by heating the

desire single stranded sequences at 95 °C for 10 min and gradually cooling down to room

temperature over 2 h.

Synthesis of Pt complexes

[Pt(bpa)](PF6)2 (1). The bpa ligand (bpa-H+) was protonated by dissolving in trifluoroacetic acid

and precipitated by diethyl ether. Pt(dmso)2Cl2 (20.9 mg, 0.05 mmol) and bpa-H+ (24.5 mg, 0.05

mmol) were dissolved in ethylene glycol. The mixture was stirred at 125 °C for 1.5 h. Orange solid

was precipitated by adding an excessive amount of saturated KPF6 solution. The precipitate was

washed with water, cold methanol and ethyl ether, each solvent for 3 times. The product yield is

76%. Crystal of 1 was grown by diffusing acetone into the saturated solution of 1 in dimethyl

sulfoxide at r.t. and orange needle crystals were obtained. ESI-MS (MeOH) m/z: 567.44 (calc 568.10

for C24H16N5Pt).

4,7-Dimethyl-1,10-phenanthroline-N-oxide (a). To a solution of 4,7-dimethyl-1,10-

phenanthroline (480 mg) in acetic acid (8 mL) was added 30% H2O2 (580 μL). The temperature was

maintained at 70 oC for 3 h, after which an additional portion of H2O2 (580 μL) was added and

continued stirring for an additional 3 h. After cooling to r.t., a third portion of H2O2 (360 μL) was

added and the reaction mixture was stirred for 6 h. Then, the solution was concentrated under

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vacuum to a total volume of approximately 3 mL, which was followed by addition of water and re-

concentration back to a volume of 4 ml. To the resulting dark-brown oil, a solid K2CO3 was added

and the mixture was extracted by CHCl3 for 3 h. To the CHCl3 solution, a charcoal and MgSO4 were

added. Subsequently, solids were filtered of and the solvent was removed to afford the product (280

mg, 56% yield). 1H NMR (300 MHz, CDCl3) δ: 9.09 (d, J = 4.1 Hz, 1H), 8.60 (d, J = 6.3 Hz, 1H),

7.85 (dd, J = 53.4, 9.4 Hz, 2H), 7.44 (d, J = 4.0 Hz, 1H), 7.24 (d, J = 6.3 Hz, 1H), 2.68 (d, J = 24.7

Hz, 6H). ESI-MS (MeOH) m/z: 225.15.

2-Chloro-4,7-dimethyl-1,10-phenanthroline (b). To a mixture of a (226 mg) and NaCl (1.2 g) in

DMF (8 mL), a neat POCl3 (0.3 ml) was added by syringe at 0 ºC. Then, the reaction mixture was

stirred at 100 oC for 6 h. After cooling to r.t., water (5 mL) was added and the mixture was basified

with aqueous ammonia and saturated with NaCl. Solids were filtered and solution was extracted by

CHCl3 (3 × 5 mL). The combined extracts were washed with brine (3 × 5 mL), dried over MgSO4

before the solvent was removed under vacuum. The crude residue was purified by flash

chromatography (SiO2; 3:2, EtOAc/hexanes; Rf = 0.3) to afford the product as a yellow solid (90

mg, 35% yield). 1H NMR (300 MHz, CDCl3) δ: 9.05 (d, J = 4.5 Hz, 1H), 7.95 (dd, J = 22.9, 9.3 Hz,

2H), 7.44 (d, J = 7.3 Hz, 2H), 2.75 (d, J = 10.3 Hz, 6H). ESI-MS (MeOH) m/z: 243.14.

2-Amino-4,7-dimethyl-1,10-phenanthroline (c). A mixture of b (90 mg), acetamide (0.5 g) and

K2CO3 (355 mg) was heated to 180 oC and stirred for 2 h. After cooling to r.t., water (50 mL) was

added and resulting mixture was extracted with CH2Cl2 (4 × 10 mL). The combined organic extracts

were washed with brine (3 × 10 ml), dried over MgSO4 before the solvent was removed under

vacuum. The crude residue was purified by flash chromatography (SiO2; 3:7:90, NH4OH

(25%)/MeOH/EtOAc; Rf = 0.3) to afford c as a yellow powder (37 mg, 47% yield). 1H NMR (300

MHz, CDCl3) δ: 8.95 (d, J = 4.4 Hz, 1H), 7.81 (dd, J = 39.1, 9.2 Hz, 2H), 7.37 (d, J = 4.2 Hz, 2H),

2.71 (d, J = 25.4 Hz, 6H). ESI-MS (MeOH) m/z: 224.21.

N,N-Bis-(4,7-dimethyl-1,10-phenanthrolin-2-yl)amine (dmbpa). A mixture of c (37 mg) and

NaH (40 mg) were dissolved in DMA (2 mL) and stirred at 100 oC for 0.5 h. After that, a solution

of b (40 mg) in DMA (2 mL) was added by dropwise to the mixture. The mixture was stirred for 6

h with refluxing. DMA was distilled out and the crude product residue was purified by flash

chromatography (SiO2; 1:2:20, TEA /MeOH/DCM). The product was acidified with TFA and

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precipitate by addition of diethyl ether to afford a yellow powder (42 mg, 51% yield). 1H NMR (300

MHz, MeOD) δ: 8.89 (d, J = 4.5 Hz, 2H), 8.19 (q, J = 9.3 Hz, 4H), 7.73 (d, J = 4.4 Hz, 2H), 7.45

(s, 2H), 2.88 (d, J = 5.7 Hz, 12H). ESI-MS (MeOH) m/z: 430.40.

[Pt(dmbpa)](PF6)2 (2). Pt(dmso)2Cl2 (15.8 mg, 0.04 mmol) and dmbpa-H+ (20.3 mg, 0.04 mmol)

were dissolved in ethylene glycol and water (in 2:1 ratio). The mixture was heated at 100 °C for 2

h. Yellow solid was precipitated by adding an excessive amount of saturated KPF6 solution. The

precipitate was washed with water, cold methanol and ethyl ether 3 times for each to provide the

product in 61% yield. ESI-MS (MeOH) m/z: 623.59 (calc 624.16 for C28H23N5Pt).

Circular dichroism (CD) measurement

CD spectra were recorded on a Jasco J-1500 spectropolarimeter using a 1 cm path length cuvette.

TERRA RNA stock solutions were diluted to 6 μM, with 10 mM potassium phosphate buffer

(pH=7.4) with 15 mM KCl. CD spectra of TERRA, in the absence and presence of 3 μM of 1 or 2,

were recorded from 220-320 nm at a scan rate of 200 nm/min. These CD spectra were baseline-

corrected, and each curve represented of five averaged scans taken at 25 °C.

For thermal melting assay, ellipticity at 263 nm for TERRA and EBNA1 were monitored upon

temperature elevation from 15 °C to 95 °C with heating ramp of 0.5 °C/min. Final analysis of the

data was carried out using Origin 8.0 (OriginLab Corp.). Standard deviation over three repeat

experiments were used as error bars.

Fluorescence resonance energy transfer (FRET) melting studies

A GQ folding sequence, c-myc, labeled with 6-FAM at 5’ and TAMRA at 3’ was chosen for FRET

evaluation. F-c-myc-T was diluted to 0.4 μM in 10 mM lithium cacodylate buffer with 1 mM KCl

(pH = 7.4) and then annealed by heating to 90°C for 5 min, followed by slowly cooling down to

25°C at 1 °C/min. 20 μL of 0.4 μM F-c-myc-T with 20 μL 2 μM of 1 or TMPyP4 in cacodylate

buffer were applied into several wells of 96-well plate to a total volume of 40 μL. For the

competition assay, 10μL 8 μM ds26 was added to the mixed solution of 20μL 0.4 μM F-c-myc-T

and 10μL 4 μM 1 or TMPyP4 to a total volume of 40 μL. Each concentration data point was

performed in triplicate. The fluorescence of FAM was recorded on Biorad CT1000 Quantity PCR

instrument with excitation at λ = 450-490 nm and detection at λ = 515-530 nm. The melting profiles

S5

were collected over 25°C to 100°C with an interval of 0.3 °C. A constant temperature maintained

for 30 s prior to each reading to ensure a stable value.

Fluorescent emission measurement

Fluorescent spectra were recorded on Shimadzu RF-5301 Spectrofluorophotometer. All the samples

were excited at 450 nm. For aqueous samples, the spectra were collected from 520 to 700 nm. For

non-aqueous samples, the spectra were collected from 510 nm to 700 nm. For anaerobic samples,

the aqueous solutions of 1 (5 μM in 10 mM potassium phosphate buffer, pH 7.4) with and without

GQ-folding structure (5 μM) were prepared by blowing N2 gas for 30 min. For nucleic acid

titrations, aliquots of a high concentration of DNA/RNA stock solution (500 μM) were added to the

solution of 1 (1 μM). The equilibrium dissociation constant (Kd) of 1 on different nucleic acid was

obtained by fitting the fluorescent titration curves with the Hyperbl function. For albumin

interaction, 1.8 g/mL of albumin was added to 1 (1 μM) when needed. The emission quantum yield

was measured with Ru(bpy)3Cl2 (Φ = 0.055) as reference.

Ultraviolet–visible (UV-vis) absorption measurement

Absorption spectra of 1 (25 μM in 10 mM potassium phosphate buffer, pH=7.4) in the absence and

presence of different RNA structures (25 μM) were recorded on a Shimadzu UV-1800

spectrophotometer at 25 °C. The UV-vis spectrum were scanned from 350 nm to 550 nm.

Time-resolved transient absorption spectroscopy

Time-resolved nanosecond transient absorption measurements were performed using an Applied

Photophysics LKS.60 Nanosecond Laser Flash Photolysis Spectrometer with 430 nm excitation

pulses generated from a seed Nd:YAG, Q-switched laser, and the probe was a 150-W Xe lamp

aligned normal to the excitation source. Each time-resolved trace was acquired by averaging 20

laser shots at a repetition rate of 1 Hz. The typical laser pump fluence used is ~60 μJcm-2. The data

were collected in transmission geometry with the sample positioned at 90° with respect to both the

excitation source and probe light.

Lifetime of excited states

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Time-resolved transient absorption of 1 (50 μM) in the presence and absence of GQ-folding

structure (50 μM) was collected by pump-probe method with a short laser pulse excitation at 430

nm and a delayed pulse probed at 550 nm. The time dependence of the absorption changes was

measured and used to analyze the lifetime of excited state with Origin 8.0 software.

Phosphorescent emission measurement

Photoluminescence spectra were recorded on a Cary Eclipse spectrofluorometer system. 5 μM of 1

in 10 mM potassium phosphate buffer (pH 7.4) with and without GQ-folding structure (5 μM) were

prepared by blowing N2 gas for 30 min. The delay and gate time were 0.1 and 0.04 ms, respectively.

The samples were excited at 450 nm and the spectra were collected from 500 to 750 nm.

Cell viability studies

HeLa cells were seeded in 96-well plates (5000 cells per well) and incubated in DMEM media

containing 10% fetal bovine serum at 37 °C, with 5% CO2 atmosphere for 24 h. 1 was dissolved in

bio-grade DMSO, diluted with DMEM medium to the required concentrations (contains <1% v/v

DMSO) and added to the wells in the dark respectively. After another 24 h treatment, 10 μL of 5

mg/mL methyl thiazolyl tetrazolium (MTT) solution was added to each well, and the cells were

further incubated for 4 h. Then the medium was removed and 100 μL DMSO was added into each

well. After the plates were shaken for 10 mins, the optical density of each well was measured on a

plate spectrophotometer at a wavelength of 570 nm with background subtraction at 650nm. All

experiments were performed in parallel and in quintuplicate.

Fixed cell staining experiment

The HeLa cells were cultured in DMEM medium containing 10% fetal bovine serum at 37 °C, with

5% CO2 atmosphere. 40000 cells were seeded into an imaging dish (ibidi, μ-dish 35 mm, high) and

cultured for 24 h. Cells were fixed with 4% paraformaldehyde/PBS at r.t. for 15 min and

permeabilized with 0.5% TritonX-100/PBS at 37 °C for 30 min. For enzymatic treatments, cells

were treated with 0.12 U μL-1 DNase I or with 100 μg mL-1 RNase A for 3 h before staining with 5

μM of 1 and 1 μg mL-1 of Hoechst 33258 for 20 min. For immunofluorescence experiment, cells

were blocked with 5% bovine serum albumin/PBS at 37 °C for 1 h. After that, the cells were

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incubated with BG4 antibody at 37 °C for 2 h, followed by incubating with anti-His-tag antibody

(#12698S, Cell Signaling Technology) at 37 °C for 2 h, and then incubated with Alexa 647-

conjugated antibody (A21244, Thermo Fisher Scientific) at 37°C for 1 h. After each antibody

incubation, the cells were rinsed five times with DEPC-PBS. At last, cells were stained with 5 μM

of 1 and the emission of 1 was collected under excitation at 488 nm, and that of BG4 antibody was

collected under excitation at 640 nm. Digital images were recorded by Carl ZEISS LSM 800

confocal laser scanning microscope with a 63× objective lens.

Live cell staining experiment

The HeLa, MCF-7 and A549 cells were cultured in DMEM media, CHO (Chinese Hamster Ovary)

cells were cultured in DMEM/Ham’s F-12 media, MCF 10A cells were cultured in DMEM/Ham’s

F-12 media (containing of 0.5‰ hydrocortisone, 1‰ insulin and 2‰ EGF) and MRC-5 cells were

cultured in EMEM media. All the media used were containing 10% fetal bovine serum at 37 °C,

with 5% CO2 atmosphere. 40,000 cells were seeded into an imaging dish (ibidi, μ-dish 35 mm, high)

and cultured for 24 h. Cells were incubated with 5 μM of 1 for 24 hrs, and then rinsed by culture

medium. For the competitive assay, HeLa cells were incubated with 1 (5 μM) and different

concentrations of BRACO19 or PhenDC3 for 24 hrs at 37 °C. The cell nuclei were then stained with

Hoechst 33258 and rinsed by culture medium. Digital images were recorded by ZEISS LSM 800

confocal laser scanning microscope with a 63× objective lens and analyzed with Volocity 6.3

software (PerkinElmer Corp.).

To track the un/folding dynamics of RNA GQs in live cells, we continuously image the above-

mentioned cells over 1 minute with a 1s interval. 60 images of each cell were reconstructed into a

video by ZEN 2.3 software. The fluorescence intensity of 1 per cell ( ) in each image 𝐹𝑐𝑖, 𝑖 = 1~60𝑠

were analyzed with Volocity 6.3 software.

In order to display the dynamic folding behavior of RNA GQ in live cells, the fluorescent intensity

fluctuation of 1 in live cells were further analyzed. The fluorescence intensities of 1 per cell ( ) 𝐹𝑐𝑖

over 60 s were plotted. An ExpDec1 model from Origin 8.0 (OriginLab Corp.) was selected to fit

the data. We take the fitting curve as a reference and set the most deviated data points from the

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fitting curve as a boundary on both sides to curb the data set as a fluctuation zone. To quantify and

compare the fluorescent fluctuation differences, the standard deviation (σ) and the mean (μ) of 𝐹𝑐𝑖

in both cancerous and healthy cells in the first 40 s were further calculated by statistical analysis

with Origin 8.0, since the value of has not increased rapidly within the first 40 s. The relative 𝐹𝑐𝑖

variability (coefficient of variation) of was set as the ratio of σ to μ.𝐹𝑐𝑖

To explore and compare the folding dynamics of RNA GQ in the three pairs of live cancerous and

normal cell lines, we read each image as a 2D frame with pixels in both and axis to build a 100 𝑥 𝑦

matrix of the fluorescent intensity, where is the time index and ( is the 𝑀𝑡,𝑥𝑖,𝑦𝑗 𝑡 𝑥𝑖,𝑦𝑗 𝑖,𝑗 = 1~100)

pixel index in a single frame. We set the “ON” threshold to be the mean of all the positive elements

in the matrix:

𝑇𝑂𝑁 = ⟨𝑀𝑡,𝑥𝑖,𝑦𝑗> 0⟩

Such a threshold was set to offset the significant difference in the fluorescent intensity of 1 among

each of the cell lines. Thereafter, we can convert the matrix into a matrix of a state function 𝑀𝑡,𝑥𝑖,𝑦𝑗

with the same size as .𝑆𝑡,𝑥𝑖,𝑦𝑗 𝑀

𝑆𝑡,𝑥𝑖,𝑦𝑗= { 1, 𝑀𝑡,𝑥𝑖,𝑦𝑗

> 𝑇𝑂𝑁

0, 𝑀𝑡,𝑥𝑖,𝑦𝑗≤ 𝑇𝑂𝑁 �

For , if , while , we consider the RNA GQ have a residing (𝑥𝑖,𝑦𝑗) 𝑆𝑡,𝑥𝑖,𝑦𝑗

= 1 𝑆𝑡 ‒ 1,𝑥𝑖,𝑦𝑗= 𝑆𝑡 + 1,𝑥𝑖,𝑦𝑗

= 0

duration of 1s. The total number of such pixel points was set as the frequency count for 𝑆𝑡,𝑥𝑖,𝑦𝑗

RNA GQ with the lifetime of 1s. The counts for RNA GQ with longer lifetime were analyzed

similarly and were plotted against duration time to form a histogram of the unfolding dynamics of

RNA GQ in the six cell lines.

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Fig S1. 1H NMR spectrum of 1 (A) and 2 (B) in DMSO-d6 with variation of temperatures.

Fig S2. (A) CD melting curve of TERRA (6 μM, black) with addition 1 (3 μM, red), 2 (3 μM, blue) and 3 (3 μM, green, from Ref [1]); (B) CD melting curve of EBNA1 (3 μM, black) with addition 1 (3 μM, red), 2 (3 μM, blue) and 5 equiv. amount of 1 (15 μM, green).

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N

N

N

N

N

Pt

2+

Fig S3. Chemical structures of Pt complex 3

Fig S4. Pt complex 1 selectively binds to G-quadruplex. (A) FRET melting curves of GQ in the absence and presence of 1, with and without the excess amount of ds26. TMPyP4, a well-known GQ stabilizer, was used as a control molecule. Tm of each solution was summarized in the table. (B) Emission intensities of 1 (1 μM) in the presence of different nucleic acid with varies [NA] / [1] ratios (Left). The equilibrium dissociation constants (Kd) of 1 on different nucleic acids was obtained from the emission curves and were listed in the table (Right).

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Fig S5. CD spectrum of TERRA (6 μM) in the absence (black) and presence of 1 (3 μM, red) and 2 (3 μM, blue).

Fig S6. Emission spectrum of 2 (1 μM) in the presence of 20 equivalent amounts of GQs and ssRNA, 5 U of human HeLa total RNA (blue) and 5 U of tRNA from bovine liver (green) (λex = 450 nm).

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Fig S7. UV-vis spectra of 1 (25 μM) in the absence and presence of different RNAs (25 μM).

Fig S8. Transient absorption spectrum of 1 in buffer (pH=7).

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500 550 600 650 700 7500

25

50

75

Rel

ativ

e In

tens

ity

(nm)

1 1+GQ

Fig S9. Phosphorescent spectrum of 1 in the absence (black) and presence of GQ (red).

Fig S10. Emission spectrum of (A) 1 and (B) 2 in different solvents (λex = 450 nm).

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Fig S11. Plot of the luminescent intensity for 1 versus solvent (a mixture of methanol as glycerol in different ratios) viscosity. Logarithm of luminescent intensity were correlated with logarithm of solvent viscosity.

Fig S12. Emission spectrum of 1 μM 1 in the absence (black and blue) and presence of GQ (red and green) at different viscosities

Fig S13. (A) Steady state emission spectrum of 1 in the absence and (B) presence of GQ before (black) and after deaeration (red). (C) Transient absorption decay kinetics of 1 in the presence of GQ before (black) and after deaeration (red).

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Fig S14. Confocal imaging of fixed Hela cells stained with 1, nucleus dye Hoechst 33258, bright field and merged images before and after treatment of cells with RNase A and DNase I, respectively.

Fig S15. (A) Analysis of the values in Hela and CHO cells over 60 s-observation 𝐹𝑐time period. (B) Comparison of the relative variability (coefficient of variation) of the fluorescence of 1 in cancerous and normal cells in A.

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Fig S16. Live-cell imaging of A549, MCR-5, MCF-7 and MCF 10A cells stained with 1 (red). The nucleus were stained with Hoechst 33258 (green).

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Fig S17. Emission spectrum of 1 μM 1 in the absence (black) and presence of 20 equivalent amounts of GQ structure (red) and 1.8 g/mL of albumin (blue) (λex = 450 nm).

Fig S18. Comparison of the photostability of 1 (black) and thiazole orange (red) as GQ fluorescent probes. The luminescence intensity of 1 (black) at 580 nm and thiazole orange (red) at 535 nm under continuous excitation at 450 nm and 500 nm, respectively.

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1 10 1000

20

40

60

80

100

HeLa A549 MCF-7

Cel

l via

bilit

y (%

)[1] (M)

Fig S19. Viability of HeLa, A549 and MCF-7 cells exposed to variant concentration of 1 for 24 h determined by MTT assay.

To avoid false positive results, the endocytosis ability of different cells to 1 was investigated. As shown in Fig S20, both cancerous and healthy cells have similar endocytosis ability (around 90%) to 1, indicated that the differences of fluorescent foci number and intensity among these cell lines reflect the features of RNA GQs.

Fig S20. Investigation of endocytosis ability of different cells to 1. UV-vis spectrum of the remaining amount of ligand in the cell medium after incubation with different cell lines.

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Table S1a. Crystal data, collection and structure refinement parameters for 1.

Chemical formula C51H34F12N10OP2Pt2

Formula weight (g mol-1) 1483.00

Crystal system monoclinic

Space group P 1 21/c 1

Crystal size (mm3) 0.020 x 0.040 x 0.240

Temperature (K) 103(2)

a (Å) 6.8545(3)

b (Å) 39.3855(16)

c (Å) 17.3529(7) Å

α (deg) 90

β (deg) 98.991(2)

γ (deg) γ = 90

V (Å3) 4627.2(3)

Z 4

ρcalcd (g cm-3) 2.129

μ (mm-1) 12.723

F(000) 2848

Data / restraints / parameters 8170 / 72 / 705

R1 [I > 2σ(I)] 0.0438

wR2 [I > 2σ(I)] 0.0999

R1 (all data) 0.0472

wR2 (all data) 0.1014

Goodness-of-fit on F2 1.220

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Table S1b. Selected Bond Lengths (Å) and Bond Angles (deg) for 1.

Pt1-N1 2.062(5)

Pt1-N2 1.981(6)

Pt1-N4 1.992(5)

Pt1-N5 2.062(5)

N1-Pt1-N2 81.2(2)

N1-Pt1-N4 172.7(2)

N1-Pt1-N5 106.2(2)

N2-Pt1-N4 91.6(2)

N2-Pt1-N5 172.6(2)

N4-Pt1-N5 81.1(2)

Table S2. RNA used in this work.

Oligomer Sequence*

TERRA 5’-uaggguuagggu-3’

EBNA1[2] 5’-ggggcaggagcaggagga-3’

ssRNA 5’-gggaaggccagggaaucuuccc-3’

tRNA transfer RNA

HtRNA human HeLa cell total RNA

ds26 5’-CAATCGGATCGAATTCGATCCGATTG-3’ (a duplex DNA sequence)

F-c-myc-T 5’-FAM-TGGGGAGGGTGGGGAGGGTGGGGAAGG-TAMRA-3' (a DNA GQ sequence with 5’ and 3’ labeling)

*Low case letters are used for RNA sequences.

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Table S3. Quantum yield of 1 and 2 in the absence and presence of different RNAs.Φem

RNA1 2

without RNA 0.0241 (4) 0.0084 (4)

TERRA 0.366 (7) 0.456 (9)

EBNA1 0.312 (5) 0.378 (8)

ssRNA 0.063 (4) 0.112 (6)

HeLa total RNA 0.050 (7) 0.117 (9)

tRNA 0.073 (8) 0.134 (6)

Table S4. Quantum yield of 1 and 2 in different solvents at room temperature.

ΦemSolvent

1 2

Buffer pH=7 0.0241 (4) 0.0084 (4)

Dichloromethane 0.0173 (8) 0.0244 (9)

Acetonitrile 0.0086 (4) 0.0180 (8)

Methanol 0.0082 (3) 0.013 (1)

References[1] He, L.; Meng, Z.; Xie, Y.; Chen, X.; Li, T.; Shao, F. J. Inorg. Biochem. 2017, 166, 135.[2] Lista, M. J.; Martins, R. P.; Billant, O.; Contesse, M.-A.; Findakly, S.; Pochard, P.; Daskalogianni, C.; Beauvineau, C.; Guetta, C.; Jamin, C.; Teulade-Fichou, M.-P.; Fåhraeus, R.; Voisset, C.; Blondel, M. Nat. Commun. 2017, 8, 16043.