Supplementary Data

Biological Evaluation of Water Soluble Arene Ru(II) Enantiomers with Amino-Oxime

Ligands

Isabel de la Cueva-Aliqueǂ Sara Sierraǂ Laura Muntildeoz-Morenodagger Adriaacuten Peacuterez-Redondoǂ Ana M

Bajo dagger Isabel Marzo Lourdes Gudeǂ Tomaacutes Cuencaǂ Eva Royoǂ

ǂ Departamento de Quiacutemica Orgaacutenica y Quiacutemica Inorgaacutenica Instituto de Investigacioacuten Quiacutemica

Andreacutes M del Riacuteo (IQAR) Universidad de Alcalaacute 28805 Alcalaacute de Henares Madrid Spain

Departamento de Bioquiacutemica y Biologiacutea Molecular y Celular Universidad de Zaragoza 50009

Zaragoza Spain

dagger Departamento de Biologiacutea de Sistemas Facultad de Medicina y Ciencias de la Salud Universidad de

Alcalaacute 28805 Alcalaacute de Henares Madrid Spain

S1

Selected experimental data

1 Elemental analysis of compound 1a

2 Figures S1-S2 NMR spectra of proligands a and arsquo

3 Figures S3-S8 NMR spectra of 1a and 1arsquo

4 Figures S9 Time-dependent 1H NMR of 1arsquo in PBSD2O

5 Figures S10 UV-vis spectra of arsquomiddotHCl in water

6 Figure S11 UV-vis spectrum of 1arsquo in water

7 Figure S12 Circular dichroism spectra of amiddotHCl and arsquomiddotHCl in water

Selected crystallographic data

1 Table S1 Selected crystallographic data for 1arsquo

2 Figure S13 ORTEP drawing of 1arsquo cation

3 Table S2 Selected bond and angles for 1arsquo

Selected biological data

1 Figure S14 Morphological changes induced by compounds 1a and 1arsquo (25 microM) on A-549

cells

2 Figure S15 Effect of compounds 1a and 1arsquo on JurKatpLVTHM and Jurkat shBak cell viability

3 Figure S16 FRET DNA melting curves with a∙HCl 1a and arsquo∙HCl 1arsquo at 10 M

4 Figure S17 Effect of treatment of PC-3 cells with 1a on the tumor growth

S2

Elemental analysis CHN for 1a

Anal Calcd for C27H38Cl2N2ORu C 5605 H 662 N 484 Found C 5598 H

667 N 479

S3

Selected NMR spectra

Figure S11H NMR of a in CDCl3

Figure S21H NMR of arsquo in CDCl3

S4

Figure S31H NMR of 1a in CD3OD

Figure S41H NMR of 1arsquo in CD3OD

S5

Figure S51H NMR of 1arsquo in CDCl3

Figure S6 1H NMR of 1arsquo in D2O

S6

Figure S713C-APT NMR of 1arsquo in CD3OD

S7

Figure S815N-1H HMBC of 1arsquo in CD3OD

Figure S9 Time dependent 1H NMR of 1arsquo in a PBS solution (D2O) at pH = 740

S8

S9

Figure S10 UV-vis spectrum of arsquomiddotHCl in water (500M)

Figure S11 UV-vis spectrum of 1arsquo in water (500M)

S10

Figure S12 Circular dichroism spectra of amiddotHCl and arsquomiddotHCl in water (500 M)

-1

-08

-06

-04

-02

0

02

04

06

08

220 240 260 280 300 320Δε

λ (nm)

amiddotHCl amiddotHCl

S11

Table S1 Crystallographic data for complex 1arsquo

Compound 1arsquomiddot2CHCl3

Empirical formula C29H40Cl8N2ORu

Formula weight 81730

Temperature (K) 200(2)

Wavelength (Aring) 071073

Crystal system Monoclinic

Space group P21

a (Aring) (ordm) 8865(2)

b (Aring) (ordm) 21916(2) 9049(1)

c (Aring) (ordm) 18183(2)

Volume (Aring3) 35327(8)

Z 4

Density calculated (g cmndash3) 1537

Absorption coefficient (mmndash1) 1075

F(000) 1664

Crystal size (mm3) 031 times 013 times 012

Theta range for data collection (ordm) 300 to 2524

Index ranges ndash10 to 10 ndash26 to 26ndash21 to 21

Reflections collected 66843

Independent reflections 12759 [R(int) = 0174]

Reflections [Igt2(I)] 8747

Completeness to theta 998

Goodness-of-fit on F2 1085

Final R indices [Igt2(I)] R1 = 0071 wR2 = 0114

R indices (all data) R1 = 0132 wR2 = 0134

Largest diff peak and hole (emiddotAringndash3) 0699 and ndash0588

S12

Figure S13 ORTEP drawing of the cation in the second independent molecule of the asymmetric unit

for compound 1arsquo with 50 probability ellipsoids Hydrogen bonded to carbon atoms have been

omitted for clarity

Table S2 Selected lengths (Aring) and angles (ordm) of the two independent molecules of the asymmetric

unit for 1arsquo

Ru(1)-Cl(1) 2404(4) Ru(1)-N(11) 2067(10)

Ru(1)-N(12) 2173(10) N(11)-O(11) 1381(13)

N(11)-C(21) 1284(15) N(12)-C(22) 1496(16)

C(21)-C(22) 1503(16) Ru(1)-Ct(1)a 1674

Ru(2)-Cl(3) 2396(4) Ru(2)-N(21) 2065(11)

Ru(2)-N(22) 2156(11) N(21)-O(21) 1388(13)

N(21)-C(61) 1263(16) N(22)-C(62) 1508(17)

C(61)-C(62) 1504(17) Ru(2)-Ct(2)b 1700

Cl(1)-Ru(1)-N(11) 829(3) Cl(1)-Ru(1)-N(12) 818(3)

N(11)-Ru(1)-N(12) 758(4) Ru(1)-N(11)-C(21) 1216(8)

N(11)-C(21)-C(22) 1157(11) C(21)-C(22)-N(12) 1087(10)

Ru(1)-N(12)-C(22) 1111(7) Ru(1)-N(12)-C(31) 1206(8)

C(22)-N(12)-C(31) 1127(10) Ru(1)-N(11)-O(11) 1240(8)

C(21)-N(11)-O(11) 1144(10) Cl(3)-Ru(2)-N(21) 820(3)

Cl(3)-Ru(2)-N(22) 820(3) N(21)-Ru(2)-N(22) 751(4)

Ru(2)-N(21)-C(61) 1225(9) N(21)-C(61)-C(62) 1156(12)

C(61)-C(62)-N(22) 1075(10) Ru(2)-N(22)-C(62) 1114(7)

Ru(2)-N(22)-C(71) 1204(9) C(62)-N(22)-C(71) 1113(10)

Ru(2)-N(21)-O(21) 1236(8) C(61)-N(21)-O(21) 1139(10)a Ct(1) centroid of the C(11)-C(16) ring b Ct(2) centroid of the C(51)-C(56) ring

S13

Fig S14 Morphological changes induced by compounds 1a and 1arsquo (25 microM) A549 cells were seeded and 24 h later compounds were added at the

indicated concentrations Cell morphology was evaluated by optical microscopy after 24 h

S14

Fig S15 Effect of enantiomers 1a and 1arsquo on JurKatpLVTHM and Jurkat shBak cell viability after 24 h of exposure The results are expressed as

a percentage of live cells compared to control

S15

Figure S16 FRET DNA melting curves at 10 M of A) aHCl and 1a B) arsquoHCl and 1arsquo The concentration of F10T is 02 M

S16

Fig S17 Effect of treatment of PC-3 cells with 1a on the tumor growth Androgen-independent

prostate cancer cells were incubated in the absence or presence of 1a (25 microM) for 24 h The

cell suspension was mixed with Matrigelreg and injected subcutaneously into the right flank of

nude mice (5 x 106 cellsmouse) Ten mice were used in each group A representative tumor

sample from each group after 43 days treatment is shown P lt 005 P lt 0001 vs control

group

S17

• Z

Selected experimental data

1 Elemental analysis of compound 1a

2 Figures S1-S2 NMR spectra of proligands a and arsquo

3 Figures S3-S8 NMR spectra of 1a and 1arsquo

4 Figures S9 Time-dependent 1H NMR of 1arsquo in PBSD2O

5 Figures S10 UV-vis spectra of arsquomiddotHCl in water

6 Figure S11 UV-vis spectrum of 1arsquo in water

7 Figure S12 Circular dichroism spectra of amiddotHCl and arsquomiddotHCl in water

Selected crystallographic data

1 Table S1 Selected crystallographic data for 1arsquo

2 Figure S13 ORTEP drawing of 1arsquo cation

3 Table S2 Selected bond and angles for 1arsquo

Selected biological data

1 Figure S14 Morphological changes induced by compounds 1a and 1arsquo (25 microM) on A-549

cells

2 Figure S15 Effect of compounds 1a and 1arsquo on JurKatpLVTHM and Jurkat shBak cell viability

3 Figure S16 FRET DNA melting curves with a∙HCl 1a and arsquo∙HCl 1arsquo at 10 M

4 Figure S17 Effect of treatment of PC-3 cells with 1a on the tumor growth

S2

Elemental analysis CHN for 1a

Anal Calcd for C27H38Cl2N2ORu C 5605 H 662 N 484 Found C 5598 H

667 N 479

S3

Selected NMR spectra

Figure S11H NMR of a in CDCl3

Figure S21H NMR of arsquo in CDCl3

S4

Figure S31H NMR of 1a in CD3OD

Figure S41H NMR of 1arsquo in CD3OD

S5

Figure S51H NMR of 1arsquo in CDCl3

Figure S6 1H NMR of 1arsquo in D2O

S6

Figure S713C-APT NMR of 1arsquo in CD3OD

S7

Figure S815N-1H HMBC of 1arsquo in CD3OD

Figure S9 Time dependent 1H NMR of 1arsquo in a PBS solution (D2O) at pH = 740

S8

S9

Figure S10 UV-vis spectrum of arsquomiddotHCl in water (500M)

Figure S11 UV-vis spectrum of 1arsquo in water (500M)

S10

Figure S12 Circular dichroism spectra of amiddotHCl and arsquomiddotHCl in water (500 M)

-1

-08

-06

-04

-02

0

02

04

06

08

220 240 260 280 300 320Δε

λ (nm)

amiddotHCl amiddotHCl

S11

Table S1 Crystallographic data for complex 1arsquo

Compound 1arsquomiddot2CHCl3

Empirical formula C29H40Cl8N2ORu

Formula weight 81730

Temperature (K) 200(2)

Wavelength (Aring) 071073

Crystal system Monoclinic

Space group P21

a (Aring) (ordm) 8865(2)

b (Aring) (ordm) 21916(2) 9049(1)

c (Aring) (ordm) 18183(2)

Volume (Aring3) 35327(8)

Z 4

Density calculated (g cmndash3) 1537

Absorption coefficient (mmndash1) 1075

F(000) 1664

Crystal size (mm3) 031 times 013 times 012

Theta range for data collection (ordm) 300 to 2524

Index ranges ndash10 to 10 ndash26 to 26ndash21 to 21

Reflections collected 66843

Independent reflections 12759 [R(int) = 0174]

Reflections [Igt2(I)] 8747

Completeness to theta 998

Goodness-of-fit on F2 1085

Final R indices [Igt2(I)] R1 = 0071 wR2 = 0114

R indices (all data) R1 = 0132 wR2 = 0134

Largest diff peak and hole (emiddotAringndash3) 0699 and ndash0588

S12

Figure S13 ORTEP drawing of the cation in the second independent molecule of the asymmetric unit

for compound 1arsquo with 50 probability ellipsoids Hydrogen bonded to carbon atoms have been

omitted for clarity

Table S2 Selected lengths (Aring) and angles (ordm) of the two independent molecules of the asymmetric

unit for 1arsquo

Ru(1)-Cl(1) 2404(4) Ru(1)-N(11) 2067(10)

Ru(1)-N(12) 2173(10) N(11)-O(11) 1381(13)

N(11)-C(21) 1284(15) N(12)-C(22) 1496(16)

C(21)-C(22) 1503(16) Ru(1)-Ct(1)a 1674

Ru(2)-Cl(3) 2396(4) Ru(2)-N(21) 2065(11)

Ru(2)-N(22) 2156(11) N(21)-O(21) 1388(13)

N(21)-C(61) 1263(16) N(22)-C(62) 1508(17)

C(61)-C(62) 1504(17) Ru(2)-Ct(2)b 1700

Cl(1)-Ru(1)-N(11) 829(3) Cl(1)-Ru(1)-N(12) 818(3)

N(11)-Ru(1)-N(12) 758(4) Ru(1)-N(11)-C(21) 1216(8)

N(11)-C(21)-C(22) 1157(11) C(21)-C(22)-N(12) 1087(10)

Ru(1)-N(12)-C(22) 1111(7) Ru(1)-N(12)-C(31) 1206(8)

C(22)-N(12)-C(31) 1127(10) Ru(1)-N(11)-O(11) 1240(8)

C(21)-N(11)-O(11) 1144(10) Cl(3)-Ru(2)-N(21) 820(3)

Cl(3)-Ru(2)-N(22) 820(3) N(21)-Ru(2)-N(22) 751(4)

Ru(2)-N(21)-C(61) 1225(9) N(21)-C(61)-C(62) 1156(12)

C(61)-C(62)-N(22) 1075(10) Ru(2)-N(22)-C(62) 1114(7)

Ru(2)-N(22)-C(71) 1204(9) C(62)-N(22)-C(71) 1113(10)

Ru(2)-N(21)-O(21) 1236(8) C(61)-N(21)-O(21) 1139(10)a Ct(1) centroid of the C(11)-C(16) ring b Ct(2) centroid of the C(51)-C(56) ring

S13

Fig S14 Morphological changes induced by compounds 1a and 1arsquo (25 microM) A549 cells were seeded and 24 h later compounds were added at the

indicated concentrations Cell morphology was evaluated by optical microscopy after 24 h

S14

Fig S15 Effect of enantiomers 1a and 1arsquo on JurKatpLVTHM and Jurkat shBak cell viability after 24 h of exposure The results are expressed as

a percentage of live cells compared to control

S15

Figure S16 FRET DNA melting curves at 10 M of A) aHCl and 1a B) arsquoHCl and 1arsquo The concentration of F10T is 02 M

S16

Fig S17 Effect of treatment of PC-3 cells with 1a on the tumor growth Androgen-independent

prostate cancer cells were incubated in the absence or presence of 1a (25 microM) for 24 h The

cell suspension was mixed with Matrigelreg and injected subcutaneously into the right flank of

nude mice (5 x 106 cellsmouse) Ten mice were used in each group A representative tumor

sample from each group after 43 days treatment is shown P lt 005 P lt 0001 vs control

group

S17

• Z

Elemental analysis CHN for 1a

Anal Calcd for C27H38Cl2N2ORu C 5605 H 662 N 484 Found C 5598 H

667 N 479

S3

Selected NMR spectra

Figure S11H NMR of a in CDCl3

Figure S21H NMR of arsquo in CDCl3

S4

Figure S31H NMR of 1a in CD3OD

Figure S41H NMR of 1arsquo in CD3OD

S5

Figure S51H NMR of 1arsquo in CDCl3

Figure S6 1H NMR of 1arsquo in D2O

S6

Figure S713C-APT NMR of 1arsquo in CD3OD

S7

Figure S815N-1H HMBC of 1arsquo in CD3OD

Figure S9 Time dependent 1H NMR of 1arsquo in a PBS solution (D2O) at pH = 740

S8

S9

Figure S10 UV-vis spectrum of arsquomiddotHCl in water (500M)

Figure S11 UV-vis spectrum of 1arsquo in water (500M)

S10

Figure S12 Circular dichroism spectra of amiddotHCl and arsquomiddotHCl in water (500 M)

-1

-08

-06

-04

-02

0

02

04

06

08

220 240 260 280 300 320Δε

λ (nm)

amiddotHCl amiddotHCl

S11

Table S1 Crystallographic data for complex 1arsquo

Compound 1arsquomiddot2CHCl3

Empirical formula C29H40Cl8N2ORu

Formula weight 81730

Temperature (K) 200(2)

Wavelength (Aring) 071073

Crystal system Monoclinic

Space group P21

a (Aring) (ordm) 8865(2)

b (Aring) (ordm) 21916(2) 9049(1)

c (Aring) (ordm) 18183(2)

Volume (Aring3) 35327(8)

Z 4

Density calculated (g cmndash3) 1537

Absorption coefficient (mmndash1) 1075

F(000) 1664

Crystal size (mm3) 031 times 013 times 012

Theta range for data collection (ordm) 300 to 2524

Index ranges ndash10 to 10 ndash26 to 26ndash21 to 21

Reflections collected 66843

Independent reflections 12759 [R(int) = 0174]

Reflections [Igt2(I)] 8747

Completeness to theta 998

Goodness-of-fit on F2 1085

Final R indices [Igt2(I)] R1 = 0071 wR2 = 0114

R indices (all data) R1 = 0132 wR2 = 0134

Largest diff peak and hole (emiddotAringndash3) 0699 and ndash0588

S12

Figure S13 ORTEP drawing of the cation in the second independent molecule of the asymmetric unit

for compound 1arsquo with 50 probability ellipsoids Hydrogen bonded to carbon atoms have been

omitted for clarity

Table S2 Selected lengths (Aring) and angles (ordm) of the two independent molecules of the asymmetric

unit for 1arsquo

Ru(1)-Cl(1) 2404(4) Ru(1)-N(11) 2067(10)

Ru(1)-N(12) 2173(10) N(11)-O(11) 1381(13)

N(11)-C(21) 1284(15) N(12)-C(22) 1496(16)

C(21)-C(22) 1503(16) Ru(1)-Ct(1)a 1674

Ru(2)-Cl(3) 2396(4) Ru(2)-N(21) 2065(11)

Ru(2)-N(22) 2156(11) N(21)-O(21) 1388(13)

N(21)-C(61) 1263(16) N(22)-C(62) 1508(17)

C(61)-C(62) 1504(17) Ru(2)-Ct(2)b 1700

Cl(1)-Ru(1)-N(11) 829(3) Cl(1)-Ru(1)-N(12) 818(3)

N(11)-Ru(1)-N(12) 758(4) Ru(1)-N(11)-C(21) 1216(8)

N(11)-C(21)-C(22) 1157(11) C(21)-C(22)-N(12) 1087(10)

Ru(1)-N(12)-C(22) 1111(7) Ru(1)-N(12)-C(31) 1206(8)

C(22)-N(12)-C(31) 1127(10) Ru(1)-N(11)-O(11) 1240(8)

C(21)-N(11)-O(11) 1144(10) Cl(3)-Ru(2)-N(21) 820(3)

Cl(3)-Ru(2)-N(22) 820(3) N(21)-Ru(2)-N(22) 751(4)

Ru(2)-N(21)-C(61) 1225(9) N(21)-C(61)-C(62) 1156(12)

C(61)-C(62)-N(22) 1075(10) Ru(2)-N(22)-C(62) 1114(7)

Ru(2)-N(22)-C(71) 1204(9) C(62)-N(22)-C(71) 1113(10)

Ru(2)-N(21)-O(21) 1236(8) C(61)-N(21)-O(21) 1139(10)a Ct(1) centroid of the C(11)-C(16) ring b Ct(2) centroid of the C(51)-C(56) ring

S13

Fig S14 Morphological changes induced by compounds 1a and 1arsquo (25 microM) A549 cells were seeded and 24 h later compounds were added at the

indicated concentrations Cell morphology was evaluated by optical microscopy after 24 h

S14

Fig S15 Effect of enantiomers 1a and 1arsquo on JurKatpLVTHM and Jurkat shBak cell viability after 24 h of exposure The results are expressed as

a percentage of live cells compared to control

S15

Figure S16 FRET DNA melting curves at 10 M of A) aHCl and 1a B) arsquoHCl and 1arsquo The concentration of F10T is 02 M

S16

Fig S17 Effect of treatment of PC-3 cells with 1a on the tumor growth Androgen-independent

prostate cancer cells were incubated in the absence or presence of 1a (25 microM) for 24 h The

cell suspension was mixed with Matrigelreg and injected subcutaneously into the right flank of

nude mice (5 x 106 cellsmouse) Ten mice were used in each group A representative tumor

sample from each group after 43 days treatment is shown P lt 005 P lt 0001 vs control

group

S17

• Z

Selected NMR spectra

Figure S11H NMR of a in CDCl3

Figure S21H NMR of arsquo in CDCl3

S4

Figure S31H NMR of 1a in CD3OD

Figure S41H NMR of 1arsquo in CD3OD

S5

Figure S51H NMR of 1arsquo in CDCl3

Figure S6 1H NMR of 1arsquo in D2O

S6

Figure S713C-APT NMR of 1arsquo in CD3OD

S7

Figure S815N-1H HMBC of 1arsquo in CD3OD

Figure S9 Time dependent 1H NMR of 1arsquo in a PBS solution (D2O) at pH = 740

S8

S9

Figure S10 UV-vis spectrum of arsquomiddotHCl in water (500M)

Figure S11 UV-vis spectrum of 1arsquo in water (500M)

S10

Figure S12 Circular dichroism spectra of amiddotHCl and arsquomiddotHCl in water (500 M)

-1

-08

-06

-04

-02

0

02

04

06

08

220 240 260 280 300 320Δε

λ (nm)

amiddotHCl amiddotHCl

S11

Table S1 Crystallographic data for complex 1arsquo

Compound 1arsquomiddot2CHCl3

Empirical formula C29H40Cl8N2ORu

Formula weight 81730

Temperature (K) 200(2)

Wavelength (Aring) 071073

Crystal system Monoclinic

Space group P21

a (Aring) (ordm) 8865(2)

b (Aring) (ordm) 21916(2) 9049(1)

c (Aring) (ordm) 18183(2)

Volume (Aring3) 35327(8)

Z 4

Density calculated (g cmndash3) 1537

Absorption coefficient (mmndash1) 1075

F(000) 1664

Crystal size (mm3) 031 times 013 times 012

Theta range for data collection (ordm) 300 to 2524

Index ranges ndash10 to 10 ndash26 to 26ndash21 to 21

Reflections collected 66843

Independent reflections 12759 [R(int) = 0174]

Reflections [Igt2(I)] 8747

Completeness to theta 998

Goodness-of-fit on F2 1085

Final R indices [Igt2(I)] R1 = 0071 wR2 = 0114

R indices (all data) R1 = 0132 wR2 = 0134

Largest diff peak and hole (emiddotAringndash3) 0699 and ndash0588

S12

Figure S13 ORTEP drawing of the cation in the second independent molecule of the asymmetric unit

for compound 1arsquo with 50 probability ellipsoids Hydrogen bonded to carbon atoms have been

omitted for clarity

Table S2 Selected lengths (Aring) and angles (ordm) of the two independent molecules of the asymmetric

unit for 1arsquo

Ru(1)-Cl(1) 2404(4) Ru(1)-N(11) 2067(10)

Ru(1)-N(12) 2173(10) N(11)-O(11) 1381(13)

N(11)-C(21) 1284(15) N(12)-C(22) 1496(16)

C(21)-C(22) 1503(16) Ru(1)-Ct(1)a 1674

Ru(2)-Cl(3) 2396(4) Ru(2)-N(21) 2065(11)

Ru(2)-N(22) 2156(11) N(21)-O(21) 1388(13)

N(21)-C(61) 1263(16) N(22)-C(62) 1508(17)

C(61)-C(62) 1504(17) Ru(2)-Ct(2)b 1700

Cl(1)-Ru(1)-N(11) 829(3) Cl(1)-Ru(1)-N(12) 818(3)

N(11)-Ru(1)-N(12) 758(4) Ru(1)-N(11)-C(21) 1216(8)

N(11)-C(21)-C(22) 1157(11) C(21)-C(22)-N(12) 1087(10)

Ru(1)-N(12)-C(22) 1111(7) Ru(1)-N(12)-C(31) 1206(8)

C(22)-N(12)-C(31) 1127(10) Ru(1)-N(11)-O(11) 1240(8)

C(21)-N(11)-O(11) 1144(10) Cl(3)-Ru(2)-N(21) 820(3)

Cl(3)-Ru(2)-N(22) 820(3) N(21)-Ru(2)-N(22) 751(4)

Ru(2)-N(21)-C(61) 1225(9) N(21)-C(61)-C(62) 1156(12)

C(61)-C(62)-N(22) 1075(10) Ru(2)-N(22)-C(62) 1114(7)

Ru(2)-N(22)-C(71) 1204(9) C(62)-N(22)-C(71) 1113(10)

Ru(2)-N(21)-O(21) 1236(8) C(61)-N(21)-O(21) 1139(10)a Ct(1) centroid of the C(11)-C(16) ring b Ct(2) centroid of the C(51)-C(56) ring

S13

Fig S14 Morphological changes induced by compounds 1a and 1arsquo (25 microM) A549 cells were seeded and 24 h later compounds were added at the

indicated concentrations Cell morphology was evaluated by optical microscopy after 24 h

S14

Fig S15 Effect of enantiomers 1a and 1arsquo on JurKatpLVTHM and Jurkat shBak cell viability after 24 h of exposure The results are expressed as

a percentage of live cells compared to control

S15

Figure S16 FRET DNA melting curves at 10 M of A) aHCl and 1a B) arsquoHCl and 1arsquo The concentration of F10T is 02 M

S16

Fig S17 Effect of treatment of PC-3 cells with 1a on the tumor growth Androgen-independent

prostate cancer cells were incubated in the absence or presence of 1a (25 microM) for 24 h The

cell suspension was mixed with Matrigelreg and injected subcutaneously into the right flank of

nude mice (5 x 106 cellsmouse) Ten mice were used in each group A representative tumor

sample from each group after 43 days treatment is shown P lt 005 P lt 0001 vs control

group

S17

• Z

Figure S31H NMR of 1a in CD3OD

Figure S41H NMR of 1arsquo in CD3OD

S5

Figure S51H NMR of 1arsquo in CDCl3

Figure S6 1H NMR of 1arsquo in D2O

S6

Figure S713C-APT NMR of 1arsquo in CD3OD

S7

Figure S815N-1H HMBC of 1arsquo in CD3OD

Figure S9 Time dependent 1H NMR of 1arsquo in a PBS solution (D2O) at pH = 740

S8

S9

Figure S10 UV-vis spectrum of arsquomiddotHCl in water (500M)

Figure S11 UV-vis spectrum of 1arsquo in water (500M)

S10

Figure S12 Circular dichroism spectra of amiddotHCl and arsquomiddotHCl in water (500 M)

-1

-08

-06

-04

-02

0

02

04

06

08

220 240 260 280 300 320Δε

λ (nm)

amiddotHCl amiddotHCl

S11

Table S1 Crystallographic data for complex 1arsquo

Compound 1arsquomiddot2CHCl3

Empirical formula C29H40Cl8N2ORu

Formula weight 81730

Temperature (K) 200(2)

Wavelength (Aring) 071073

Crystal system Monoclinic

Space group P21

a (Aring) (ordm) 8865(2)

b (Aring) (ordm) 21916(2) 9049(1)

c (Aring) (ordm) 18183(2)

Volume (Aring3) 35327(8)

Z 4

Density calculated (g cmndash3) 1537

Absorption coefficient (mmndash1) 1075

F(000) 1664

Crystal size (mm3) 031 times 013 times 012

Theta range for data collection (ordm) 300 to 2524

Index ranges ndash10 to 10 ndash26 to 26ndash21 to 21

Reflections collected 66843

Independent reflections 12759 [R(int) = 0174]

Reflections [Igt2(I)] 8747

Completeness to theta 998

Goodness-of-fit on F2 1085

Final R indices [Igt2(I)] R1 = 0071 wR2 = 0114

R indices (all data) R1 = 0132 wR2 = 0134

Largest diff peak and hole (emiddotAringndash3) 0699 and ndash0588

S12

Figure S13 ORTEP drawing of the cation in the second independent molecule of the asymmetric unit

for compound 1arsquo with 50 probability ellipsoids Hydrogen bonded to carbon atoms have been

omitted for clarity

Table S2 Selected lengths (Aring) and angles (ordm) of the two independent molecules of the asymmetric

unit for 1arsquo

Ru(1)-Cl(1) 2404(4) Ru(1)-N(11) 2067(10)

Ru(1)-N(12) 2173(10) N(11)-O(11) 1381(13)

N(11)-C(21) 1284(15) N(12)-C(22) 1496(16)

C(21)-C(22) 1503(16) Ru(1)-Ct(1)a 1674

Ru(2)-Cl(3) 2396(4) Ru(2)-N(21) 2065(11)

Ru(2)-N(22) 2156(11) N(21)-O(21) 1388(13)

N(21)-C(61) 1263(16) N(22)-C(62) 1508(17)

C(61)-C(62) 1504(17) Ru(2)-Ct(2)b 1700

Cl(1)-Ru(1)-N(11) 829(3) Cl(1)-Ru(1)-N(12) 818(3)

N(11)-Ru(1)-N(12) 758(4) Ru(1)-N(11)-C(21) 1216(8)

N(11)-C(21)-C(22) 1157(11) C(21)-C(22)-N(12) 1087(10)

Ru(1)-N(12)-C(22) 1111(7) Ru(1)-N(12)-C(31) 1206(8)

C(22)-N(12)-C(31) 1127(10) Ru(1)-N(11)-O(11) 1240(8)

C(21)-N(11)-O(11) 1144(10) Cl(3)-Ru(2)-N(21) 820(3)

Cl(3)-Ru(2)-N(22) 820(3) N(21)-Ru(2)-N(22) 751(4)

Ru(2)-N(21)-C(61) 1225(9) N(21)-C(61)-C(62) 1156(12)

C(61)-C(62)-N(22) 1075(10) Ru(2)-N(22)-C(62) 1114(7)

Ru(2)-N(22)-C(71) 1204(9) C(62)-N(22)-C(71) 1113(10)

Ru(2)-N(21)-O(21) 1236(8) C(61)-N(21)-O(21) 1139(10)a Ct(1) centroid of the C(11)-C(16) ring b Ct(2) centroid of the C(51)-C(56) ring

S13

Fig S14 Morphological changes induced by compounds 1a and 1arsquo (25 microM) A549 cells were seeded and 24 h later compounds were added at the

indicated concentrations Cell morphology was evaluated by optical microscopy after 24 h

S14

Fig S15 Effect of enantiomers 1a and 1arsquo on JurKatpLVTHM and Jurkat shBak cell viability after 24 h of exposure The results are expressed as

a percentage of live cells compared to control

S15

Figure S16 FRET DNA melting curves at 10 M of A) aHCl and 1a B) arsquoHCl and 1arsquo The concentration of F10T is 02 M

S16

Fig S17 Effect of treatment of PC-3 cells with 1a on the tumor growth Androgen-independent

prostate cancer cells were incubated in the absence or presence of 1a (25 microM) for 24 h The

cell suspension was mixed with Matrigelreg and injected subcutaneously into the right flank of

nude mice (5 x 106 cellsmouse) Ten mice were used in each group A representative tumor

sample from each group after 43 days treatment is shown P lt 005 P lt 0001 vs control

group

S17

• Z

Figure S51H NMR of 1arsquo in CDCl3

Figure S6 1H NMR of 1arsquo in D2O

S6

Figure S713C-APT NMR of 1arsquo in CD3OD

S7

Figure S815N-1H HMBC of 1arsquo in CD3OD

Figure S9 Time dependent 1H NMR of 1arsquo in a PBS solution (D2O) at pH = 740

S8

S9

Figure S10 UV-vis spectrum of arsquomiddotHCl in water (500M)

Figure S11 UV-vis spectrum of 1arsquo in water (500M)

S10

Figure S12 Circular dichroism spectra of amiddotHCl and arsquomiddotHCl in water (500 M)

-1

-08

-06

-04

-02

0

02

04

06

08

220 240 260 280 300 320Δε

λ (nm)

amiddotHCl amiddotHCl

S11

Table S1 Crystallographic data for complex 1arsquo

Compound 1arsquomiddot2CHCl3

Empirical formula C29H40Cl8N2ORu

Formula weight 81730

Temperature (K) 200(2)

Wavelength (Aring) 071073

Crystal system Monoclinic

Space group P21

a (Aring) (ordm) 8865(2)

b (Aring) (ordm) 21916(2) 9049(1)

c (Aring) (ordm) 18183(2)

Volume (Aring3) 35327(8)

Z 4

Density calculated (g cmndash3) 1537

Absorption coefficient (mmndash1) 1075

F(000) 1664

Crystal size (mm3) 031 times 013 times 012

Theta range for data collection (ordm) 300 to 2524

Index ranges ndash10 to 10 ndash26 to 26ndash21 to 21

Reflections collected 66843

Independent reflections 12759 [R(int) = 0174]

Reflections [Igt2(I)] 8747

Completeness to theta 998

Goodness-of-fit on F2 1085

Final R indices [Igt2(I)] R1 = 0071 wR2 = 0114

R indices (all data) R1 = 0132 wR2 = 0134

Largest diff peak and hole (emiddotAringndash3) 0699 and ndash0588

S12

Figure S13 ORTEP drawing of the cation in the second independent molecule of the asymmetric unit

for compound 1arsquo with 50 probability ellipsoids Hydrogen bonded to carbon atoms have been

omitted for clarity

Table S2 Selected lengths (Aring) and angles (ordm) of the two independent molecules of the asymmetric

unit for 1arsquo

Ru(1)-Cl(1) 2404(4) Ru(1)-N(11) 2067(10)

Ru(1)-N(12) 2173(10) N(11)-O(11) 1381(13)

N(11)-C(21) 1284(15) N(12)-C(22) 1496(16)

C(21)-C(22) 1503(16) Ru(1)-Ct(1)a 1674

Ru(2)-Cl(3) 2396(4) Ru(2)-N(21) 2065(11)

Ru(2)-N(22) 2156(11) N(21)-O(21) 1388(13)

N(21)-C(61) 1263(16) N(22)-C(62) 1508(17)

C(61)-C(62) 1504(17) Ru(2)-Ct(2)b 1700

Cl(1)-Ru(1)-N(11) 829(3) Cl(1)-Ru(1)-N(12) 818(3)

N(11)-Ru(1)-N(12) 758(4) Ru(1)-N(11)-C(21) 1216(8)

N(11)-C(21)-C(22) 1157(11) C(21)-C(22)-N(12) 1087(10)

Ru(1)-N(12)-C(22) 1111(7) Ru(1)-N(12)-C(31) 1206(8)

C(22)-N(12)-C(31) 1127(10) Ru(1)-N(11)-O(11) 1240(8)

C(21)-N(11)-O(11) 1144(10) Cl(3)-Ru(2)-N(21) 820(3)

Cl(3)-Ru(2)-N(22) 820(3) N(21)-Ru(2)-N(22) 751(4)

Ru(2)-N(21)-C(61) 1225(9) N(21)-C(61)-C(62) 1156(12)

C(61)-C(62)-N(22) 1075(10) Ru(2)-N(22)-C(62) 1114(7)

Ru(2)-N(22)-C(71) 1204(9) C(62)-N(22)-C(71) 1113(10)

Ru(2)-N(21)-O(21) 1236(8) C(61)-N(21)-O(21) 1139(10)a Ct(1) centroid of the C(11)-C(16) ring b Ct(2) centroid of the C(51)-C(56) ring

S13

Fig S14 Morphological changes induced by compounds 1a and 1arsquo (25 microM) A549 cells were seeded and 24 h later compounds were added at the

indicated concentrations Cell morphology was evaluated by optical microscopy after 24 h

S14

Fig S15 Effect of enantiomers 1a and 1arsquo on JurKatpLVTHM and Jurkat shBak cell viability after 24 h of exposure The results are expressed as

a percentage of live cells compared to control

S15

Figure S16 FRET DNA melting curves at 10 M of A) aHCl and 1a B) arsquoHCl and 1arsquo The concentration of F10T is 02 M

S16

Fig S17 Effect of treatment of PC-3 cells with 1a on the tumor growth Androgen-independent

prostate cancer cells were incubated in the absence or presence of 1a (25 microM) for 24 h The

cell suspension was mixed with Matrigelreg and injected subcutaneously into the right flank of

nude mice (5 x 106 cellsmouse) Ten mice were used in each group A representative tumor

sample from each group after 43 days treatment is shown P lt 005 P lt 0001 vs control

group

S17

• Z

Figure S713C-APT NMR of 1arsquo in CD3OD

S7

Figure S815N-1H HMBC of 1arsquo in CD3OD

Figure S9 Time dependent 1H NMR of 1arsquo in a PBS solution (D2O) at pH = 740

S8

S9

Figure S10 UV-vis spectrum of arsquomiddotHCl in water (500M)

Figure S11 UV-vis spectrum of 1arsquo in water (500M)

S10

Figure S12 Circular dichroism spectra of amiddotHCl and arsquomiddotHCl in water (500 M)

-1

-08

-06

-04

-02

0

02

04

06

08

220 240 260 280 300 320Δε

λ (nm)

amiddotHCl amiddotHCl

S11

Table S1 Crystallographic data for complex 1arsquo

Compound 1arsquomiddot2CHCl3

Empirical formula C29H40Cl8N2ORu

Formula weight 81730

Temperature (K) 200(2)

Wavelength (Aring) 071073

Crystal system Monoclinic

Space group P21

a (Aring) (ordm) 8865(2)

b (Aring) (ordm) 21916(2) 9049(1)

c (Aring) (ordm) 18183(2)

Volume (Aring3) 35327(8)

Z 4

Density calculated (g cmndash3) 1537

Absorption coefficient (mmndash1) 1075

F(000) 1664

Crystal size (mm3) 031 times 013 times 012

Theta range for data collection (ordm) 300 to 2524

Index ranges ndash10 to 10 ndash26 to 26ndash21 to 21

Reflections collected 66843

Independent reflections 12759 [R(int) = 0174]

Reflections [Igt2(I)] 8747

Completeness to theta 998

Goodness-of-fit on F2 1085

Final R indices [Igt2(I)] R1 = 0071 wR2 = 0114

R indices (all data) R1 = 0132 wR2 = 0134

Largest diff peak and hole (emiddotAringndash3) 0699 and ndash0588

S12

Figure S13 ORTEP drawing of the cation in the second independent molecule of the asymmetric unit

for compound 1arsquo with 50 probability ellipsoids Hydrogen bonded to carbon atoms have been

omitted for clarity

Table S2 Selected lengths (Aring) and angles (ordm) of the two independent molecules of the asymmetric

unit for 1arsquo

Ru(1)-Cl(1) 2404(4) Ru(1)-N(11) 2067(10)

Ru(1)-N(12) 2173(10) N(11)-O(11) 1381(13)

N(11)-C(21) 1284(15) N(12)-C(22) 1496(16)

C(21)-C(22) 1503(16) Ru(1)-Ct(1)a 1674

Ru(2)-Cl(3) 2396(4) Ru(2)-N(21) 2065(11)

Ru(2)-N(22) 2156(11) N(21)-O(21) 1388(13)

N(21)-C(61) 1263(16) N(22)-C(62) 1508(17)

C(61)-C(62) 1504(17) Ru(2)-Ct(2)b 1700

Cl(1)-Ru(1)-N(11) 829(3) Cl(1)-Ru(1)-N(12) 818(3)

N(11)-Ru(1)-N(12) 758(4) Ru(1)-N(11)-C(21) 1216(8)

N(11)-C(21)-C(22) 1157(11) C(21)-C(22)-N(12) 1087(10)

Ru(1)-N(12)-C(22) 1111(7) Ru(1)-N(12)-C(31) 1206(8)

C(22)-N(12)-C(31) 1127(10) Ru(1)-N(11)-O(11) 1240(8)

C(21)-N(11)-O(11) 1144(10) Cl(3)-Ru(2)-N(21) 820(3)

Cl(3)-Ru(2)-N(22) 820(3) N(21)-Ru(2)-N(22) 751(4)

Ru(2)-N(21)-C(61) 1225(9) N(21)-C(61)-C(62) 1156(12)

C(61)-C(62)-N(22) 1075(10) Ru(2)-N(22)-C(62) 1114(7)

Ru(2)-N(22)-C(71) 1204(9) C(62)-N(22)-C(71) 1113(10)

Ru(2)-N(21)-O(21) 1236(8) C(61)-N(21)-O(21) 1139(10)a Ct(1) centroid of the C(11)-C(16) ring b Ct(2) centroid of the C(51)-C(56) ring

S13

Fig S14 Morphological changes induced by compounds 1a and 1arsquo (25 microM) A549 cells were seeded and 24 h later compounds were added at the

indicated concentrations Cell morphology was evaluated by optical microscopy after 24 h

S14

Fig S15 Effect of enantiomers 1a and 1arsquo on JurKatpLVTHM and Jurkat shBak cell viability after 24 h of exposure The results are expressed as

a percentage of live cells compared to control

S15

Figure S16 FRET DNA melting curves at 10 M of A) aHCl and 1a B) arsquoHCl and 1arsquo The concentration of F10T is 02 M

S16

Fig S17 Effect of treatment of PC-3 cells with 1a on the tumor growth Androgen-independent

prostate cancer cells were incubated in the absence or presence of 1a (25 microM) for 24 h The

cell suspension was mixed with Matrigelreg and injected subcutaneously into the right flank of

nude mice (5 x 106 cellsmouse) Ten mice were used in each group A representative tumor

sample from each group after 43 days treatment is shown P lt 005 P lt 0001 vs control

group

S17

• Z

Figure S815N-1H HMBC of 1arsquo in CD3OD

Figure S9 Time dependent 1H NMR of 1arsquo in a PBS solution (D2O) at pH = 740

S8

S9

Figure S10 UV-vis spectrum of arsquomiddotHCl in water (500M)

Figure S11 UV-vis spectrum of 1arsquo in water (500M)

S10

Figure S12 Circular dichroism spectra of amiddotHCl and arsquomiddotHCl in water (500 M)

-1

-08

-06

-04

-02

0

02

04

06

08

220 240 260 280 300 320Δε

λ (nm)

amiddotHCl amiddotHCl

S11

Table S1 Crystallographic data for complex 1arsquo

Compound 1arsquomiddot2CHCl3

Empirical formula C29H40Cl8N2ORu

Formula weight 81730

Temperature (K) 200(2)

Wavelength (Aring) 071073

Crystal system Monoclinic

Space group P21

a (Aring) (ordm) 8865(2)

b (Aring) (ordm) 21916(2) 9049(1)

c (Aring) (ordm) 18183(2)

Volume (Aring3) 35327(8)

Z 4

Density calculated (g cmndash3) 1537

Absorption coefficient (mmndash1) 1075

F(000) 1664

Crystal size (mm3) 031 times 013 times 012

Theta range for data collection (ordm) 300 to 2524

Index ranges ndash10 to 10 ndash26 to 26ndash21 to 21

Reflections collected 66843

Independent reflections 12759 [R(int) = 0174]

Reflections [Igt2(I)] 8747

Completeness to theta 998

Goodness-of-fit on F2 1085

Final R indices [Igt2(I)] R1 = 0071 wR2 = 0114

R indices (all data) R1 = 0132 wR2 = 0134

Largest diff peak and hole (emiddotAringndash3) 0699 and ndash0588

S12

Figure S13 ORTEP drawing of the cation in the second independent molecule of the asymmetric unit

for compound 1arsquo with 50 probability ellipsoids Hydrogen bonded to carbon atoms have been

omitted for clarity

Table S2 Selected lengths (Aring) and angles (ordm) of the two independent molecules of the asymmetric

unit for 1arsquo

Ru(1)-Cl(1) 2404(4) Ru(1)-N(11) 2067(10)

Ru(1)-N(12) 2173(10) N(11)-O(11) 1381(13)

N(11)-C(21) 1284(15) N(12)-C(22) 1496(16)

C(21)-C(22) 1503(16) Ru(1)-Ct(1)a 1674

Ru(2)-Cl(3) 2396(4) Ru(2)-N(21) 2065(11)

Ru(2)-N(22) 2156(11) N(21)-O(21) 1388(13)

N(21)-C(61) 1263(16) N(22)-C(62) 1508(17)

C(61)-C(62) 1504(17) Ru(2)-Ct(2)b 1700

Cl(1)-Ru(1)-N(11) 829(3) Cl(1)-Ru(1)-N(12) 818(3)

N(11)-Ru(1)-N(12) 758(4) Ru(1)-N(11)-C(21) 1216(8)

N(11)-C(21)-C(22) 1157(11) C(21)-C(22)-N(12) 1087(10)

Ru(1)-N(12)-C(22) 1111(7) Ru(1)-N(12)-C(31) 1206(8)

C(22)-N(12)-C(31) 1127(10) Ru(1)-N(11)-O(11) 1240(8)

C(21)-N(11)-O(11) 1144(10) Cl(3)-Ru(2)-N(21) 820(3)

Cl(3)-Ru(2)-N(22) 820(3) N(21)-Ru(2)-N(22) 751(4)

Ru(2)-N(21)-C(61) 1225(9) N(21)-C(61)-C(62) 1156(12)

C(61)-C(62)-N(22) 1075(10) Ru(2)-N(22)-C(62) 1114(7)

Ru(2)-N(22)-C(71) 1204(9) C(62)-N(22)-C(71) 1113(10)

Ru(2)-N(21)-O(21) 1236(8) C(61)-N(21)-O(21) 1139(10)a Ct(1) centroid of the C(11)-C(16) ring b Ct(2) centroid of the C(51)-C(56) ring

S13

Fig S14 Morphological changes induced by compounds 1a and 1arsquo (25 microM) A549 cells were seeded and 24 h later compounds were added at the

indicated concentrations Cell morphology was evaluated by optical microscopy after 24 h

S14

Fig S15 Effect of enantiomers 1a and 1arsquo on JurKatpLVTHM and Jurkat shBak cell viability after 24 h of exposure The results are expressed as

a percentage of live cells compared to control

S15

Figure S16 FRET DNA melting curves at 10 M of A) aHCl and 1a B) arsquoHCl and 1arsquo The concentration of F10T is 02 M

S16

Fig S17 Effect of treatment of PC-3 cells with 1a on the tumor growth Androgen-independent

prostate cancer cells were incubated in the absence or presence of 1a (25 microM) for 24 h The

cell suspension was mixed with Matrigelreg and injected subcutaneously into the right flank of

nude mice (5 x 106 cellsmouse) Ten mice were used in each group A representative tumor

sample from each group after 43 days treatment is shown P lt 005 P lt 0001 vs control

group

S17

• Z

S9

Figure S10 UV-vis spectrum of arsquomiddotHCl in water (500M)

Figure S11 UV-vis spectrum of 1arsquo in water (500M)

S10

Figure S12 Circular dichroism spectra of amiddotHCl and arsquomiddotHCl in water (500 M)

-1

-08

-06

-04

-02

0

02

04

06

08

220 240 260 280 300 320Δε

λ (nm)

amiddotHCl amiddotHCl

S11

Table S1 Crystallographic data for complex 1arsquo

Compound 1arsquomiddot2CHCl3

Empirical formula C29H40Cl8N2ORu

Formula weight 81730

Temperature (K) 200(2)

Wavelength (Aring) 071073

Crystal system Monoclinic

Space group P21

a (Aring) (ordm) 8865(2)

b (Aring) (ordm) 21916(2) 9049(1)

c (Aring) (ordm) 18183(2)

Volume (Aring3) 35327(8)

Z 4

Density calculated (g cmndash3) 1537

Absorption coefficient (mmndash1) 1075

F(000) 1664

Crystal size (mm3) 031 times 013 times 012

Theta range for data collection (ordm) 300 to 2524

Index ranges ndash10 to 10 ndash26 to 26ndash21 to 21

Reflections collected 66843

Independent reflections 12759 [R(int) = 0174]

Reflections [Igt2(I)] 8747

Completeness to theta 998

Goodness-of-fit on F2 1085

Final R indices [Igt2(I)] R1 = 0071 wR2 = 0114

R indices (all data) R1 = 0132 wR2 = 0134

Largest diff peak and hole (emiddotAringndash3) 0699 and ndash0588

S12

Figure S13 ORTEP drawing of the cation in the second independent molecule of the asymmetric unit

for compound 1arsquo with 50 probability ellipsoids Hydrogen bonded to carbon atoms have been

omitted for clarity

Table S2 Selected lengths (Aring) and angles (ordm) of the two independent molecules of the asymmetric

unit for 1arsquo

Ru(1)-Cl(1) 2404(4) Ru(1)-N(11) 2067(10)

Ru(1)-N(12) 2173(10) N(11)-O(11) 1381(13)

N(11)-C(21) 1284(15) N(12)-C(22) 1496(16)

C(21)-C(22) 1503(16) Ru(1)-Ct(1)a 1674

Ru(2)-Cl(3) 2396(4) Ru(2)-N(21) 2065(11)

Ru(2)-N(22) 2156(11) N(21)-O(21) 1388(13)

N(21)-C(61) 1263(16) N(22)-C(62) 1508(17)

C(61)-C(62) 1504(17) Ru(2)-Ct(2)b 1700

Cl(1)-Ru(1)-N(11) 829(3) Cl(1)-Ru(1)-N(12) 818(3)

N(11)-Ru(1)-N(12) 758(4) Ru(1)-N(11)-C(21) 1216(8)

N(11)-C(21)-C(22) 1157(11) C(21)-C(22)-N(12) 1087(10)

Ru(1)-N(12)-C(22) 1111(7) Ru(1)-N(12)-C(31) 1206(8)

C(22)-N(12)-C(31) 1127(10) Ru(1)-N(11)-O(11) 1240(8)

C(21)-N(11)-O(11) 1144(10) Cl(3)-Ru(2)-N(21) 820(3)

Cl(3)-Ru(2)-N(22) 820(3) N(21)-Ru(2)-N(22) 751(4)

Ru(2)-N(21)-C(61) 1225(9) N(21)-C(61)-C(62) 1156(12)

C(61)-C(62)-N(22) 1075(10) Ru(2)-N(22)-C(62) 1114(7)

Ru(2)-N(22)-C(71) 1204(9) C(62)-N(22)-C(71) 1113(10)

Ru(2)-N(21)-O(21) 1236(8) C(61)-N(21)-O(21) 1139(10)a Ct(1) centroid of the C(11)-C(16) ring b Ct(2) centroid of the C(51)-C(56) ring

S13

Fig S14 Morphological changes induced by compounds 1a and 1arsquo (25 microM) A549 cells were seeded and 24 h later compounds were added at the

indicated concentrations Cell morphology was evaluated by optical microscopy after 24 h

S14

Fig S15 Effect of enantiomers 1a and 1arsquo on JurKatpLVTHM and Jurkat shBak cell viability after 24 h of exposure The results are expressed as

a percentage of live cells compared to control

S15

Figure S16 FRET DNA melting curves at 10 M of A) aHCl and 1a B) arsquoHCl and 1arsquo The concentration of F10T is 02 M

S16

Fig S17 Effect of treatment of PC-3 cells with 1a on the tumor growth Androgen-independent

prostate cancer cells were incubated in the absence or presence of 1a (25 microM) for 24 h The

cell suspension was mixed with Matrigelreg and injected subcutaneously into the right flank of

nude mice (5 x 106 cellsmouse) Ten mice were used in each group A representative tumor

sample from each group after 43 days treatment is shown P lt 005 P lt 0001 vs control

group

S17

• Z

Figure S10 UV-vis spectrum of arsquomiddotHCl in water (500M)

Figure S11 UV-vis spectrum of 1arsquo in water (500M)

S10

Figure S12 Circular dichroism spectra of amiddotHCl and arsquomiddotHCl in water (500 M)

-1

-08

-06

-04

-02

0

02

04

06

08

220 240 260 280 300 320Δε

λ (nm)

amiddotHCl amiddotHCl

S11

Table S1 Crystallographic data for complex 1arsquo

Compound 1arsquomiddot2CHCl3

Empirical formula C29H40Cl8N2ORu

Formula weight 81730

Temperature (K) 200(2)

Wavelength (Aring) 071073

Crystal system Monoclinic

Space group P21

a (Aring) (ordm) 8865(2)

b (Aring) (ordm) 21916(2) 9049(1)

c (Aring) (ordm) 18183(2)

Volume (Aring3) 35327(8)

Z 4

Density calculated (g cmndash3) 1537

Absorption coefficient (mmndash1) 1075

F(000) 1664

Crystal size (mm3) 031 times 013 times 012

Theta range for data collection (ordm) 300 to 2524

Index ranges ndash10 to 10 ndash26 to 26ndash21 to 21

Reflections collected 66843

Independent reflections 12759 [R(int) = 0174]

Reflections [Igt2(I)] 8747

Completeness to theta 998

Goodness-of-fit on F2 1085

Final R indices [Igt2(I)] R1 = 0071 wR2 = 0114

R indices (all data) R1 = 0132 wR2 = 0134

Largest diff peak and hole (emiddotAringndash3) 0699 and ndash0588

S12

Figure S13 ORTEP drawing of the cation in the second independent molecule of the asymmetric unit

for compound 1arsquo with 50 probability ellipsoids Hydrogen bonded to carbon atoms have been

omitted for clarity

Table S2 Selected lengths (Aring) and angles (ordm) of the two independent molecules of the asymmetric

unit for 1arsquo

Ru(1)-Cl(1) 2404(4) Ru(1)-N(11) 2067(10)

Ru(1)-N(12) 2173(10) N(11)-O(11) 1381(13)

N(11)-C(21) 1284(15) N(12)-C(22) 1496(16)

C(21)-C(22) 1503(16) Ru(1)-Ct(1)a 1674

Ru(2)-Cl(3) 2396(4) Ru(2)-N(21) 2065(11)

Ru(2)-N(22) 2156(11) N(21)-O(21) 1388(13)

N(21)-C(61) 1263(16) N(22)-C(62) 1508(17)

C(61)-C(62) 1504(17) Ru(2)-Ct(2)b 1700

Cl(1)-Ru(1)-N(11) 829(3) Cl(1)-Ru(1)-N(12) 818(3)

N(11)-Ru(1)-N(12) 758(4) Ru(1)-N(11)-C(21) 1216(8)

N(11)-C(21)-C(22) 1157(11) C(21)-C(22)-N(12) 1087(10)

Ru(1)-N(12)-C(22) 1111(7) Ru(1)-N(12)-C(31) 1206(8)

C(22)-N(12)-C(31) 1127(10) Ru(1)-N(11)-O(11) 1240(8)

C(21)-N(11)-O(11) 1144(10) Cl(3)-Ru(2)-N(21) 820(3)

Cl(3)-Ru(2)-N(22) 820(3) N(21)-Ru(2)-N(22) 751(4)

Ru(2)-N(21)-C(61) 1225(9) N(21)-C(61)-C(62) 1156(12)

C(61)-C(62)-N(22) 1075(10) Ru(2)-N(22)-C(62) 1114(7)

Ru(2)-N(22)-C(71) 1204(9) C(62)-N(22)-C(71) 1113(10)

Ru(2)-N(21)-O(21) 1236(8) C(61)-N(21)-O(21) 1139(10)a Ct(1) centroid of the C(11)-C(16) ring b Ct(2) centroid of the C(51)-C(56) ring

S13

Fig S14 Morphological changes induced by compounds 1a and 1arsquo (25 microM) A549 cells were seeded and 24 h later compounds were added at the

indicated concentrations Cell morphology was evaluated by optical microscopy after 24 h

S14

Fig S15 Effect of enantiomers 1a and 1arsquo on JurKatpLVTHM and Jurkat shBak cell viability after 24 h of exposure The results are expressed as

a percentage of live cells compared to control

S15

Figure S16 FRET DNA melting curves at 10 M of A) aHCl and 1a B) arsquoHCl and 1arsquo The concentration of F10T is 02 M

S16

Fig S17 Effect of treatment of PC-3 cells with 1a on the tumor growth Androgen-independent

prostate cancer cells were incubated in the absence or presence of 1a (25 microM) for 24 h The

cell suspension was mixed with Matrigelreg and injected subcutaneously into the right flank of

nude mice (5 x 106 cellsmouse) Ten mice were used in each group A representative tumor

sample from each group after 43 days treatment is shown P lt 005 P lt 0001 vs control

group

S17

• Z

Figure S12 Circular dichroism spectra of amiddotHCl and arsquomiddotHCl in water (500 M)

-1

-08

-06

-04

-02

0

02

04

06

08

220 240 260 280 300 320Δε

λ (nm)

amiddotHCl amiddotHCl

S11

Table S1 Crystallographic data for complex 1arsquo

Compound 1arsquomiddot2CHCl3

Empirical formula C29H40Cl8N2ORu

Formula weight 81730

Temperature (K) 200(2)

Wavelength (Aring) 071073

Crystal system Monoclinic

Space group P21

a (Aring) (ordm) 8865(2)

b (Aring) (ordm) 21916(2) 9049(1)

c (Aring) (ordm) 18183(2)

Volume (Aring3) 35327(8)

Z 4

Density calculated (g cmndash3) 1537

Absorption coefficient (mmndash1) 1075

F(000) 1664

Crystal size (mm3) 031 times 013 times 012

Theta range for data collection (ordm) 300 to 2524

Index ranges ndash10 to 10 ndash26 to 26ndash21 to 21

Reflections collected 66843

Independent reflections 12759 [R(int) = 0174]

Reflections [Igt2(I)] 8747

Completeness to theta 998

Goodness-of-fit on F2 1085

Final R indices [Igt2(I)] R1 = 0071 wR2 = 0114

R indices (all data) R1 = 0132 wR2 = 0134

Largest diff peak and hole (emiddotAringndash3) 0699 and ndash0588

S12

Figure S13 ORTEP drawing of the cation in the second independent molecule of the asymmetric unit

for compound 1arsquo with 50 probability ellipsoids Hydrogen bonded to carbon atoms have been

omitted for clarity

Table S2 Selected lengths (Aring) and angles (ordm) of the two independent molecules of the asymmetric

unit for 1arsquo

Ru(1)-Cl(1) 2404(4) Ru(1)-N(11) 2067(10)

Ru(1)-N(12) 2173(10) N(11)-O(11) 1381(13)

N(11)-C(21) 1284(15) N(12)-C(22) 1496(16)

C(21)-C(22) 1503(16) Ru(1)-Ct(1)a 1674

Ru(2)-Cl(3) 2396(4) Ru(2)-N(21) 2065(11)

Ru(2)-N(22) 2156(11) N(21)-O(21) 1388(13)

N(21)-C(61) 1263(16) N(22)-C(62) 1508(17)

C(61)-C(62) 1504(17) Ru(2)-Ct(2)b 1700

Cl(1)-Ru(1)-N(11) 829(3) Cl(1)-Ru(1)-N(12) 818(3)

N(11)-Ru(1)-N(12) 758(4) Ru(1)-N(11)-C(21) 1216(8)

N(11)-C(21)-C(22) 1157(11) C(21)-C(22)-N(12) 1087(10)

Ru(1)-N(12)-C(22) 1111(7) Ru(1)-N(12)-C(31) 1206(8)

C(22)-N(12)-C(31) 1127(10) Ru(1)-N(11)-O(11) 1240(8)

C(21)-N(11)-O(11) 1144(10) Cl(3)-Ru(2)-N(21) 820(3)

Cl(3)-Ru(2)-N(22) 820(3) N(21)-Ru(2)-N(22) 751(4)

Ru(2)-N(21)-C(61) 1225(9) N(21)-C(61)-C(62) 1156(12)

C(61)-C(62)-N(22) 1075(10) Ru(2)-N(22)-C(62) 1114(7)

Ru(2)-N(22)-C(71) 1204(9) C(62)-N(22)-C(71) 1113(10)

Ru(2)-N(21)-O(21) 1236(8) C(61)-N(21)-O(21) 1139(10)a Ct(1) centroid of the C(11)-C(16) ring b Ct(2) centroid of the C(51)-C(56) ring

S13

Fig S14 Morphological changes induced by compounds 1a and 1arsquo (25 microM) A549 cells were seeded and 24 h later compounds were added at the

indicated concentrations Cell morphology was evaluated by optical microscopy after 24 h

S14

Fig S15 Effect of enantiomers 1a and 1arsquo on JurKatpLVTHM and Jurkat shBak cell viability after 24 h of exposure The results are expressed as

a percentage of live cells compared to control

S15

Figure S16 FRET DNA melting curves at 10 M of A) aHCl and 1a B) arsquoHCl and 1arsquo The concentration of F10T is 02 M

S16

Fig S17 Effect of treatment of PC-3 cells with 1a on the tumor growth Androgen-independent

prostate cancer cells were incubated in the absence or presence of 1a (25 microM) for 24 h The

cell suspension was mixed with Matrigelreg and injected subcutaneously into the right flank of

nude mice (5 x 106 cellsmouse) Ten mice were used in each group A representative tumor

sample from each group after 43 days treatment is shown P lt 005 P lt 0001 vs control

group

S17

• Z

Table S1 Crystallographic data for complex 1arsquo

Compound 1arsquomiddot2CHCl3

Empirical formula C29H40Cl8N2ORu

Formula weight 81730

Temperature (K) 200(2)

Wavelength (Aring) 071073

Crystal system Monoclinic

Space group P21

a (Aring) (ordm) 8865(2)

b (Aring) (ordm) 21916(2) 9049(1)

c (Aring) (ordm) 18183(2)

Volume (Aring3) 35327(8)

Z 4

Density calculated (g cmndash3) 1537

Absorption coefficient (mmndash1) 1075

F(000) 1664

Crystal size (mm3) 031 times 013 times 012

Theta range for data collection (ordm) 300 to 2524

Index ranges ndash10 to 10 ndash26 to 26ndash21 to 21

Reflections collected 66843

Independent reflections 12759 [R(int) = 0174]

Reflections [Igt2(I)] 8747

Completeness to theta 998

Goodness-of-fit on F2 1085

Final R indices [Igt2(I)] R1 = 0071 wR2 = 0114

R indices (all data) R1 = 0132 wR2 = 0134

Largest diff peak and hole (emiddotAringndash3) 0699 and ndash0588

S12

Figure S13 ORTEP drawing of the cation in the second independent molecule of the asymmetric unit

for compound 1arsquo with 50 probability ellipsoids Hydrogen bonded to carbon atoms have been

omitted for clarity

Table S2 Selected lengths (Aring) and angles (ordm) of the two independent molecules of the asymmetric

unit for 1arsquo

Ru(1)-Cl(1) 2404(4) Ru(1)-N(11) 2067(10)

Ru(1)-N(12) 2173(10) N(11)-O(11) 1381(13)

N(11)-C(21) 1284(15) N(12)-C(22) 1496(16)

C(21)-C(22) 1503(16) Ru(1)-Ct(1)a 1674

Ru(2)-Cl(3) 2396(4) Ru(2)-N(21) 2065(11)

Ru(2)-N(22) 2156(11) N(21)-O(21) 1388(13)

N(21)-C(61) 1263(16) N(22)-C(62) 1508(17)

C(61)-C(62) 1504(17) Ru(2)-Ct(2)b 1700

Cl(1)-Ru(1)-N(11) 829(3) Cl(1)-Ru(1)-N(12) 818(3)

N(11)-Ru(1)-N(12) 758(4) Ru(1)-N(11)-C(21) 1216(8)

N(11)-C(21)-C(22) 1157(11) C(21)-C(22)-N(12) 1087(10)

Ru(1)-N(12)-C(22) 1111(7) Ru(1)-N(12)-C(31) 1206(8)

C(22)-N(12)-C(31) 1127(10) Ru(1)-N(11)-O(11) 1240(8)

C(21)-N(11)-O(11) 1144(10) Cl(3)-Ru(2)-N(21) 820(3)

Cl(3)-Ru(2)-N(22) 820(3) N(21)-Ru(2)-N(22) 751(4)

Ru(2)-N(21)-C(61) 1225(9) N(21)-C(61)-C(62) 1156(12)

C(61)-C(62)-N(22) 1075(10) Ru(2)-N(22)-C(62) 1114(7)

Ru(2)-N(22)-C(71) 1204(9) C(62)-N(22)-C(71) 1113(10)

Ru(2)-N(21)-O(21) 1236(8) C(61)-N(21)-O(21) 1139(10)a Ct(1) centroid of the C(11)-C(16) ring b Ct(2) centroid of the C(51)-C(56) ring

S13

Fig S14 Morphological changes induced by compounds 1a and 1arsquo (25 microM) A549 cells were seeded and 24 h later compounds were added at the

indicated concentrations Cell morphology was evaluated by optical microscopy after 24 h

S14

Fig S15 Effect of enantiomers 1a and 1arsquo on JurKatpLVTHM and Jurkat shBak cell viability after 24 h of exposure The results are expressed as

a percentage of live cells compared to control

S15

Figure S16 FRET DNA melting curves at 10 M of A) aHCl and 1a B) arsquoHCl and 1arsquo The concentration of F10T is 02 M

S16

Fig S17 Effect of treatment of PC-3 cells with 1a on the tumor growth Androgen-independent

prostate cancer cells were incubated in the absence or presence of 1a (25 microM) for 24 h The

cell suspension was mixed with Matrigelreg and injected subcutaneously into the right flank of

nude mice (5 x 106 cellsmouse) Ten mice were used in each group A representative tumor

sample from each group after 43 days treatment is shown P lt 005 P lt 0001 vs control

group

S17

• Z

Figure S13 ORTEP drawing of the cation in the second independent molecule of the asymmetric unit

for compound 1arsquo with 50 probability ellipsoids Hydrogen bonded to carbon atoms have been

omitted for clarity

Table S2 Selected lengths (Aring) and angles (ordm) of the two independent molecules of the asymmetric

unit for 1arsquo

Ru(1)-Cl(1) 2404(4) Ru(1)-N(11) 2067(10)

Ru(1)-N(12) 2173(10) N(11)-O(11) 1381(13)

N(11)-C(21) 1284(15) N(12)-C(22) 1496(16)

C(21)-C(22) 1503(16) Ru(1)-Ct(1)a 1674

Ru(2)-Cl(3) 2396(4) Ru(2)-N(21) 2065(11)

Ru(2)-N(22) 2156(11) N(21)-O(21) 1388(13)

N(21)-C(61) 1263(16) N(22)-C(62) 1508(17)

C(61)-C(62) 1504(17) Ru(2)-Ct(2)b 1700

Cl(1)-Ru(1)-N(11) 829(3) Cl(1)-Ru(1)-N(12) 818(3)

N(11)-Ru(1)-N(12) 758(4) Ru(1)-N(11)-C(21) 1216(8)

N(11)-C(21)-C(22) 1157(11) C(21)-C(22)-N(12) 1087(10)

Ru(1)-N(12)-C(22) 1111(7) Ru(1)-N(12)-C(31) 1206(8)

C(22)-N(12)-C(31) 1127(10) Ru(1)-N(11)-O(11) 1240(8)

C(21)-N(11)-O(11) 1144(10) Cl(3)-Ru(2)-N(21) 820(3)

Cl(3)-Ru(2)-N(22) 820(3) N(21)-Ru(2)-N(22) 751(4)

Ru(2)-N(21)-C(61) 1225(9) N(21)-C(61)-C(62) 1156(12)

C(61)-C(62)-N(22) 1075(10) Ru(2)-N(22)-C(62) 1114(7)

Ru(2)-N(22)-C(71) 1204(9) C(62)-N(22)-C(71) 1113(10)

Ru(2)-N(21)-O(21) 1236(8) C(61)-N(21)-O(21) 1139(10)a Ct(1) centroid of the C(11)-C(16) ring b Ct(2) centroid of the C(51)-C(56) ring

S13

Fig S14 Morphological changes induced by compounds 1a and 1arsquo (25 microM) A549 cells were seeded and 24 h later compounds were added at the

indicated concentrations Cell morphology was evaluated by optical microscopy after 24 h

S14

Fig S15 Effect of enantiomers 1a and 1arsquo on JurKatpLVTHM and Jurkat shBak cell viability after 24 h of exposure The results are expressed as

a percentage of live cells compared to control

S15

Figure S16 FRET DNA melting curves at 10 M of A) aHCl and 1a B) arsquoHCl and 1arsquo The concentration of F10T is 02 M

S16

Fig S17 Effect of treatment of PC-3 cells with 1a on the tumor growth Androgen-independent

prostate cancer cells were incubated in the absence or presence of 1a (25 microM) for 24 h The

cell suspension was mixed with Matrigelreg and injected subcutaneously into the right flank of

nude mice (5 x 106 cellsmouse) Ten mice were used in each group A representative tumor

sample from each group after 43 days treatment is shown P lt 005 P lt 0001 vs control

group

S17

• Z

Fig S14 Morphological changes induced by compounds 1a and 1arsquo (25 microM) A549 cells were seeded and 24 h later compounds were added at the

indicated concentrations Cell morphology was evaluated by optical microscopy after 24 h

S14

Fig S15 Effect of enantiomers 1a and 1arsquo on JurKatpLVTHM and Jurkat shBak cell viability after 24 h of exposure The results are expressed as

a percentage of live cells compared to control

S15

Figure S16 FRET DNA melting curves at 10 M of A) aHCl and 1a B) arsquoHCl and 1arsquo The concentration of F10T is 02 M

S16

Fig S17 Effect of treatment of PC-3 cells with 1a on the tumor growth Androgen-independent

prostate cancer cells were incubated in the absence or presence of 1a (25 microM) for 24 h The

cell suspension was mixed with Matrigelreg and injected subcutaneously into the right flank of

nude mice (5 x 106 cellsmouse) Ten mice were used in each group A representative tumor

sample from each group after 43 days treatment is shown P lt 005 P lt 0001 vs control

group

S17

• Z

Fig S15 Effect of enantiomers 1a and 1arsquo on JurKatpLVTHM and Jurkat shBak cell viability after 24 h of exposure The results are expressed as

a percentage of live cells compared to control

S15

Figure S16 FRET DNA melting curves at 10 M of A) aHCl and 1a B) arsquoHCl and 1arsquo The concentration of F10T is 02 M

S16

Fig S17 Effect of treatment of PC-3 cells with 1a on the tumor growth Androgen-independent

prostate cancer cells were incubated in the absence or presence of 1a (25 microM) for 24 h The

cell suspension was mixed with Matrigelreg and injected subcutaneously into the right flank of

nude mice (5 x 106 cellsmouse) Ten mice were used in each group A representative tumor

sample from each group after 43 days treatment is shown P lt 005 P lt 0001 vs control

group

S17

• Z

Figure S16 FRET DNA melting curves at 10 M of A) aHCl and 1a B) arsquoHCl and 1arsquo The concentration of F10T is 02 M

S16

Fig S17 Effect of treatment of PC-3 cells with 1a on the tumor growth Androgen-independent

prostate cancer cells were incubated in the absence or presence of 1a (25 microM) for 24 h The

cell suspension was mixed with Matrigelreg and injected subcutaneously into the right flank of

nude mice (5 x 106 cellsmouse) Ten mice were used in each group A representative tumor

sample from each group after 43 days treatment is shown P lt 005 P lt 0001 vs control

group

S17

• Z

Fig S17 Effect of treatment of PC-3 cells with 1a on the tumor growth Androgen-independent

prostate cancer cells were incubated in the absence or presence of 1a (25 microM) for 24 h The

cell suspension was mixed with Matrigelreg and injected subcutaneously into the right flank of

nude mice (5 x 106 cellsmouse) Ten mice were used in each group A representative tumor

sample from each group after 43 days treatment is shown P lt 005 P lt 0001 vs control

group

S17

• Z