Synthesis and in vivo PET Imaging of Hyaluronan Conjugates of Oligonucleotides

Transcription

1 Supporting information to: Synthesis and in vivo PET Imaging of Hyaluronan Conjugates of ligonucleotides Satish Jadhav, a Meeri Käkelä, b Jussi Mäkilä, b,c Max Kiugel, b Heidi Liljenbäck, b,d Jenni Virta, b Päivi Poijärvi-Virta, a Tiina Laitala-Leinonen, b Ville Kytö e, Sirpa Jalkanen f, Antti Saraste b,e,g, Anne Roivainen, b,d Harri Lönnberg a and Pasi Virta a* a Department of Chemistry, University of Turku, FI Turku, Finland b Turku PET Centre, University of Turku and Turku University Hospital, FI Turku, Finland c Department of Cell Biology and Anatomy, University of Turku, FI Turku, Finland d Turku Center for Disease Modeling, University of Turku, FI Turku, Finland e Heart Center, Turku University Hospital, University of Turku, FI Turku, Finland f MediCity Research Laboratory and Department of Medical Microbiology and Immunology, University of Turku, FI Turku, Finland g Institute of Clinical Medicine, University of Turku, FI Turku, Finland S1

2 Contents Figure S1. 1 H NMR (500 MHz, CDCl 3 ) spectrum of 3 Figure S2. 13 C NMR (100 MHz, CDCl 3 ) spectrum of 3 Figure S3. 1 H NMR (400 MHz, CDCl 3 ) spectrum of 4 Figure S4. 13 C NMR (100 MHz, CDCl 3 ) spectrum of 4 Figure S5. 1 H NMR (500 MHz, CDCl 3 ) spectrum of 5 Figure S6. 13 C NMR (125 MHz, CDCl 3 ) spectrum of 5 Figure S7. 1 H NMR (500 MHz, CDCl 3 ) spectrum of 6 Figure S8. 13 C NMR (125 MHz, CDCl 3 ) spectrum of 6 S3 S3 S4 S4 S5 S5 S6 S6 Figure S9. 1 H NMR (500 MHz, CDCl 3 ) spectrum of 7 7 Figure S C NMR (100 MHz, CDCl 3 ) spectrum of 7 7 Figure S11. 1 H NMR (500 MHz, CDCl 3 ) spectrum of 8 Figure S C NMR (125 MHz, CDCl 3 ) spectrum of 8 Figure S13. 1 H NMR (500 MHz, CDCl 3 ) spectrum of 10 Figure S C NMR (125 MHz, CDCl 3 ) spectrum of 10 Figure S15. 1 H NMR (400 MHz, CDCl 3 ) spectrum of 9 Figure S C NMR (100 MHz, CDCl 3 ) spectrum of 9 Figure S17. HSQC and 13 C NMR (100 MHz, CDCl 3 ) spectra of 9 Figure S18. HSQC and 13 C NMR (100 MHz, CDCl3) spectra of 9 Figure S19. RP HPLC chromatogram of 9 Figure S20. MS(ESI-TF) spectrum of 9 S8 S8 S9 S9 S10 S10 S11 S12 S13 S13 S2

3 (CH 3 ) 3 CSi C 2 Me Lev (CH 3 ) 2 Si H-3' arom CHPh H-4' H-2' H-1 H-1' H-3 H-6 H-5' H-4 H-6 H-5 H-2 Lev Lev Figure S1. 1 H NMR (500 MHz, CDCl 3 ) spectrum of PPM file: F:\Satish\HA 6 paper NMR\TBSdisaccharide \TBSHAdiasacchlev \2\fid expt: <ph_zgpg45> number of scans: 2000 LB: GB: Figure S2. 13 C NMR (100 MHz, CDCl 3 ) spectrum of 3. S3

4 SpinWorks 3: no title Me 2 C Bz Ph Bz PPM file:...r\tcadisaccharide \1\fid expt: <zg30> transmitter freq.: MHz width: Hz = ppm = Hz/pt number of scans: 16 freq. of 0 ppm: MHz LB: GF: Hz/cm: ppm/cm: Figure S3. 1 H NMR (400 MHz, CDCl 3 ) spectrum of 4. Me 2 C Bz Ph Bz arom C= Lev C 2Me C= Bz C= Bz C= Lev C= TCA C= TCA CHPh C-1 C-1' C-2 C-5' C-3' C-2' C-4' C-3 C-6C-5 Me Lev Lev Lev Figure S4. 13 C NMR (100 MHz, CDCl 3 ) spectrum of 4. S4

5 Figure S5. 1 H NMR (500 MHz, CDCl 3 ) spectrum of PPM file: F:\Satish\HA 6 paper NMR\Azidopropyldisaccharide \2\fid expt: <zgpg30> transmitter freq.: MHz width: Hz = ppm = Hz/pt number of scans: 1024 freq. of 0 ppm: MHz LB: GB: Figure S6. 13 C NMR (125 MHz, CDCl 3 ) spectrum of 5. S5

6 Figure S7. 1 H NMR (500 MHz, CDCl 3 ) spectrum of 6. Figure S8. 13 C NMR (125 MHz, CDCl 3 ) spectrum of 6. S6

7 Me2C Bz Ph Bz Me 2 C Bz Ph Bz N 3 Figure S9. 1 H NMR (500 MHz, CDCl 3 ) spectrum of Me2C Bz Ph Bz Me 2 C Bz Ph Bz N 3 PPM file: F:\Satish\HA 6 paper NMR\tetrasaccharide with Lev\HA tetrasachhalev \3\fid expt: <ph_zgpg45> number of scans: LB: GB: Figure S C NMR (100 MHz, CDCl 3 ) spectrum of 7. S7

8 Me Me arom arom H-2''' CHPh CHPh H-2' H-3''' H-4' H-1 H-6'' H-1' H-3' H-1'' H-1''' H-3'' H-3 H-4''' CH2 H-5' H-6 H-6' H-5''' H-4'' CH 2 H-4 H-2'' CH 2 H-5 H-2 H-5'' H H-6 * CH 2 * * = solvent traces and impurities * Figure S11. 1 H NMR (500 MHz, CDCl 3 ) of 8. C-3'' C-3 C-4' C-3''' C: TCA, Bz, C2Me arom CHPh CHPh C-1' C-1''' C-1'' C-1 C-4'' C-4 C-5''' C-5' C-3' C-2''' C-2' C-4''' C-6'' C-6 CH 2 C-5'' C-5 C-2'' C-2 C2Me C 2Me CH 2 CH2 Figure S C NMR (125 MHz, CDCl 3 ) spectrum of 8. S8

9 Figure S13. 1 H NMR (500 MHz, CDCl 3 ) spectrum of 10. Figure S C NMR (125 MHz, CDCl 3 ) spectrum of 10. S9

10 PPM file: F:\Satish\HA 6 paper NMR\HPLCPurifiedHA-6-I \HA \1\fid expt: <zg30> transmitter freq.: MHz width: Hz = ppm = Hz/pt number of scans: 160 freq. of 0 ppm: MHz LB: GB: Figure S15. 1 H NMR (400 MHz, CDCl 3 ) of PPM file: F:\Satish\HA 6 paper NMR\HPLCPurifiedHA-6-I \HA \2\fid expt: <ph_zgpg45> number of scans: LB: GB: Figure S C NMR (100 MHz, CDCl 3 ) spectrum of 9. S10

11 PPM file: D:\Satish\HA 6 paper NMR\HPLCPurifiedHA-6-I \HA \2\fid expt: <ph_zgpg45> LB: GB: number of scans: file: D:\Satish\HA 6 paper NMR\HPLCPurifiedHA-6-I \HA \2\fid expt: <ph_zgpg45> number of scans: LB: GB: PPM (F2) PPM (F1) file: D:\Satish\HA 6 paper NMR\HPLCPurifiedHA-6-I \4\ser expt: <hsqcetgpsi2> transmitter freq.: MHz time domain size: 1024 by 256 points width: Hz = ppm = Hz/pt number of scans: 5 F2: freq. of 0 ppm: MHz processed size: 1024 complex points window func tion: Sine Squared shift: 90.0 degrees F1: freq. of 0 ppm: MHz processed size: 1024 complex points window function: Sine Squared shift: 90.0 degrees PPM (F2) PPM (F1) file: D:\Satish\HA 6 paper NMR\HPLCPurifiedHA-6-I \4\ser expt: <hsqcetgpsi2> transmitter freq.: MHz time domain size: 1024 by 256 points width: Hz = ppm = Hz/pt number of scans: 5 F2: freq. of 0 ppm: MHz processed size: 1024 complex points window function: Sine Squared shift: 90.0 degrees F1: freq. of 0 ppm: MHz processed size: 1024 complex points window function: Sine Squared shift: 90.0 degrees PPM Figure S17. HSQC and of 13 C NMR (100 MHz, CDCl 3 ) spectra of 10. S11

12 PPM file: D:\Satish\HA 6 paper NMR\HPLCPurifiedHA-6-I \HA \2\fid expt: <ph_zgpg45> number of scans: LB: GB: PPM file: D:\Satish\HA 6 paper NMR\HPLCPurifiedHA-6-I \HA \2\fid expt: <ph_zgpg45> number of scans: LB: GB: PPM (F2) PPM (F1) file: D:\Satish\HA 6 paper NMR\HPLCPurifiedHA-6-I \4\ser expt: <hsqcetgpsi2> transmitter freq.: MHz time domain size: 1024 by 256 points width: Hz = ppm = Hz/pt number of scans: 5 F2: freq. of 0 ppm: MHz processed size: 1024 complex points window function: Sine Squared shift: 90.0 degrees F1: freq. of 0 ppm: MHz processed size: 1024 complex points window function: Sine Squared shift: 90.0 degrees Figure S18. HSQC and of 13 C NMR (100 MHz, CDCl 3 ) spectra of 9. S12

13 Figure S19. An RP HPLC chromatogram of purified 9. Conditions: An analytical RP HPLC column (C18, mm, 5 µm), a linear gradient from 0 to 100% acetonitrile in 0.01 mol L -1 aqueous Et 3 + Ac - (0-30 min), flow rate 1.0 ml min -1, detection at λ = 220 nm. Figure S20. MS(ESI-TF) spectrum (negative mode) of 9 (m/z calcd. for C 95 H 96 Cl 9 N 6 42 [M 2H] ). S13