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Alkyl sulfonyl derivatized PAMAM-G2 dendrimers as nonviral gene delivery vectors with improved transfection
efficiencies Julia Morales-Sanfrutos, Alicia Megia-Fernandez, Fernando Hernandez-Mateo, Mª Dolores Giron-Gonzalez, Rafael Salto-Gonzalez and Francisco Santoyo-Gonzalez*
Departamento de Química Orgánica, Facultad de Ciencias, Instituto de Biotecnología, Universidad de Granada, 18071 Granada, Spain.
Departamento de Bioquímica y Biología Molecular II, Facultad de Farmacia, Universidad de Granada, 18071 Granada, Spain
E-mail: [email protected]
Table of Contents
1. 1H- and 13C-NMR spectra for compounds 4a-d, 5a-d, 6a-d, 8a-b, 8d, 9a-d, 10d, 11d and 13
2. Figure S1
3. Figure S2
4. Figure S3
5. Table S1
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1. 1H- and 13C-NMR spectra for compounds 4a-d, 5a-d, 6a-d, 8a-b, 8d, 9a-d, 10d, 11d and 13:
1H-NMR spectra for compound 4a
13C-NMR spectra for compound 4a
SOH
SOH
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1H-NMR spectra for compound 4b
13C-NMR spectra for compound 4b
SOH
11
SOH
11
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1H-NMR spectra for compound 4c
13C-NMR spectra for compound 4c
SOH
17
SOH
17
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1H-NMR spectra for compound 4d
13C-NMR spectra for compound 4d
SOH
SOH
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1H-NMR spectra for compound 5a
13C-NMR spectra for compound 5a
O2S
OH
O2S
OH
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1H-NMR spectra for compound 5b
13C-NMR spectra for compound 5b
O2S
OH11
O2S
OH11
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1H-NMR spectra for compound 5c
13C-NMR spectra for compound 5c
O2S
OH17
O2S
OH17
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1H-NMR spectra for compound 5d
13C-NMR spectra for compound 5d
SO2
OH
SO2
OH
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1H-NMR spectra for compound 6a
13C-NMR spectra for compound 6a
SO2
SO2
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1H-NMR spectra for compound 6b
13C-NMR spectra for compound 6b
O2S
11
O2S
11
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1H-NMR spectra for compound 6c
13C-NMR spectra for compound 6c
O2S
17
O2S
17
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1H-NMR spectra for compound 6d
13C-NMR spectra for compound 6d
SO2
SO2
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1H-NMR spectra for compound 8a
13C-NMR spectra for compound 8a
NH
OOH
NH
OOH
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1H-NMR spectra for compound 8b
13C-NMR spectra for compound 8b
NH
OOH
10
NH
OOH
10
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13C-NMR spectra for compound 8d
NH
OOH
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1H-NMR spectra for compound 9a
13C-NMR spectra for compound 9a
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1H-NMR spectra for compound 9b
13C-NMR spectra for compound 9b
O
NH
OSO210
O
NH
OSO210
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1H-NMR spectra for compound 9c
13C-NMR spectra for compound 9c
O
NH
OSO216
O
NH
OSO216
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1H-NMR spectra for compound 9d
13C-NMR spectra for compound 9d
NH
OO
SO2
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1H-NMR spectra for compound 10d
13C-NMR spectra for compound 10d
NH
OS
2
NH
OS
2
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1H-NMR spectra for compound 11d
13C-NMR spectra for compound 11d
NH
OS
SO2
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1H-NMR spectra for compound 13
13C-NMR spectra for compound 13
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9d
9d-G2(III)
2. Figure S1 1H NMR spectra (400 MHz) proving functionalization of PAMAM-G2 on the reaction with vinyl sulfone derivatives in selected cases: A, Disappearance of the vinyl sulfone proton signals of compound 9d in the reaction with PAMAM-G2 for the preparation of 9d-G2(III) dendrimer; B, Appearance of signals corresponding to the alkyl chains and the vinyl residue in the preparation of 6c-G2(I), 6d-G2(I), 9d-G2(III), 11d-G2(III), 6cd-G2(I)-Rho and 9d-G2(III)-Rho.
A
B
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3. Figure S2. Gel Electrophoresis Shift Assays. A.- Gel shift assays showing PAMAM-G2 or PAMAM-G2 derivatives-pEGFP-N3 binding at N/P ratios between 0 (pEGFP-N3 alone) and 10. The absence of plasmid band in the wells correlates with the inhibition of the plasmid DNA electrophoretic mobility. B.- Rhodamine labeled PAMAM or PAMAM-G2 derivatives gel shift assays.
Supplementary Material (ESI) for Organic & Biomolecular ChemistryThis journal is (c) The Royal Society of Chemistry 2010
4. Figure S3. DNase I protection experiments. Representative agarose electrophoresis of samples corresponding to pEGFP-N3 incubated in the absence (-) or presence (+) of DNase I as controls. pEGFP-N samples that have been complexed with the PAMAM-G2 derivatives before the DNase I treatment have been run in parallel. B.- Samples of PAMAM-G2 derivatives labeled with Rhodamine has been processed as above and the DNase I protection has been assayed by gel electrophoresis. (a) Dendrimer-pDNA complexes, (b) labeled dendrimer alone. Since the dendrimers are labeled with fluorescence, the gels have been photographed before (UV) and after ethidium bromide staining (EtBr). Notice that due to the interaction of the free labeled dendrimer with the SDS used in the sample preparation, the free labeled dendrimers have a net negative charge in the electrophoresis.
Supplementary Material (ESI) for Organic & Biomolecular ChemistryThis journal is (c) The Royal Society of Chemistry 2010
5. Table S1. Hydrodynamic Diameter and ζ Potential for PAMAM-G2 derivatives and their complexes with DNA.
Z Potential (mV)
Hydrodynamic Diameter (nm) Compound
Without DNA With DNA Without DNA With DNA
N/P
6b-G2(I) + 25.8 ± 0.9 + 14.4 ± 0.3 367 ± 20 734 ± 33 2.5 6c-G2(I) + 29.6 ± 0.7 + 14.8 ± 0.2 536 ± 84 524 ± 19 2.5 6d-G2-(I) + 29.9 ± 0.2 + 12.2 ± 0.2 394 ± 41 736 ± 68 2.5 9b-G2-(I) + 29.5 ± 0.5 + 21.8 ± 0.8 709 ± 95 515 ± 10 10 9c-G2(I) + 32.1 ± 0.6 + 13.2 ± 1.6 627 ± 27 2.5 9d-G2-(III) + 38.1 ± 0.6 + 28.2 ± 0.6 131 ± 26 226 ± 3 10 11d-G2-(III) + 27.3 ± 0.5 + 17.8 ± 0.7 272 ± 24 288 ± 5 10 13-G2(III) + 23.5 ± 0.4 + 21.3 ± 0.6 528 ± 92 329 ± 15 10 G2-Rho + 27.3 ± 3.2 + 15.2 ± 0.7 267 ± 66 326 ± 19 2.5 6c-G2(I)-Rho + 33.1 ± 1.2 + 24.3 ± 1.6 393 ± 18 207 ± 10 2.5 9d-G2-(III)-Rho + 44.9 ± 7.6 + 28.2 ± 2.4 282 ± 22 273 ± 9 10
Supplementary Material (ESI) for Organic & Biomolecular ChemistryThis journal is (c) The Royal Society of Chemistry 2010
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