The Mode of SN38 Derivatives Interacting with Nicked DNA Mimics Biological Targeting of Topo I Poisons
Abstract
:1. Introduction
2. Results and Discussion
2.1. DOSY Experiments of Molecular Complexes
2.2. NOESY Experiment of the Molecular C Complex
2.3. Molecular Modeling
2.4. The Reaction Products Analysis
2.5. The Topo I Relaxation Activity Test
3. Materials and Methods
3.1. Chemical Substrates
3.2. Sample Preparation
3.3. HPLC Analysis
3.4. NMR Experiments
3.5. ESI–MS Experiments
3.6. MALDI–MS Experiments
3.7. MALDI Sample Preparation
3.8. Calculating Binding Constants from the Diffusion Coefficients
3.9. Molecular Dynamics Calculations
3.9.1. Calculating Binding Free Energies (Enthalpies) Using MM-PBSA and MM-GBSA Methods
3.9.2. PM7 Semi-Empirical Calculations
3.9.3. Cluster Analysis
3.10. The Topo I Relaxation Activity Assay
4. Conclusions
Supplementary Materials
Author Contributions
Funding
Data Availability Statement
Conflicts of Interest
References
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Sample | Concentration (mM) | 1 (T7-CH3) Integral | 2 (19-CH3) Integral | Di of 1 (10−10 m2 s−1) | Di of 2 (10−10 m2 s−−1) | Ka (mM−1) | % of Complex | |
---|---|---|---|---|---|---|---|---|
1 | 2 | |||||||
Complex of 1 + 2 after 5 days | 1.5 a | 3.25 b | 3 | 6.5 | 1.147 | 1.475 | 4.07 | 41% c 89% d |
Compound 2 | - | 2.0 | - | - | - | 3.07 | - | - |
Decamer 1 | 0.96 | - | - | - | 1.13 | - | - | - |
Compound 2 | Free; δ (ppm) | Complex 1 + 2; δ (ppm) | Δ (ppm) | Δ (Hz) |
---|---|---|---|---|
19-CH3 | 0.96 | 0.993 | 0.033 | 16.5 |
23-CH3 | 1.50 | 1.377 | −0.123 | −61.5 |
18-CH2 | 1.95 | 1.908 | −0.042 | −21 |
N–CH3 | 2.82 | 2.894 | 0.074 | 37 |
22-CH2 | 3.09 | 2.911 | −0.179 | −89.5 |
24-CH2 | 4.54 | 4.512 | −0.028 | −14 |
24-CH2 | 4.62 | 4.512 | −0.108 | −54 |
5-CH2 | 5.02 | 4.65 | −0.370 | −185 |
17-CH2 | 5.36 | 5.259 | −0.101 | −50.5 |
17-CH2 | 5.51 | 5.259 | −0.251 | −125.5 |
14-CH | 7.18 | 7.041 | −0.139 | −69.5 |
11-CH | 7.32 | 7.081 | −0.239 | −119.5 |
12-CH | 7.71 | 7.333 | −0.377 | −188.5 |
Base | H1’ | H2’ | H2” | H3’ | H4’ | H5’, H5” | H6/8 | H2/5/Me |
---|---|---|---|---|---|---|---|---|
T4 | −0.008 | −0.026 | −0.048 | 0.014 | −0.035 | -/−0.003 | −0.005 | −0.023 |
T5 | 0.020 | 0.052 | −0.046 | −0.006 | −0.009 | −0.001/- | 0.067 | −0.012 |
nick | ----------- | ---------- | ---------- | ----------- | ---------- | ------------------ | ----------- | ----------- |
G6 | −0.342 | −0.284 | −0.068 | 0.017 | −0.037 | −0.076/−0.222 | 0.020 | - |
T7 | −0.045 | −0.022 | −0.037 | −0.018 | −0.082 | -/0.013 | −0.204 | −0.025 |
A14 | −0.044 | −0.035 | −0.128 | −0.118 | −0.057 | -/- | −0.093 | −0.058 |
C15 | 0.170 | 0.010 | −0.199 | - | 0.021 | -/0.000 | 0.027 | −0.245 |
A16 | −0.230 | 0.024 | −0.093 | −0.043 | 0.029 | -/- | −0.224 | −0.204 |
A17 | −0.040 | −0.048 | −0.031 | −0.026 | −0.015 | -/0.073 | −0.069 | −0.081 |
Deriv. 2 | DNA 1 | NOE (a) | Structure 1 (b) | Structure 2 (b) | Structure 3 (b) | Structure 4 (b) | ||||||||||||
---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|
<6Å | <5Å | <4Å | <3Å | <6Å | <5Å | <4Å | <3Å | <6Å | <5Å | <4Å | <3Å | <6Å | <5Å | <4Å | <3Å | |||
11 | C15H2” | 8.0% | ++++ | ++++ | ++++ | ++ | ++++ | +++ | + | * | ||||||||
12 | A16H8 | 8.4% | ++++ | ++++ | +++ | +++ | + | |||||||||||
14 | A16H2 | 23.5% | ++++ | ++++ | ++++ | +++ | ++ | ++++ | ++++ | ++++ | + | |||||||
17a | G6H1′ | 11.0% | ++++ | ++++ | ++++ | +++ | + | |||||||||||
17b | ++++ | ++++ | +++ | . | ++++ | ++ | * | |||||||||||
17a | G6H4′ | 16.2% | ++++ | ++++ | ++++ | +++ | ++ | . | ||||||||||
17b | ++++ | ++++ | ++++ | + | ++++ | ++ | ++ | * | ||||||||||
17a | T5H1′ | 6.9% | ++++ | +++ | * | + | * | |||||||||||
17b | ++++ | ++++ | +++ | + | + | |||||||||||||
17a | T5H2” | 14.8% | ++++ | ++++ | +++ | +++ | ++ | + | * | |||||||||
17b | ++++ | ++++ | ++++ | +++ | ++++ | +++ | + | |||||||||||
17a | T5H2′ | 9.4% | ++++ | +++ | + | ++++ | +++ | ++ | * | |||||||||
17b | ++++ | ++++ | ++++ | ++ | ++++ | ++++ | + | |||||||||||
17a | T5H6 | 4.7% | ++++ | + | ++++ | ++++ | ++ | |||||||||||
17b | ++++ | ++++ | +++ | ++++ | ++++ | * | ||||||||||||
18 | G6H1’ | 12.3% | ++++ | ++++ | +++ | ++ | * | + | * | |||||||||
18 | G6H2” | 9.1% | +++ | ++ | * | |||||||||||||
18 | G6H4′ | 11.7% | ++++ | +++ | ++ | + | * | |||||||||||
18 | T7H4′ | 5.7% | ++++ | +++ | ++ | * | * | * | ||||||||||
18 | T7H5” | 4.7% | ++++ | +++ | + | * | ||||||||||||
18 | T7H5′ | 11.5% | ++++ | ++++ | +++ | ++ | * | * | ||||||||||
19 | A16H2 | 2.3% | +++ | * | ++++ | ++++ | ++ | * | ||||||||||
19 | G6H1′ | 5.4% | ++++ | +++ | ++ | * | * | |||||||||||
19 | G6H2” | 4.1% | +++ | ++ | * | |||||||||||||
19 | G6H4′ | 7.9% | ++++ | +++ | ++ | * | * | |||||||||||
19 | T7H4′ | 12.1% | ++++ | ++++ | ++++ | + | * | * | ||||||||||
19 | T7H5” | 5.7% | ++++ | +++ | ++ | * | * | |||||||||||
19 | T7H5′ | 10.9% | ++++ | ++++ | +++ | ++ | * | * | ||||||||||
24 | C15H5 | 5.7% | ++++ | ++++ | ++ | + | ||||||||||||
N–CH3 | C15H6 | 1.7% | ++ | * | * |
Atom Names | Atom Distances in Structure 1 in (Å) | |||
DNA | Comp. 2 | 100% Cluster a | 50% Cluster a | 25% Cluster a |
C15-N4 | -CH2NHCH3 | 7.23 ± 12.64 | 3.90 ± 0.33 | 3.62 ± 0.21 |
A16-N7 | -CH2NHCH3 | 7.53 ± 12.78 | 4.27 ± 0.32 | 4.01 ± 0.25 |
A16-N6 | -CH2NHCH3 | 8.47 ± 12.79 | 5.12 ± 0.49 | 4.75 ± 0.44 |
Atom Distances in Structure 3 in (Å) | ||||
G6-N2 | -CH2NHCH3 | 6.43 ± 10.88 | 4.03 ± 0.32 | 3.76 ± 0.21 |
A16-N3 | -CH2NHCH3 | 5.51 ± 10.65 | 3.37 ± 0.13 | 3.26 ± 0.09 |
IC50 (µM) | |||
CPT | SN38 | 2 | 3 |
92.9 ± 16.1 | 40.3 ± 13.17 | 100.7 ± 10.0 | 48.7 ± 8.2 |
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Bocian, W.; Naumczuk, B.; Urbanowicz, M.; Sitkowski, J.; Bierczyńska-Krzysik, A.; Bednarek, E.; Wiktorska, K.; Milczarek, M.; Kozerski, L. The Mode of SN38 Derivatives Interacting with Nicked DNA Mimics Biological Targeting of Topo I Poisons. Int. J. Mol. Sci. 2021, 22, 7471. https://0-doi-org.brum.beds.ac.uk/10.3390/ijms22147471
Bocian W, Naumczuk B, Urbanowicz M, Sitkowski J, Bierczyńska-Krzysik A, Bednarek E, Wiktorska K, Milczarek M, Kozerski L. The Mode of SN38 Derivatives Interacting with Nicked DNA Mimics Biological Targeting of Topo I Poisons. International Journal of Molecular Sciences. 2021; 22(14):7471. https://0-doi-org.brum.beds.ac.uk/10.3390/ijms22147471
Chicago/Turabian StyleBocian, Wojciech, Beata Naumczuk, Magdalena Urbanowicz, Jerzy Sitkowski, Anna Bierczyńska-Krzysik, Elżbieta Bednarek, Katarzyna Wiktorska, Małgorzata Milczarek, and Lech Kozerski. 2021. "The Mode of SN38 Derivatives Interacting with Nicked DNA Mimics Biological Targeting of Topo I Poisons" International Journal of Molecular Sciences 22, no. 14: 7471. https://0-doi-org.brum.beds.ac.uk/10.3390/ijms22147471