Design and Synthesis of 4-(Heterocyclic Substituted Amino)-1H-Pyrazole-3-Carboxamide Derivatives and Their Potent Activity against Acute Myeloid Leukemia (AML)
Abstract
:1. Introduction
2. Results and Discussion
2.1. Chemistry
2.2. Structure-Activity Relationship Study
2.3. Molecular Modeling of Compound 8t with CDK2 and FLT3
2.4. Kinase Profiling
2.5. In vitro Cell Assays
2.6. Cellular Mode of Action
3. Materials and Methods
3.1. Procedure A For the Synthesis of Compounds 4a and 4b
3.2. Procedure B For the Synthesis of Compounds 4c–4h
3.3. Procedure C For the Synthesis of Compounds 2a–2c, 5a–5h, and 7a–7h
3.4. Procedure D for the Synthesis of Compounds 1a–1c and 6a–6h
3.5. Procedure E for the Synthesis of Compounds 3a–3c, 8a–8g, and 8r
3.6. Procedure F for the Synthesis of Compounds 3d, 8h–8q and 8s–8t
3.7. Kinase Inhibition Assay
3.8. Cell Growth Inhibition Assay
3.9. Cell Apoptosis Assay
3.10. Molecular Modeling
4. Conclusions
Supplementary Materials
Author Contributions
Funding
Acknowledgments
Conflicts of Interest
Abbreviations
FLT3 | Fms-like receptor tyrosine kinase 3 |
AML | acute myeloid leukemia |
ALL | acute lymphoblastic leukemia |
ITD | internal-tandem duplication |
TKD | tyrosine kinase domain |
CDK | cyclin-dependent kinases |
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FLT3 Inhibitors | Selectivity | Generation | Targets | Phases of Development (AML) | |
---|---|---|---|---|---|
Sunitinib (Type I) | Non-selective | First-generation | c-KIT, KDR, PDGFR, and FLT3 | Phase II | |
Lestaurtinib (Type I) | Non-selective | First-generation | Mutant and wild-type FLT3, JAK2, and FLT3 | Phase II | |
Midostaurin (Type I) | Non-selective | First-generation | FLT3, FLT3-ITD and FLT3-TKD | Launched | |
Sorafenib (Type II) | Non-selective | First-generation | RAF-1, VEGFR, PDGFR, c-KIT and FLT3 | Phase II | |
FN-1501 (Type I) | Non-selective | First-generation | FLT3, CDKs | Phase I 1 | |
Quizartinib (Type II) | Selective | Second-generation | PDGFR, c-KIT, FLT3, CSF-1R and RET | Launched (In Japan) | |
Gilteritinib (Type II) | Selective | Second-generation | FLT3 and AXL | Launched | |
Crenolanib (Type II) | Selective | Second-generation | FLT3 and PDGFR α/β | Phase III |
Cpd. | R 1 | IC50 (nM) 1 | ||
---|---|---|---|---|
CDK2 | CDK4 | FLT3 | ||
FN-1501 | 2.33 ± 0.02 | 1.02 ± 0.16 | 0.39 ± 0.07 | |
3a | 0.20 ± 0.01 | 34.13 ± 0.94 | 5.10 ± 0.46 | |
3b | 4.31 ± 0.91 | 54.24 ± 1.26 | 5.83 ± 0.74 | |
3c | 32.81 ± 1.34 | 87.07 ± 1.26 | 88.76 ± 1.06 | |
3d | 63.21 ± 0.91 | 77.37 ± 1.10 | 74.30 ± 1.21 |
Cpd. | Structure | IC50 (nM) 1 | IC50 (nM) 2 | ||||
---|---|---|---|---|---|---|---|
R 1 | R 2 | A | CDK2 | CDK4 | FLT3 | MV4-11 | |
FN-1501 | H | NH | 2.33 ± 0.02 | 1.02 ± 0.16 | 0.39 ± 0.07 | 9 ± 0.27 | |
8a | H | S | 10.39 ± 0.41 | 32.99 ± 0.94 | 17.81 ± 0.89 | 33.10 ± 0. 17 | |
8b | H | S | 8.42 ± 0.52 | 30.14 ± 0.99 | 19.18 ± 0.18 | 35.21 ± 0.83 | |
8c | H | S | 3.51 ± 0.19 | 2.41 ± 0.21 | 0.176 ± 0.09 | 4.28 ± 0.35 | |
8d | H | NH | 2.32 ± 0.014 | 5.32 ± 0.31 | 0.262 ± 0.01 | 9.5 ± 0.01 | |
8e | H | NH | 5.49 ± 0.42 | 51.035 ± 0.88 | 2.71 ± 0.31 | 38.3 ± 1.21 | |
8f | H | S | 31.7 ± 0.55 | 67.28 ± 1.09 | 8.07 ± 0.21 | 54.15 ± 1.73 | |
8g | H | NH | 3.74 ± 0.16 | 10.605 ± 0.24 | 1.945 ± 0.013 | 16.02 ± 0.43 | |
8h | H | NH | 0.282 ± 0.013 | 1.19 ± 0.09 | 0.038 ± 0.001 | 7.3 ± 0.33 | |
8i | H | S | 24.53 ± 0.57 | 9.165 ± 0.33 | 3.24 ± 0.14 | 27.21 ± 0.43 | |
8j | H | NH | 9.64 ± 0.46 | 17.79 ± 0.82 | 2.81 ± 0.11 | 21.35 ± 0.56 |
Cpd. | Structure | IC50 (nM) 1 | IC50 (nM) 2 | ||||
---|---|---|---|---|---|---|---|
R 1 | A | R 2 | CDK2 | CDK4 | FLT3 | MV4-11 | |
8k | N | 315.21 ± 2.30 | 22.3 ± 0.92 | 6.03 ± 0.16 | 51.09 ± 1.34 | ||
8l | N | 156.54 ± 3.22 | 23.11 ± 0.55 | 13.83 ± 0.37 | 133.50 ± 1.64 | ||
8m | N | 133.37 ± 1.74 | 17.53 ± 0.88 | 23.69 ± 0.65 | 143.50 ± 1.21 | ||
8n | N | 98.72 ± 1.33 | 4.85 ± 0.20 | 1.88 ± 0.09 | 19.92 ± 0.74 | ||
8o | N | 109.21 ± 1.01 | 1.81 ± 0.023 | 8.28 ± 0.12 | 45.44 ± 1.07 | ||
8p | N | 86.36 ± 1.36 | 3.81 ± 0.26 | 7.45 ± 0.19 | 27.04 ± 0. 31 | ||
8q | N | 508.94 ± 10.33 | 123.37 ± 1.09 | 20.23 ± 0.35 | 458.32 ± 9.20 | ||
8r | CH | 9.29 ± 0.64 | 24.95 ± 0.29 | 3.80 ± 0.10 | 3.28 ± 0.19 | ||
8s | CH | 5.43 ± 0.41 | 4.36 ± 0.19 | 0.82 ± 0.003 | 9.13 ± 0.11 | ||
8t | CH | 0.719 ± 0.064 | 0.770 ± 0.007 | 0.089 ± 0.013 | 1.22 ± 0.06 |
Kinase | IC50 (nM) |
---|---|
Compound 8t | |
CDK1/cyclin B | 138.03 ± 1.24 |
CDK2/cyclin A | 2.56 ± 0.31 |
CDK3/cyclin E | 6.88 ± 0.25 |
CDK4/cyclin D1 | 0.78 ± 0.04 |
CDK5/p35 | 9.64 ± 0.81 |
CDK6/cyclin D1 | 0.59 ± 0.09 |
CDK7/cyclin H | 21.31 ± 1.01 |
CDK9/cyclin K | 14.20 ± 0.82 |
ERK7/MAPK15 | 9.57 ± 0.23 |
FLT1/VEGFR1 | 12.41 ± 0.19 |
FLT3 | 0.035 ± 0.01 |
FLT3 (D835Y) | 0.75 ± 0.04 |
FLT3 (F594_R595insR) | 0.63 ± 0.09 |
FLT3 (F594_R595insREY) | 1.38 ± 0.11 |
FLT3 (ITD)-NPOS | 4.32 ± 0.23 |
FLT3 (ITD)-W51 | 0.94 ± 0.09 |
FLT3 (R595_E596insEY) | 1.59 ± 0.14 |
FLT3 (Y591-V592insVDFREYEYD) | 0.66 ± 0.08 |
FLT3 (ITD)-F691L | 0.60 ± 0.01 |
FLT4/VEGFR3 | 4.16 ± 0.19 |
GSK3β | 11.99 ± 1.20 |
KDR/VEGFR2 | 8.32 ± 0.54 |
Panel | Cell Line | GI50 (μM) | Panel | Cell Line | GI50 (μM) |
---|---|---|---|---|---|
Leukemia | CCRF-CEM | 0.22 | Melanoma | LOX IMVI | 0.20 |
HL-60(TB) | 1.15 | MALME-3M | 0.24 | ||
K-562 | 0.12 | M14 | 0.12 | ||
MOLT-4 | 0.08 | MDA-MB-435 | 0.11 | ||
RPMI-8226 | 0.89 | SK-MEL-2 | 1.16 | ||
SR | 0.06 | SK-MEL-5 | 0.25 | ||
Non-Small Cell Lung Cancer | A549/ATCC | 0.14 | SK-MEL-28 | 1.13 | |
EKVX | 0.06 | UACC-257 | 0.48 | ||
HOP-62 | 0.37 | UACC-62 | 1.09 | ||
HOP-92 | 0.04 | Ovarian Cancer | IGROV1 | 0.12 | |
NCI-H226 | 0.32 | OVCAR-3 | 0.23 | ||
NCI-H23 | 0.26 | OVCAR-4 | 0.22 | ||
NCI-H322M | 0.14 | OVCAR-5 | 0.27 | ||
NCI-H460 | 0.06 | OVCAR-8 | 0.36 | ||
NCI-H522 | 0.21 | NCI/ADR-RES | 1.48 | ||
Colon Cancer | COLO 205 | 0.12 | SK-OV-3 | 0.26 | |
HCC-2998 | 1.20 | Renal Cancer | 786-0 | 0.36 | |
HCT-116 | 0.11 | A498 | 0.14 | ||
HCT-15 | 0.22 | ACHN | 0.16 | ||
HT29 | 0.26 | CAKI-1 | 0.08 | ||
SW-620 | 0.12 | RXF 393 | 0.39 | ||
KM12 | 0.17 | SN12C | 0.45 | ||
CNS Cancer | SF-268 | 0.72 | TK-10 | 0.47 | |
SF-295 | 0.27 | UO-31 | 0.40 | ||
SF-539 | 0.39 | Breast Cancer | MCF7 | 0.16 | |
SNB-19 | 0.24 | MDA-MB-231 | 1.93 | ||
SNB-75 | 0.05 | HS 578T | 0.10 | ||
U251 | 0.11 | BT-549 | 1.02 | ||
Prostate Cancer | PC-3 | 0.18 | T-47D | 0.59 | |
DU-145 | 0.30 | MDA-MB-468 | 0.42 |
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Zhi, Y.; Wang, Z.; Yao, C.; Li, B.; Heng, H.; Cai, J.; Xiang, L.; Wang, Y.; Lu, T.; Lu, S. Design and Synthesis of 4-(Heterocyclic Substituted Amino)-1H-Pyrazole-3-Carboxamide Derivatives and Their Potent Activity against Acute Myeloid Leukemia (AML). Int. J. Mol. Sci. 2019, 20, 5739. https://doi.org/10.3390/ijms20225739
Zhi Y, Wang Z, Yao C, Li B, Heng H, Cai J, Xiang L, Wang Y, Lu T, Lu S. Design and Synthesis of 4-(Heterocyclic Substituted Amino)-1H-Pyrazole-3-Carboxamide Derivatives and Their Potent Activity against Acute Myeloid Leukemia (AML). International Journal of Molecular Sciences. 2019; 20(22):5739. https://doi.org/10.3390/ijms20225739
Chicago/Turabian StyleZhi, Yanle, Zhijie Wang, Chao Yao, Baoquan Li, Hao Heng, Jiongheng Cai, Li Xiang, Yue Wang, Tao Lu, and Shuai Lu. 2019. "Design and Synthesis of 4-(Heterocyclic Substituted Amino)-1H-Pyrazole-3-Carboxamide Derivatives and Their Potent Activity against Acute Myeloid Leukemia (AML)" International Journal of Molecular Sciences 20, no. 22: 5739. https://doi.org/10.3390/ijms20225739