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Transformed Mammary Epithelial Cells (MDAMB-231) Nontransformed Mammary Epithelial Cells (NMuMG) Fibroblastic osteosarcoma cells (U2OS) Nontransformed Fibroblast (NIH3T3) Colon Cancer Line (HT29)*
GI50: 7.7nM LC50: 117nM GI50: 122nM LC50: 272nM GI50: 55nM LC50: 94nM GI50: 480nM LC50: >8M GI50: 12nM LC50: 22nM
Neuroblastoma (IMR-32)*
Ying, Y.; Taori, K.; Kim, H.; Hong, J.; Luesch, H. Journal of the American Chemical Society 2008, 130, 8455 *Taori, K.; Paul, V. J.; Luesch, H. Journal of the American Chemical Society 2008, 130, 1806
Thiol Analogues
HCT-116 growth HCT-116 HDAC inhibition (nM) cellular assay (nM) 4410 513 HeLa nuclear extracts HDACs (nM) 3711
385
20915
4229
332
5018
5227
Ying, Y.; Taori, K.; Kim, H.; Hong, J.; Luesch, H. Journal of the American Chemical Society 2008, 130, 8455
> 10000
> 10000
> 10000
Ying, Y.; Taori, K.; Kim, H.; Hong, J.; Luesch, H. Journal of the American Chemical Society 2008, 130, 8455
HCT-116 cell line growth inhibition experiment HDAC assays were performed with cell permeable fluorescent substrate Cytotoxicity for HT-116 cells not reported
Ying, Y.; Taori, K.; Kim, H.; Hong, J.; Luesch, H. Journal of the American Chemical Society 2008, 130, 8455
Importance of Thiol
Need thiol to chelate Zn2+ Analogue is active if hydrolysis to thiol is possible Structural similarity to FK228
Ying, Y.; Taori, K.; Kim, H.; Hong, J.; Luesch, H. Journal of the American Chemical Society 2008, 130, 8455
Bowers, A.; West, N.; Taunton, J.; Schreiber, S. L.; Bradner, J. E.; Williams, R. M. Journal of the American Chemical Society 2008, 130, 11219
Proliferation studies of various malignant human melanoma cell lines Cell viability assay based on ATP content
Bowers, A.; West, N.; Taunton, J.; Schreiber, S. L.; Bradner, J. E.; Williams, R. M. Journal of the American Chemical Society 2008, 130, 11219
Ying, Y.; Liu, Y.; Byeon, S. R.; Kim, H.; Luesch, H.; Hong, J. Organic Letters 2008, 10, 4021
Linker Length
n 1 2 3 4 HCT-116 growth inhibition (nM) >10000 6.80.6 62050 2500600 HeLa nuclear extracts HDACs (nM) >20000 3213 7600900 4100430
3900450
>20000
>600
>600
>600
>600
-Breast cancer cell line vs. normal mammary epithelial cells -Crystal Violet Dye
Nasveschuk, C. G.; Ungermannova, D.; Liu, X.; Phillips, A. J. Organic Letters 2008, 10, 3595
Linker Analogues
No free thiol can be formed. Synthetic Largazole showed weak selectivity for the cancer cell line GI50 = 49 vs 127nM Largazole Thiol showed weaker potency but better selectivity GI50 = 126 vs 1200nM
Seiser, T.; Kamena, F.; Cramer, N. Angewandte Chemie International Edition 2008, 47, 6483
60nM
38nM
50nM
70nM
-ED50 values from reporter gene assay in HEK293 cell promoted by cytomegalovirus (CMV) (nM) -Ability of being hydrolized to free thiol
Numajiri, Y.; Takahashi, T.; Takagi, M.; Shin-ya, K.; Doi, T. Synlett 2008, 2008, 2483
SEAP (Secreted Alkaline Phosphatase) reporter gene Cytomegalovirus promoter Induction of promoter by largazole analogues Measured Phosphatase units to calculate ED50 values
Numajiri, Y.; Takahashi, T.; Takagi, M.; Shin-ya, K.; Doi, T. Synlett 2008, 2008, 2483
Isostere Analogues
HDAC 1 Largazole Free Thiol 40 0.1 >3000 HDAC 2 HDAC 3 42 0.8 >3000 96 1 >3000 HDAC 6 >1000 40 >3000
0.9
1500
-Purified Human HDACs IC50 values in nM -Probably due to changed conformational space on nitrogen vs. oxygen
Bowers, A. A.; Greshock, T. J.; West, N.; Estiu, G.; Schreiber, S. L.; Wiest, O.; Williams, R. M.; Bradner, J. E. Journal of the American Chemical Society 2009, 131, 2900
>30
>30
>30
>30
Bowers, A. A.; West, N.; Newkirk, T. L.; Troutman-Youngman, A. E.; Schreiber, S. L.; Wiest, O.; Bradner, J. E.; Williams, R. M. Organic Letters 2009, 11, 1301
Bowers, A. A.; West, N.; Newkirk, T. L.; Troutman-Youngman, A. E.; Schreiber, S. L.; Wiest, O.; Bradner, J. E.; Williams, R. M. Organic Letters 2009, 11, 1301
23
0.67
0.27
HDAC1 (M)
Continued
29 14 >30
0.077
0.12
0.085
>30
0.00032
0.00086
0.0011
0.029
Disulfide Homodimer
0.00069
0.0017
0.0015
0.045
Bowers, A. A.; West, N.; Newkirk, T. L.; Troutman-Youngman, A. E.; Schreiber, S. L.; Wiest, O.; Bradner, J. E.; Williams, R. M. Organic Letters 2009, 11, 1301
Continued
Pyridine substitution of thiazol resulted in increased potency Methyl group in C-7 position is not important Linker analogues did not result in increased potency Enantiomer potency decreased by 3 orders of magnitude
C-7 demethylation
HDAC1 HDAC2 HDAC3 HDAC6 Largazole 0.0137 0.190 2.0 18.5 0.245 16.8 11.5 14.7 HCT-116 0.0250.0004 24.13.0
NA
NA
NA
NA
>100
Macrocycle dimer
NA
51.8
22.6
NA
26.83.7
-Purified Human HDACs. IC50 Values in M -C-7 demethylation decreases potency -C17 stereochemistry is crucial -Macrocycle dimer was surprisingly active in HDAC 2 and 3 but not 1
Chen, F.; Gao, A.-H.; Li, J.; Nan, F.-J. ChemMedChem 2009, 4, 1269
colorectal carcinoma cell line HCT-116 Largazole R = i-Pr, 18Z R = i-Bu, 18E R = i-Bu, 18Z R = Bn, 18E R = Bn, 18Z R = p-OH-Bn, 18E R = p-OH-Bn, 18Z 0.08 >10 0.56 >10 0.26 >10 0.39 >10
human lung cancer cell line A549 0.32 >10 3.28 >10 0.77 >10 1.46 >10
human embryonic kidney cell line HEK293 1.36 >10 8.95 >10 2.57 >10 >100 >10
human embryonic lung fibroblast cell line HLF 0.98 >10 6.12 >10 1.43 >10 >100 >10
Zeng, X.; Yin, B.; Hu, Z.; Liao, C.; Liu, J.; Li, S.; Li, Z.; Nicklaus, M. C.; Zhou, G.; Jiang, S. Organic Letters 2010, 12, 1368
C-7 Variations
IC50 HDAC1 (nM) Largazole X=H X = Et X = Bn 11.4 13.7 161.3 4.5 4.7
-Purified Recombinant HDACs -Size of substituent on C-7 position does not affect activity
Souto, J. A.; Vaz, E.; Lepore, I.; Poppler, A.-C.; Franci, G.; Alvarez, R.; Altucci, L.; de Lera, A. n. R. Journal of Medicinal Chemistry 2010, 53, 4654
Largazole 57229
Benelkebir, H.; Marie, S.; Hayden, A. L.; Lyle, J.; Loadman, P. M.; Crabb, S. J.; Packham, G.; Ganesan, A. Bioorganic & Medicinal Chemistry, In Press, Corrected Proof
3.150.35
0.170.05
Valine, glycine, and thiazoline groups could be replaced with slight decrease in activity Cell growth inhibition does not correlate necessarily with HDAC inhibition Half life of compounds is a concern No specification on what HDAC was used
Benelkebir, H.; Marie, S.; Hayden, A. L.; Lyle, J.; Loadman, P. M.; Crabb, S. J.; Packham, G.; Ganesan, A. Bioorganic & Medicinal Chemistry, In Press, Corrected Proof
Recent Analogues