S. Gapil Tiamas, et al.
Bioorganic&MedicinalChemistryLettersxxx(xxxx)xxxx
Scheme 4. Attempt of demthylation of car-
boxamide (+)-4h and cyclisation of 1,5-
diene of fislatifolic acid (+)- and (−)-1: a)
BBr3 1 M in CH2Cl2 (2 equiv.), CH2Cl2, -78
°C to r.t., 3 h; b) BF3.OEt2 (2 equiv.), CH2Cl2,
0 °C to r.t., 4 h; c) MeONHMe·HCl (1.1
equiv.), HATU (1.1 equiv.), N,N-diisopro-
pylethylamine (2 equiv.), DMF, r.t., 2 h. (+)
Enantiomers correspond to 1S,2S stereo-
chemistry and (−) enantiomers to 1R,2R.
Table 1
Biological evaluation of synthesized compounds on Bcl-xL/Bak, Mcl-1/Bid and
Bcl-2/Bim displacement assays.
Cpd.
(+)-1
5.7
0.7
0.6
4.0
0.5
0.6
0.1
> 23
> 23
(−)-1
9.6
8.6
Scheme 5. Synthesis of carboxamide derivatives (+)- and (−)-4k by peptidic
coupling on (+)- and (−)-fislatifolic acid 1: a) 4-Aminophenol (1.1 equiv.),
HATU (1.1 equiv.), N,N-diisopropylethylamine (2 equiv.), DMF, r.t., 2 h. (+)
Enantiomers correspond to 1S,2S stereochemistry and (−) enantiomers to
1R,2R.
(+)-2
> 23
> 23
> 23
> 23
> 23
> 23
19.0
0.9
0.3
0.1
(−)-2
> 33
16.6
> 33
7.2
> 23
2.4
(+)-4a
(−)-4a
(+)-4b
(−)-4b
(+)-4c
(−)-4c
(+)-4d
(−)-4d
(+)-4e
(−)-4e
(+)-4f
(−)-4f
(+)-4g
(−)-4g
(+)-4h
(−)-4h
(+)-4i
(−)-4i
(+)-4j
(−)-4j
(+)-4k
(−)-4k
(+)-9
1.7
0.1
0.7
0.2
0.3
0.1
0.1
2.7
0.6
0.6
0.4
0.4
2.1
9.2
2.3
1.0
7.3
0.6
17.4
0.6
8.2
0.7
> 23
> 23
20.1
> 33
> 33
5.4
> 23
> 23
> 23
> 23
3.1
proteins. Activity of the analogues with a longer chain was also studied.
While compound (+)-4f was found to be a Mcl-1/Bcl-2 dual inhibitor
with micromolar range affinities on both proteins, its enantiomer
(−)-4f expressed only a little affinity toward Mcl-1. Compounds 4g-i
and 4k were completely inactive against all three Bcl-2 family proteins.
In contrast, (+)-4j revealed to be Mcl-1 selective and its enantiomer
(−)-4j Bcl-2 selective. Finally, both enantiomers of the cyclized pro-
ducts 9 and 10 lost their activity in comparison with fislatifolic acid 1
and Weinreb amide 2.
1.9
0.3
0.3
> 23
> 23
> 23
> 23
> 23
> 23
> 23
> 23
> 23
> 23
> 23
> 23
> 23
> 23
> 23
> 23
> 23
8.3
> 33
5.5
2.2
15.0
> 33
> 33
> 33
> 33
> 33
> 33
8.5
1.0
> 23
> 23
> 23
> 23
> 23
> 23
> 23
> 23
2.2
0.4
0.3
In terms of cytotoxicity on A549 and MCF7 cancer cell lines, some of
the most potent compounds (+)-1, (+)-2, (+)-4a, (+)-4c and (+)-4f
according to in vitro protein assays revealed moderate cytotoxic activ-
ities (Table 2), in the same range as compound (+)-4k, not active on
Bcl-2 proteins. Thus, no direct correlation could be established between
protein affinity and cytotoxicity on cancer cell lines. This result in-
dicates that another mechanism could be involved in the cell death.
However, the Weinreb amide (+)-2 is still the most active analogue on
these two cancer cell lines with 31.2 and 42.1 μM on A549 and MCF7
respectively, thus approaching the result obtained with the cisplatin on
this latter cell line (43.2 μM).
> 33
> 33
> 33
9.0
0.3
0.1
> 23
> 23
> 23
> 23
> 23
> 23
1.5
(−)-9
> 33
18.9
15.0
5.2
(+)-10
2.4
3.8
1.2
1.2
Binding affinities were measured by fluorescence polarization14 after
a
competition between the ligand and a fluorescein-labeled peptide.
b
Ki is the concentration of the ligand corresponding to 50% of the binding of
the labelled reference compound, corrected for experimental conditions.15
In conclusion, a library of 26 novel carboxamides deriving from
synthetic fislatifolic acid (+)- and (−)-1 has been prepared based on
the results previously obtained. Their evaluation on the modulation of
Bcl-xL/Bak, Mcl-1/Bid, and Bcl-2/Bim interactions have shown the in-
cidence of tiny modifications on the carboxamide part and confirmed
than the (+) enantiomers were as or more active than the (–) en-
antiomers. In this series, six compounds behaved as dual Bcl-xL/Mcl-1
inhibitors in micromolar range but the previously prepared Weinreb
amide (+)-2 is still the most promising compound deriving from nat-
ural cyclohexenyl chalcones with sub-micromolar affinities toward Mcl-
1 and Bcl-2 but with moderate cytotoxicities on A549 and MCF7 cancer
cell lines.
c
Meiogynin A was used as reference compound.9h
Acknowledgments
This research was conducted within the frame of the International
French Malaysian Natural Product Laboratory (IFM-NatProLab) estab-
lished between CNRS-ICSN and University Malaya. We are grateful to
the Bright Spark Unit, University of Malaya, Malaysia and the French
Embassy in Malaysia for the scholarship of SGT. We also thank
Investissement d’Avenir grant of the Agence Nationale de la Recherche
(CEBA: ANR-10-LABX-25-01) for its financial support.
4