Communications
The second set comprised phenylpropanoyl derivatives
(compounds 12 and 13, Figure 3b) which on the one hand
probe the significance of the Michael system for bioactivity,
and on the other hand flexibility. These derivatives, formed by
insertion of an ethane linker consisting of sp3 carbons, are con-
formationally flexible between the subfragment structure and
the N-terminal aromatic ring. It has been known for a long
time that cinnamic acids tend to cyclo-dimerize when exposed
to UV radiation.[21] Thus it seemed worthwhile to make these
analogues without the Michael system to avoid the possibility
of dimerization, as this might be expected to cause a loss of
antimicrobial activity. In derivative 14, which bears a benzoyl
group, the linking region has been deleted (Figure 3c). Synthe-
sis, coupling, and deprotection have been carried out under
the same conditions as described for albicidin for the above
set of cinnamic acids.
Figure 4. Inhibition assay of, E. coli DNA gyrase supercoiling activity for the
novel albicidins prepared. The control experiment without enzyme and drug
(lane c) shows relaxed DNA (re) and the addition of enzyme (lane –) results
in supercoiled DNA (sc). The other lanes represent the reaction with enzyme
and derivatives. a) Derivatives 3–9, with albicidin (1) in the right lane. b) De-
rivatives 10–16 with albicidin (1) in the right lane. The assay was carried out
at the IC50 of albicidin (40 nm). Values obtained from two independent ex-
periments.
Finally, acetylation (15) as well as replacement of the aro-
matic ring with a cyclohexyl-substituent (16) were investigated
(Figure 3c). We aimed to decrease the molecular weight (MW)
of the target derivatives as much as possible. Since albicidin is
a natural product that is a member of one of the orally active
therapeutic classes which are antibiotics, antifungals, vitamins,
and cardiac glycosides, Lipinski’s “rule of 5” does not strictly
apply.[22] However, this provides a useful guideline to work
from, and when molecular weights become too large, this
lowers the “drug-likeness” of the compound. Comparing albici-
din and its derivatives with Lipinski’s “rule of 5” results in an
outnumbering of each component of the rule. Of course the
number of hydrogen bond donors (HBDs), hydrogen bond ac-
ceptors (HBAs), and the MW differ from each compound, but
the MW ranges from 724 (15) to 984 (9) Da and is in every
case clearly above 500 Da. The maximum number of HBDs,
which should be 5, as well as the HBAs (ideally 10) is clearly ex-
ceeded (8 to 9 HBDs, respectively 15 to 17 HBAs).
be important for the activity. We then investigated the role of
molecular weight, in particular the decrease of MW. Initially we
synthesized the analogue 10 without a methyl substituent on
the double bond, and it revealed less activity than albicidin. In-
terestingly compound 11 (no p-OH and no methyl on the
double bond) was similar in activity to albicidin, whilst having
a reduced molecular weight. But since it is only a loss of 4% of
weight, it may not affect the permeability and the solubility
properties. To decrease the molecular weight further, we cou-
pled benzoic acids and fatty acids to the subfragment 2. The
resulting benzoyl compound 14 showed a similar activity pro-
file. Encouragingly, it was the most soluble compound among
those prepared, based upon estimations from the retention
time in HR-LCMS (gradient details available in the Supporting
Information). The retention times on HPLC systems give
a good indication of the lipophilicity. But since the solubility
issue is a more complex one, we are measuring the solubility
in a separate experiment (unpublished data). The smallest cou-
pled residue was the acetyl group that led to derivative 15
which was only a weak inhibitor of the gyrase. Attaching the
cyclohexylcarboxylic acid (16) retained the activity of albicidin
but decreased the solubility of the compound.
DNA gyrase is the primary molecular target for albicidins.
The new compounds were therefore assessed for their inhibi-
tion of DNA gyrase from E. coli. The half maximal inhibitory
concentration (IC50) value of albicidin was determined to be
approximately 40 nm.[19] The assay was then carried out at this
concentration for all newly synthesized compounds and the re-
sults are shown in Figure 4.
Initially, the location of the OH-group on the cinnamoyl resi-
due was investigated. The naturally occurring albicidin has OH
in the para-position. Derivatives with m-phenol (3) and o-
phenol (4) and also lacking a hydroxy-group (5) were synthe-
sized. The data in Figure 4a indicate that these derivatives are
virtually devoid of inhibition of gyrase supercoiling at a concen-
tration of 40 nm. Therefore we investigated several alternative
substituents in the para-position. Initial studies for albicidin
reveal a moderate microsomal instability (unpublished data)
and to enhance metabolic stability, we replaced the hydroxy
group with a fluorine (6). Increasing lipophilicity can enhance
membrane permeability, and this can lead to a better accessi-
bility to the target. For this reason we also made the analogue
with trifluoromethyl in the para-position (9), and this showed
similar activity to albicidin. Along with their lipophilicity, the
electron withdrawing effects of compound 6 and 9 could also
We also investigated the importance of the Michael system,
as mentioned above, because it is known that cinnamic acids
tend to dimerize when exposed to UV light.[21] We synthesized
the saturated analogues 12 and 13 which contain phenylpro-
panoyl acids with a hydroxy group and a fluorine in the, para-
position respectively. Both compounds were less active than
their cinnamoyl counterparts but revealed a similar trend, that
fluorine in the para-position increases the potency.
Finally, the antimicrobial activities for all compounds were
determined against a set of Gram-positive and Gram-negative
bacteria (Table 1). When we compared the obtained data from
the gyrase assay with the results from the cell-based assay we
ChemMedChem 2016, 11, 1 – 6
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