Improved potency and reduced toxicity of the antifungal peptoid AEC5 through submonomer modification

Article abstract of DOI:10.1016/j.bmcl.2018.10.001

As proteolytically stable peptidomimetics, peptoids could serve as antifungal agents to supplement a therapeutic field wrought with toxicity issues. We report the improvement of an antifungal peptoid, AEC5, through an iterative structure-activity relationship study. A sarcosine scan was used to first identify the most pharmacophorically important peptoid building blocks of AEC5, followed by sequential optimization of each building block. The optimized antifungal peptoid from this study, β-5, has improved potency towards Cryptococcus neoformans and decreased toxicity towards mammalian cells. For example, the selectivity ratio for C. neoformans over mammalian fibroblasts was improved from 8 for AEC5 to 37 for β-5.

Full text of DOI:10.1016/j.bmcl.2018.10.001

Bioorganic&MedicinalChemistryLettersxxx(xxxx)xxx–xxx
Contents lists available at ScienceDirect
Bioorganic & Medicinal Chemistry Letters
journal homepage: www.elsevier.com/locate/bmcl
Improved potency and reduced toxicity of the antifungal peptoid AEC5
through submonomer modification
Madyson P. Middleton, Scott A. Armstrong, Kevin L. Bicker⁎
Middle Tennessee State University, Department of Chemistry, 1301 E. Main St., Murfreesboro, TN 37132, United States
A R T I C L E I N F O
A B S T R A C T
Keywords:
As proteolytically stable peptidomimetics, peptoids could serve as antifungal agents to supplement a therapeutic
field wrought with toxicity issues. We report the improvement of an antifungal peptoid, AEC5, through an
iterative structure-activity relationship study. A sarcosine scan was used to first identify the most pharmaco-
phorically important peptoid building blocks of AEC5, followed by sequential optimization of each building
block. The optimized antifungal peptoid from this study, β-5, has improved potency towards Cryptococcus
neoformans and decreased toxicity towards mammalian cells. For example, the selectivity ratio for C. neoformans
over mammalian fibroblasts was improved from 8 for AEC5 to 37 for β-5.
Peptoids
Cryptococcus neoformans
Fungal infections
Antifungals
Structure activity relationship
Cryptococcus neoformans is a common infectious agent among im-
munocompromised individuals, such as transplant patients and those
infected with HIV/AIDS. Initial infection of this yeast-like fungus occurs
in the pulmonary system with central nervous system infection fol-
lowing, leading to cryptococcal meningitis.1 The most recent estimates
indicate that roughly 220,000 HIV/AIDS patients contract cryptococcal
meningitis from C. neoformans each year and that over 181,000 (82%)
of these cases will be fatal.2 A second species, C. gattii, which is endemic
to parts of the South Pacific and has been observed as an outbreak in the
Pacific Northwest regions of Canada and the United States, will infect
both immunocompromised and immunocompetent individuals.3,4
Treatment for infections due to pathogenic fungi, such as
Cryptococcus spp. vary significantly with the species of the micro-
organism and the immune status of the patient. First-line treatments,
such as flucytosine, and fluconazole, as well as last-line treatments,
such as amphotericin B carry their own risks, including high mamma-
lian cytotoxicity that results in gastrointestinal complications, vo-
miting, QT prolongation, and hepatitis.5–7 Amplifying these side effects
is the fact that most current therapeutic regimen include combination
or long-term drug treatments due to increasingly observed resistance in
many fungal pathogens.6,8 The dearth of suitable treatments necessi-
tates new therapeutic options for patients dealing with fungal infec-
tions.
microorganisms over mammalian cells and lack of observed drug re-
sistance, likely stemming from their generally accepted mode of action.
Most AMPs used against fungi target the cell membrane, forming pores
or causing changes in cell permeability that result in leakage of cyto-
plasmic components, ultimately resulting in pathogen death.9 These
AMPs are able to take advantage of key differences between mamma-
lian and fungal cell membranes to selectively target the pathogen, such
as ergosterol, a principle sterol only present in fungal cell membranes.7
Regrettably, peptides are readily recognized and degraded by proteases,
thus serving as poor clinical therapeutics for combating in vivo fungal
infections.10,11
One solution to the proteolytic instability of AMPs are N-substituted
glycines, often referred to as peptoids. In peptides the side chain R
group is attached to the α-carbon, whereas in peptoids the R group is
attached to the amide nitrogen, making peptoids unrecognizable by
proteases. This modification maintains many of the advantageous
properties of peptides while significantly reducing the proteolytic sus-
ceptibility that results in short peptide half-lives in vivo.12 The anti-
microbial utility of peptoids has been demonstrated against both bac-
teria13–22 and fungi.23–25 By the use of a high-throughput screening
assay, we have recently identified a peptoid termed AEC5 with potency
against C. neoformans similar to that of first-line clinical antifungals.23
This compound demonstrated relatively minimal toxicity against sev-
eral mammalian cell types and excellent in vitro proteolytic stability.
Herein we report the improvement of AEC5 through sequential opti-
mization of each peptoid submonomer by an iterative structure activity
Many organisms utilize antimicrobial peptides (AMPs) as a part of
their innate immune response against pathogenic bacteria and fungi.9
AMPs are advantageous given their relatively high specificity for
Abbreviations: AMP, antimicrobial peptide; MIC, minimum inhibitory concentration
⁎
Corresponding author.
E-mail address: kevin.bicker@mtsu.edu (K.L. Bicker).
https://doi.org/10.1016/j.bmcl.2018.10.001
Received 26 June 2018; Received in revised form 23 September 2018; Accepted 1 October 2018
0960-894X/©2018ElsevierLtd.Allrightsreserved.
Pleasecitethisarticleas:Middleton,M.P.,Bioorganic&MedicinalChemistryLetters,https://doi.org/10.1016/j.bmcl.2018.10.001

M.P. Middleton et al.
Bioorganic&MedicinalChemistryLettersxxx(xxxx)xxx–xxx
Fig. 1. (A.) The structure of the antifungal tripeptoid AEC5, showing distinct functionalities at each of the three submonomer positions. (B.) Compound structures for
the sarcosine scan of AEC5, replacing the submonomer at each position with sarcosine. Calculated distribution coefficient (cLogD7.4), C. neoformans antifungal
potency (MIC), and HepG2 liver cell toxicity (TD50) for AEC5 sarcosine scan compounds. MW = molecular weight; cLogD7.4 = calculated distribution coefficient at
pH 7.4; MIC = minimum inhibitory concentration; TD50 = toxic dose 50%; SR = selectivity ratio (TD50/MIC); ND = not determined.
relationship (SAR) process.
coefficient, cLogD7.4, from −1.18 for AEC5 to 1.44 for AEC5sar2. This
increase in hydrophobicity is the likely cause of increased toxicity for
AEC5sar2 and stresses the importance of this submonomer in mitigating
mammalian toxicity. Interestingly, sarcosine substitution in place of an
aromatic heterocycle at position three results in a substantial 8-fold
change in antifungal potency with little effect on mammalian toxicity
compared to AEC5. The mechanism of action of AEC5 is still being
explored, and although we have hypothesized that this compound
works in a similar manner to other antifungal peptides and peptoids,
through membrane disruption, the importance of this aromatic het-
erocycle in compound potency suggests that AEC5 may kill C. neofor-
mans through a more complex mechanism of action. These data indicate
that the alkyl tail is the most important pharmacophoric moiety in
AEC5, and we subsequently sought to optimize this group during round
one of our SAR studies.
AEC5 Sarcosine Scan. We first sought to determine the pharmaco-
logical role of each of AEC5′s three submonomers on C. neoformans
potency and mammalian cell cytotoxicity. The structure of AEC5
comprises three unique submonomers, each with a different chemical
moiety (Fig. 1A). N-terminally is a long hydrophobic tail, followed by a
cationic ammonium side chain, and a C-terminal aromatic heterocycle.
A sarcosine scan of AEC5 was completed, similar to an alanine scan of
peptides, by synthesizing three AEC5 derivatives each with sarcosine in
place of one of the submonomers (Fig. 1B). All compounds were syn-
thesized on the solid phase using peptoid submonomer26 and tradi-
tional Fmoc techniques.27 Following synthesis, compounds were
cleaved from the resin using trifluoroacetic acid (TFA) and purified by
RP-HPLC. The minimum inhibitory concentration (MIC) against C.
neoformans and the cytotoxicity against HepG2 cells was then de-
termined for these compounds, termed AEC5sar1-3, as described pre-
viously (Fig. 1C).23 The MIC of known antifungal agents fluconazole
and amphotericin B was also determined to confirm the susceptibility of
this strain of C. neoformans (Fig. S3). Substituting sarcosine into any of
the three positions had deleterious effects on antifungal potency but
mixed effects of mammalian cytotoxicity. The largest change was ob-
served for AEC5sar1 where both potency and toxicity fell above the
highest concentrations tested, 400 μg/mL and 800 μg/mL, respectively.
This is not surprising given the previously reported importance of the
alkyl chain in similar compounds.13,22 Substituting sarcosine in place of
the cationic amino group of position two caused a modest 2-fold de-
crease in antifungal potency compared to AEC5, but concomitantly
increased the mammalian cell toxicity (TD50) by 2-fold, from 56.2 μg/
mL for AEC5 to 21.1 μg/mL for AEC5sar2. Sarcosine substitution at po-
sition two results in a substantial change in the calculated distribution
Round 1 SAR. Several studies have demonstrated the importance of
alkyl tail length on potency and toxicity of lipophilic antimicrobial
peptoids.13,22 AEC5 derivatives with sixteen, ten, and eight carbon alky
tails in position one, termed α-1, α-2, and α-3, respectively, were
synthesized using peptoid submonomer methods (Fig. 2A). Given the
polyene nature of the potent antifungal amphotericin B, we hypothe-
sized that the inclusion of polyene moieties in this position might im-
prove antifungal potency. The terpene aldehydes farnesal and citral
were therefore attached to an N-terminal glycine in position one via
reductive amination to afford compounds α-4 and α-5, respectively.
The C. neoformans MIC of these compounds was evaluated (Fig. 2B),
however, significant decreases in potency were observed for com-
pounds α-2 through α-5. This decrease in response to shortening the
alkyl tail (compounds α-2 and α-3) was not surprising given literature
precedent.13,22 Although increasing the length of the alkyl tail from
2

M.P. Middleton et al.
Bioorganic&MedicinalChemistryLettersxxx(xxxx)xxx–xxx
Fig. 2. (A.) Round 1 SAR compounds with varied hydrophobic tails in the first position. (B.) Calculated distribution coefficient (cLogD7.4), C. neoformans antifungal
potency (MIC), and HepG2 liver cell toxicity (TD50) for round 1 SAR compounds. MW = molecular weight; cLogD7.4 = calculated distribution coefficient at pH 7.4;
MIC = minimum inhibitory concentration; TD50 = toxic dose 50%; SR = selectivity ratio (TD50/MIC); ND = not determined.
thirteen to sixteen yielded a compound with a 2-fold improvement in C.
neoformans potency, it unsurprisingly resulted in significantly increased
toxicity against HepG2 liver cells. Neither polyene tail resulted in im-
proved potency, however, the farnesyl tail did show a large decrease in
toxicity (> 800 μg/mL) while possessing a similar cLogD7.4, albeit with
a 16-fold decrease in antifungal potency. We thought that the sig-
nificant decrease in toxicity was important and attempted to recover
antifungal potency through modification of the aromatic heterocycle in
position three during round two of SAR. This resulted in compounds γ-
8, γ-9, and γ-10, none of which showed improved potency over α-4 or
AEC5 (Fig. S4). The use of the farnesyl tail was ultimately abandoned,
and we concluded the optimal alkyl tail in position one was the original
tridecyl tail from AEC5.
heteroatom (γ-6) resulted in a 2-fold improvement in antifungal po-
tency and no significant change in mammalian cell toxicity. A 2-fold
improvement in antifungal potency was observed with three of the
round 2 derivatives; γ-2 with a thiophene side chain, γ-4 with an indole
side chain, and γ-6, as mentioned, with a phenyl side chain. Two of
these, γ-4 and γ-6 exhibited increased or unchanged mammalian cell
toxicity compared to AEC5. However, γ-2 showed a decrease in HepG2
toxicity with a TD50 of 79.3 μg/mL compared to 56.2 μg/mL for AEC5.
With improvement in both potency and toxicity, a selectivity ratio of 25
was calculated for γ-2, compared to 9 for AEC5. A higher selectivity
ratio, defined as the TD50 divided by the MIC, is indicative of a com-
pound that is more selective for pathogen over mammalian cells. Data
from round 2 of SAR indicated that replacing the furan in position three
with a thiophene greatly improved pathogen selectivity by increasing
antifungal potency and decreasing mammalian toxicity. Therefore, the
thiophene moiety was maintained in this position through round 3 of
SAR antifungal optimization.
Round 2 SAR. Following the alkyl tail in position one, the furan in
position three appeared to have the second largest effect on antifungal
potency from the sarcosine scan. AEC5 derivatives containing a variety
of aromatic heterocycles in this position were synthesized by peptoid
submonomer methods. These included imidazole derivatized com-
pounds γ-1 and γ-3, compound γ-2 containing a thiophene side chain, γ-
4 with an indole in this position, and γ-5 displaying a pyridyl ring
(Fig. 3A). It is important to note that γ-5 required altered synthetic
parameters published by Zuckerman et al. to negate the unwanted al-
kylation reaction at the pyridyl ring.28 This was accomplished using
chloroacetic acid in place of bromoacetic acid and extending the length
of amine coupling to account for the decreased leaving group ability of
chlorine over bromine. Derivatives containing an aromatic phenyl side
chain with no heteroatom (γ-6) and a non-aromatic heterocyclic tet-
rahydrofuran (γ-7) were also synthesized to investigate the role of these
individual properties. The C. neoformans potency and HepG2 toxicity of
these compounds was determined (Fig. 3B). Derivatives γ-1, γ-3, γ-5 all
showed 4- to 5-fold decreases in antifungal potency as well as a 2- to 4-
fold decrease in mammalian cell toxicity. It is interesting to note that
this decrease in potency and toxicity accompanies a decrease in
cLogD7.4, supporting our earlier findings that overall lipophilicity of
short antimicrobial peptoids is linked to mammalian toxicity.22 The
substitution with a non-aromatic heterocycle (γ-7) gave a modest 2-fold
decrease in antifungal potency and a 3-fold decrease in mammalian cell
toxicity. Interestingly, placement of an aromatic phenyl ring with no
Round 3 SAR. The final submonomer to optimize was the amino
cation in position two. Derivatives of γ-2 were synthesized containing
various chemical moieties in position two. These include compounds
with ammonium groups similar to γ-2 attached via fewer methylene
units (β-1 and β-2), or more methylene units (β-3), as well as an argi-
nine mimic (β-4), a trimethylammonium lysine mimic (β-5), and a non-
cationic hydroxyl group (β-6) (Fig. 4A). This last derivative was syn-
thesized and characterized to evaluate the efficacy of a hydrogen
bonding moiety without cationic nature in position two. The MIC of β-6
falls off 4-fold compared to γ-2 while the toxicity against HepG2 cells
becomes more than 2-fold worse (Fig. 4B). These data, combined with
data from AECsar2, highlight the role of the cation in position two in
improving C. neoformans potency, and perhaps more importantly, ne-
gating compound toxicity against mammalian cells. Bolt et al. recently
showed that the chain length between peptoid backbone and amino
cation affects mammalian cytotoxicity and antibacterial activity, al-
though the nature of this relationship varied between organisms.29
Modifying the side-chain length of γ-2 had little effect on antifungal
activity, with only β-2, possessing the two carbon linker, showing a 2-
fold decrease in potency. However, shortening the side-chain length
was detrimental to mammalian cytotoxicity with β-1 (3 carbon linker)
3

M.P. Middleton et al.
Bioorganic&MedicinalChemistryLettersxxx(xxxx)xxx–xxx
Fig. 3. (A.) Round 2 SAR compounds utilizing the tridecyl tail from Round 1 with varied heterocyclic side chains in the third position. (B.) Calculated distribution
coefficient (cLogD7.4), C. neoformans antifungal potency (MIC), and HepG2 liver cell toxicity (TD50) for round 2 SAR compounds. MW = molecular weight;
cLogD7.4 = calculated distribution coefficient at pH 7.4; MIC = minimum inhibitory concentration; TD50 = toxic dose 50%; SR = selectivity ratio (TD50/MIC).
and β-2 having HepG2 cytotoxicity of 47.3 and 40.5 μg/mL, respec-
tively. Similarly, lengthening the side-chain to six carbons (β-3) in-
creased toxicity moderately to 56.2 μg/mL.
distribution coefficient rises sharply, indicating that these compounds
are becoming more hydrophobic, even though they possess fewer
carbon/hydrogen groups. Given that the calculated distribution coef-
ficient factors in compound ionization, an explanation for this trend can
be obtained by observing the calculated pKa values for the amino ca-
tion. These values remain relatively constant around a pKa of 10 for
compounds possessing side-chain linker lengths of 3–6 carbons. How-
ever, as the linker length becomes shorter than three, the calculated pKa
begins to drop, meaning that on average this side-chain amino group
becomes less ionized at neutral pH. Noting our observations regarding
increased mammalian cytotoxicity with complete removal of the cation
in position two, we hypothesize that this loss of ionization with shorter
An interesting correlation between side-chain length and cLogD7.4
was observed in the synthesized peptoids which was extrapolated out
using distribution coefficients and pKa values calculated in ChemAxon’s
MarvinSketch (Fig. S5).30 β-3 had the highest distribution coefficient,
as expected given the increased number of methylene units. As this
side-chain is shortened to five, four (γ-2), or three methylenes (β-1), the
distribution coefficient decreases, as anticipated. However, as this
linker is shortened further to two methylenes (β-2) or further still to
hypothetical compounds containing one or zero methylene units, the
Fig. 4. Synthesis of γ-2 derivatives with unique side chain modifications.
4

M.P. Middleton et al.
Bioorganic&MedicinalChemistryLettersxxx(xxxx)xxx–xxx
Fig. 5. (A.) Round 3 SAR compounds utilizing the tridecyl tail from Round 1 and the thiophene aromatic heterocyle from Round 2 with varied cationic moieties in
position two. (B.) Calculated distribution coefficient (cLogD7.4), C. neoformans antifungal potency (MIC), and HepG2 liver cell toxicity (TD50) for round 3 SAR
compounds. MW = molecular weight; cLogD7.4 = calculated distribution coefficient at pH 7.4; MIC = minimum inhibitory concentration; TD50 = toxic dose 50%;
SR = selectivity ratio (TD50/MIC).
Table 1
Mammalian cell toxicity profiles of γ-2 and β-5 compared to AEC5.
Cell Type
HepG2
NIH/3T3
hRBC
hRBC
Compound
AEC5
γ-2
TD50 (SR)
TD50 (SR)
HC10 (SR)
Hmax (100 μg/mL)
56.2
79.3
91.2
18.0 (9)
48.6
65.9
114.6
3.3 (8)
56.6
37.2
42.6
2.4 (9)
30.4
48.6
46.0
3.4%
8.6%
9.4%
0.35 (25)
4.7 (29)
3.3 (21)
0.83 (12)
1.6 (14)
β-5
18.7 (37)
TD50 = toxic dose 50%; HC10 = hemolysis concentration 10%; Hmax = percent hemolysis at 100 μg/mL; SR = selectivity ratio (TD50 or HC10/MIC); all concentra-
tions in µg/mL.
side-chain length makes peptoids slightly less potent against C. neo-
formans and significantly more cytotoxic to mammalian cells. This is
further confirmed with the trimethylated γ-2 derivative, β-5, which is
permanently ionized. Synthesis of this compound was achieved using
standard submonomer methods to install a 4-monomethoxytrityl (Mmt)
protected diamine in position two, followed by Boc protection of the N-
terminus, orthogonal Mmt removal with 1% TFA, and methylation
using methyl iodide (Fig. 5). Cleavage from the resin and removal of the
Boc group using 95% TFA yielded only trimethylated β-5 with no mono
or dimethylated compound observed. β-5 exhibited no change in MIC
but showed further improvement to mammalian cytotoxicity with a
TD50 of 91.2 μg/mL.
methylation (Fig. 5). Previous literature has shown that guanidinium
containing peptoids have increased mammalian cytotoxicity, as well as
improved antibacterial properties,29 however, the effect of guanidinium
moieties on antifungal potency has not been explored. We observed that
substituting an arginine mimic in place of the lysine mimic in position
two of γ-2 resulted in increased HepG2 toxicity as expected, but un-
expectedly decreased antifungal potency 4-fold. Round 3 ultimately
identified β-5 as the most promising antifungal peptoid from this study.
Broad Toxicity Evaluation. One of the most significant shortcomings
of current antifungals is their broad toxicity. We therefore sought to
evaluate the broad toxicity of the most promising antifungal peptoids
developed through SAR against NIH/3T3 mouse fibroblasts and human
erythrocytes, in addition the HepG2 testing already completed
(Table 1). As seen with liver cells, both γ-2 and β-5 exhibited decreased
toxicity towards NIH/3T3 mouse fibroblasts compared to AEC5, with
selectivity ratios improving from 8 for AEC5 to 21 and 37 for γ-2 and β-
The last derivative explored in this round of SAR possessed the
cationic gaunidinium group of an arginine mimic (β-4). This compound
was synthesized in a similar manner to β-5, but with guanidinylation
using pyrazole carboxamidine after Mmt deprotection instead of
5

M.P. Middleton et al.
Bioorganic&MedicinalChemistryLettersxxx(xxxx)xxx–xxx
5, respectively. Interestingly, both γ-2 and β-5 had slightly higher levels
of hemolysis compared to AEC5. We note that the HC10 value reported
here for AEC5 differs from that published previously. Given that human
erythrocytes are primary and originate from different donors, we re-
evaluated AEC5 along with γ-2 and β-5 to provide the most accurate
hemolytic data. Maximum hemolysis at 100 μg/mL (Hmax) further
confirmed that the AEC5 derivatives tested here are more hemolytic
than the original peptoid. Although slightly increased hemolytic prop-
erties are a drawback for γ-2 and β-5, these compounds nonetheless
maintain improved selectivity ratios for human erythrocytes compared
to AEC5 due to their improved potency against C. neoformans.
small molecules and key peptoids, as well as supplemental figures, can
be found in the supporting information. Supplementary data to this
article can be found online at https://doi.org/10.1016/j.bmcl.2018.10.
001.
References
1. Movahed E, Munusamy K, Tan GMY, Looi CY, Tay ST, Wong WF. PLoS ONE.
2015;10(9):e0137457.
2. Rajasingham R, Smith RM, Park BJ, et al. Lancet Infect Dis. 2017;17(8):873–881.
3. Smith RM, Mba-Jonas A, Tourdjman M, et al. PLoS ONE. 2014;9(2):e88875.
4. Datta K, Bartlett KH, Baer R, et al. Emerg Infect Dis. 2009;15(8):1185.
5. Pappas PG, Kauffman CA, Andes D, et al. Clin Infect Dis. 2009;48(5):503–535.
6. Perfect JR, Dismukes WE, Dromer F, et al. Clin Infect Dis. 2010;50(3):291–322.
7. Walsh TJ, Anaissie EJ, Denning DW, et al. Clin Infect Dis. 2008;46(3):327–360.
8. Hanson KE, Alexander BD, Perfect J. Antimicrobial Drug Resistance: Clinical and
Epidemiological Aspects. Totowa, NJ: Humana Press; 2009:967–985.
9. Zasloff M. Nature. 2002;415(6870):389–395.
In conclusion, we report the iterative improvement of a previously
identified antifungal lead compound through sequential optimization of
each peptoid submonomer. This work identified two beneficial subtle
modifications to AEC5. The first being a thiophene aromatic hetero-
cycle in position three in place of a furan, which improved both potency
and selectivity through unknown mechanisms. The second being tri-
methylation of the amino side-chain in position two, locking this ni-
trogen into a cationic state, and thereby further ameliorating mam-
malian cell toxicity. Current efforts are focused on elucidating the
mechanism of action of these short antifungal peptoids and in-
vestigating their in vivo therapeutic properties.
10. Latham PW. Nat Biotech. 1999;17(8):755–757.
11. Zhang L, Falla TJ. Exp Opin Pharmacother. 2006;7(6):653–663.
12. Miller SM, Simon RJ, Ng S, Zuckermann RN, Kerr JM, Moos WH. Bioorg Med Chem
Lett. 1994;4(22):2657–2662.
13. Chongsiriwatana NP, Miller TM, Wetzler M, et al. Antimicrob Agents Chemother.
2011;55(1):417–420.
14. Chongsiriwatana NP, Patch JA, Czyzewski AM, et al. Proc Natl Acad Sci USA.
2008;105(8):2794–2799.
15. Chongsiriwatana NP, Wetzler M, Barron AE. Antimicrob Agents Chemother.
2011;55(11):5399–5402.
Author contributions
16. Kapoor R, Eimerman PR, Hardy JW, Cirillo JD, Contag CH, Barron AE. Antimicrob
Agents Chemother. 2011;55(6):3058–3062.
17. Kapoor R, Wadman MW, Dohm MT, Czyzewski AM, Spormann AM, Barron AE.
Antimicrob Agents Chemother. 2011;55(6):3054–3057.
All authors designed experiments and synthesized and characterized
compounds. MPM and KLB co-wrote the manuscript.
18. Patch JA, Barron AE. J Am Chem Soc. 2003;125(40):12092–12093.
19. Lee J, Kang D, Choi J, et al. Bioorg Med Chem Lett. 2017;28(2):170–173.
20. Mojsoska B, Zuckermann RN, Jenssen H. Antimicrob Agents Chemother.
2015;59(7):4112–4120.
Notes
21. Fisher KJ, Turkett JA, Corson AE, Bicker KL. ACS Comb Sci. 2016;18:6.
22. Turkett JA, Bicker KL. ACS Comb Sci. 2017;19:4.
The authors declare no competing financial interests.
23. Corson AE, Armstrong SA, Wright ME, McClelland EE, Bicker KL. ACS Med Chem Lett.
2016;7:12.
Acknowledgments
24. Galetti MD, Cirigliano AM, Cabrera GM, Ramírez JA. Mol Divers.
2012;16(1):113–119.
25. Luo Y, Bolt HL, Eggimann GA, et al. ChemBioChem. 2017;18(1):111–118.
26. Zuckermann RN, Kerr JM, Kent SBH, Moos WH. J Am Chem Soc.
1992;114(26):10646–10647.
We thank Dr. Erin McClelland in the Department of Biology at
MTSU for her continued assistance in working with C. neoformans. This
work was supported by funds provided by MTSU and by the National
Institutes of Health AI112861.
27. Amblard M, Fehrentz J-A, Martinez J, Subra G. Mol Biotechnol. 2006;33(3):239–254.
28. Burkoth TS, Fafarman AT, Charych DH, Connolly MD, Zuckermann RN. J Am Chem
Soc. 2003;125(29):8841–8845.
29. Bolt HL, Eggimann GA, Jahoda CAB, Zuckermann RN, Sharples GJ, Cobb SL.
Medchemcomm. 2017;8(5):886–896.
Appendix A. Supplementary data
30. ChemAxon (http://www.chemaxon.com) 2016.
Synthetic methods, characterization assay details, NMR analysis of
6