Antiferroptotic Activity of Phenothiazine Analogues: A Novel Therapeutic Strategy for Oxidative Stress Related Disease

Article abstract of DOI:10.1021/acsmedchemlett.0c00293

Ferroptosis is an iron-catalyzed, nonapoptotic form of regulated necrosis that has been implicated in the pathological cell death associated with various disorders including neurodegenerative diseases (e.g., Friedreich's ataxia (FRDA), Alzheimer's disease, and Parkinson's disease), stroke, and traumatic brain injury. Recently, we showed that lipophilic methylene blue (MB) and methylene violet (MV) analogues both promoted increased frataxin levels and mitochondrial biogenesis, in addition to their antioxidant activity in cultured FRDA cells. Presently, we report the synthesis of series of lipophilic phenothiazine analogues that potently inhibit ferroptosis. The most promising compounds (1b-5b) exhibited an improved protection compared to the parent phenothiazine against erastin- and RSL3-induced ferroptotic cell death. These analogues have equivalent or better potency than ferrostatin-1 (Fer-1) and liproxstatin-1 (Lip-1), that are among the most potent inhibitors of this regulated cell death described so far. They represent novel lead compounds with therapeutic potential in relevant ferroptosis-driven disease models such as FRDA.

Full text of DOI:10.1021/acsmedchemlett.0c00293

pubs.acs.org/acsmedchemlett
Letter
Antiferroptotic Activity of Phenothiazine Analogues: A Novel
Therapeutic Strategy for Oxidative Stress Related Disease
Jun Liu, Indrajit Bandyopadhyay, Lei Zheng, Omar M. Khdour,* and Sidney M. Hecht*
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ABSTRACT: Ferroptosis is an iron-catalyzed, nonapoptotic form of
regulated necrosis that has been implicated in the pathological cell
death associated with various disorders including neurodegenerative
diseases (e.g., Friedreich’s ataxia (FRDA), Alzheimer’s disease, and
Parkinson’s disease), stroke, and traumatic brain injury. Recently, we
showed that lipophilic methylene blue (MB) and methylene violet
(MV) analogues both promoted increased frataxin levels and
mitochondrial biogenesis, in addition to their antioxidant activity in
cultured FRDA cells. Presently, we report the synthesis of series of
lipophilic phenothiazine analogues that potently inhibit ferroptosis.
The most promising compounds (1b−5b) exhibited an improved protection compared to the parent phenothiazine against erastin-
and RSL3-induced ferroptotic cell death. These analogues have equivalent or better potency than ferrostatin-1 (Fer-1) and
liproxstatin-1 (Lip-1), that are among the most potent inhibitors of this regulated cell death described so far. They represent novel
lead compounds with therapeutic potential in relevant ferroptosis-driven disease models such as FRDA.
KEYWORDS: Ferroptosis, Friedreich’s ataxia, lipid peroxidation, antiferroptotic activity, methylene blue, methylene violet
riedreich’s ataxia (FRDA) is a hereditary neurodegener-
ative disease caused by an intronic GAA repeat in the FXN
RAS and ST-expressing cells) inhibits the cystine/glutamate
transporter system xc−, leading to cysteine starvation, GSH
depletion, and ferroptosis.13,24 Additionally, a ferroptosis-
glutathione-independent pathway was recently identified for
suppressing ferroptosis.25,26 Ferroptosis-suppressor-protein 1
(FSP1), was first shown to counteract ferroptosis in the
absence of GPX4.25,26 FSP1 is an oxidoreductase, reducing
ubiquinone (coenzyme Q10) to radical scavenger ubiquinol,
thereby limiting lipid peroxidation within membranes,
independent of GPX4 or glutathione (Figure 1).26
F
gene that reduces frataxin (FXN) expression.1,2 Insufficient
frataxin impedes iron-containing mitochondrial complex
assembly, and the excess iron overloads the mitochondrial
matrix,3−7 triggering the formation of reactive oxygen species
(ROS) that contribute to cellular oxidative stress and lipid
peroxidation.8−12 These cellular and molecular features of
FRDA are also hallmark features of ferroptosis, a cell death
pathway.
Ferroptosis, originally described as a new form of cell
death,13 is an iron-dependent and ROS-dependent form of cell
death accompanied by rapid loss of plasma and mitochondrial
membrane integrity and prominent changes in mitochondrial
morphology.14−17 These abnormalities result from intense
membrane lipid peroxidation and oxidative stress, although the
precise mechanism(s) of ferroptotic cell death remains
undefined. Cells have evolved systems to block ferroptotic
cell death. One central regulators is glutathione peroxidase 4
(GPX4), that together with glutathione (GSH) and other
antioxidant defenses prevents lipid peroxidation and ferropto-
sis (Figure 1).17−20 Pathways linked to ferroptosis inhibition all
converge into GPX4/glutathione (Figure 1).17−20 RAS-
selective lethal 3 (RSL3), a ferroptosis inducer, acts directly
on GPX4 and inhibits its activity, leading to ferroptosis.13,21
GPX4 requires glutathione to reduce lipid hydroperoxides,22
while GSH synthesis is dependent on cysteine, produced from
cystine that itself is imported via the cystine/glutamate
antiporter system xc− (Figure 1).23 Erastin (eradicator of
Ferroptosis is believed to be involved in a number of
mitochondrial and neurodegenerative diseases.10,27 Evidence
from transgenic FRDA mice-derived neurons, for example,
describes a functional imbalance between respiratory chain
complexes I and II, driving free radical formation, thereby
resulting in glutathione depletion and lipid peroxidation that
contribute to neuronal death.11,12 Recently, reduction of lipid
hydroperoxides in fibroblasts and neurons from different
FRDA mouse models has also improved FRDA phenotypes
and diminished ferroptotic cell death.10−12 This suggests that
targeting ferroptosis pharmacologically, in addition to increas-
Received: May 29, 2020
Accepted: September 15, 2020
Published: September 15, 2020
https://dx.doi.org/10.1021/acsmedchemlett.0c00293
© XXXX American Chemical Society
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A

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Letter
Figure 1. Schematic representation depicting the antiferroptotic function of both GSH-dependent and GSH-independent (FSP1) pathways and the
key targeting points. Lipid peroxidation is a hallmark of ferroptosis that is triggered by inhibition of the glutamate-cystine antiporter system (system
xc−) with erastin or inhibition of GPX4 with RSL3.
ing both FXN and ATP levels, might constitute a beneficial
FRDA therapy.
of the newly prepared lipophilic phenothiazine analogues 1−5
was tested in selected biochemical and biological assays. Since
lipid peroxidation is a hallmark of ferroptotic cell death, we
established a lipid peroxidation assay in FRDA patient-derived
lymphocyte cells using the GPX4 inhibitor RSL3. After
establishing dose-dependent sensitivity to RSL3-mediated
ferroptosis, the ability of analogues 1−5 to quench lipid
peroxidation was studied in FRDA lymphocytes that had been
treated with GPX4 inhibitor RSL3. The fluorescent lipid
peroxidation-sensitive fatty acid-conjugated dye (C11-BODI-
PY581/591) probe was used as described.37 Initially, ferroptosis
sensitive FRDA cells were treated with varying concentrations
of analogues 1−5 or the validated ferroptosis inhibitors
ferrostatin-1 (Fer-1) and liproxstatin-1 (Lip-1) and then with
a lethal dose of RSL3 (2 μM). Table 1 shows the activity and
potency of tested MB/MV analogues in blocking RSL3-
induced lipid peroxidation in FRDA lymphocytes, with low
nanomolar EC50 values. As summarized in Table 1, both MB
and MV analogues were more effective in quenching lipid
peroxidation when compared to their parent compounds.
Plausibly this might be due to the influence of their side chains
on their subcellular localization or mitochondrial membrane
lipid rich environment. Interestingly, the redox core also
influenced their potency. The MV redox core exhibited better
potency than MB when tested at lower concentrations,
whereas radical-trapping antioxidants inhibit autoxidation by
donating an H atom to the peroxyl radical to give a
nonpropagating radical from both the phenolic O−H and
aminic N−H of the MV redox core. In this assay the optimal
side chain length for MB analogues was 10-carbon atoms,
while for MV analogues it was 8−12 carbon atoms (Table 1).
MV analogues 1b−5b have equivalent or better potency than
the most potent inhibitors of ferroptosis described so far,
notably ferrostatin-1 (Fer-1) and liproxstatin-1 (Lip-1), and
are superior to α-TOH (Table 1). Elevated lipid peroxyl
radical levels are associated with ferroptosis onset, and many of
Phenothiazine has a tricyclic aromatic ring containing sulfur
and nitrogen atoms and has gained importance for
pharmaceutical applications in part due to its redox proper-
ties.28 MB undergoes reversible two-electron oxidation
processes at low potential, enabling MB to donate electrons
to the electron transport chain in the absence of oxygen,
thereby enhancing ATP production and cytochrome c oxidase
activity and contributing to cell survival via bypassing
complexes I−III to generate ATP.29−32 Recently, we have
reported lipophilic phenothiazines that promote increased
frataxin levels and mitochondrial biogenesis in cells from
FRDA patients with reduced toxicity and better overall activity
than MB/MV.33,34
Improved ferroptosis inhibitors are thus of great interest. As
ferroptosis is accompanied by a burst in lipid peroxidation
within plasma and mitochondrial membranes,35 we hypothe-
sized that lipophilic MB/MV derivatives might be more
effective inhibitors of this cell death pathway. A series of
lipophilic phenothiazines were synthesized to probe their
activity. The prepared analogues were evaluated as in vitro
inhibitors of ferroptosis, and some exhibited strong anti-
ferroptosis activity, at least comparable to ferroptosis inhibitors
such as ferrostatin-1 (Fer-1) and liproxstatin-1 (Lip-1).
We have reported the importance of side chain length for
facilitating their effects on the mitochondrial respiratory chain
to achieve improved antioxidant activity.36 Accordingly, we
prepared analogues having different side chain lengths attached
to the phenothiazine redox core. The syntheses of lipophilic
MB analogues 1a−5a, having alkyl substituents on phenothia-
zine redox core position 2 (Figure 2), were carried out as
described (Scheme 1), employing a Wittig reaction as the key
step.33,34 The MV analogues (1b−5b) (Figure 2) were
obtained by basic hydrolysis of the corresponding MB
analogues 1a−5a (Scheme 1).33,34 The antiferroptotic activity
B
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Figure 2. Chemical structures of the synthesized methylene blue (MB) and methylene violet (MV) analogues (1−5).
the most potent antiferroptotic agents exhibit scavenging of
lipid peroxyl radicals. To study the relevance of this
mechanism to MB and its derivatives, we modified MB by
N-acetylation. MB acetylation blocked its redox activity and
completely eliminated its antiferroptotic effects (Table 1). To
complement the model of lethal RSL3-induced lipid
peroxidation by inhibition of GPX4, we employed a second
method of inducing ferroptosis that models the GSH depletion
and oxidative stress observed in mitochondrial disease patients.
Erastin is a classical inducer of ferroptosis; it acts by inhibition
of cystine-glutamate exchange at the plasma membrane and as
a consequence depletes the cell of GSH (Figure 1).13,24 We
investigated the effects of 1−5 on erastin-induced cell death in
primary FRDA patient-derived fibroblasts. Erastin-induced cell
death was determined by assessing the depletion of cellular
ATP using a commercially available luciferase-linked ATPase
enzymatic assay (ViaLight Plus proliferation/cytotoxicity Kit,
Lonza, Walkersville, MD). As summarized in Table 2, MB/MV
analogues potently protected FRDA cells with a trend similar
to that noted for RSL3-induced lipid peroxidation in FRDA
lymphocytes. Compounds 1b−4b were the most efficient,
having EC50 values of 15 2, 11 1, 10 1, and 15 1 nM,
respectively (Table 2). Again, in this assay the lipophilic
analogues were more effective than their redox core parents
(MB/MV). The optimal side chain length for MB analogues
was 10 carbon atoms (3a) and was 8−12 carbon atoms for MV
analogues. Compounds having a side chain with 16 carbon
atoms (5a and 5b) were less protective. These encouraging
findings demonstrated that the MV analogues had better
overall antiferroptotic activity than that of MB analogues as
well as the well-known ferroptosis inhibitors ferrostatin-1 and
liproxstatin-1.
Methylene blue is a recyclable redox core that can donate
electrons to cytochrome c in the mitochondrial electron
transport chain, contributing to increased ATP, as described
previously.29,30,33,38 Frataxin deficiency in FRDA leads to
decreased ATP production. Given the hormetic pharmaco-
logical effect of MB, exhibiting opposite effects at high and low
doses,39 we used MB in a concentration range (100 nM−2.5
μM) relevant in cellular studies, in particular to those reporting
neuroprotective effects of MB.29−34 Accordingly, MB/MV
analogues 1−5 were evaluated for their ability to support ATP
production in FRDA lymphocytes (Figure 3). Total cellular
ATP was measured in a cultured FRDA cell line as described
previously.33,34 The cells were grown on glucose-free media
supplemented with galactose. Since cells grown in galactose
C
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Scheme 1. Synthesis of Phenothiazine Analogues
Table 1. Antiferroptotic Effects of MB/MV Analogues (1−
Table 2. Effect of MB/MV Analogues (1−5) on the Cellular
5) in FRDA Lymphocytes against RSL3-Induced Lipid
Viability of Primary FRDA Fibroblasts against Erastin-
a
a
Peroxidation
Induced Ferroptotic Cell Death
Compound
MB
Acetylated MB
MV
1a
1b
2a
2b
3a
3b
4a
4b
5a
5b
Lip-1
Fer-1
α-TOH
RSL3 challenge lipid peroxidation inhibition EC50 (nM)
Compound
Erastin challenge cell survival EC50 (nM)
88 13
>2500
MB
MV
1a
1b
2a
2b
3a
3b
4a
4b
5a
5b
Lip-1
Fer-1
α-TOH
43
27
39
15
37
11
29
10
35
15
55
22
22
33
>500
3
3
3
2
3
1
3
1
1
1
6
2
4
4
b
26
76
14
53
12
38
9
3
6
1
3
1
3
1
54
16
70
25
22
48
5
1
5
3
2
2
a
>500
The calculated EC50 values were determined graphically from the
a
dose−response curves using the nonlinear regression curve fit analysis
model.
The calculated EC50 values were determined graphically from the
dose−response curves using the nonlinear regression curve fit analysis
model. N-acetyl leucomethylene blue (Figure 2).
b
rely mostly on oxidative phosphorylation (OXPHOS) to
produce their ATP, they become more sensitive to
mitochondrial respiratory chain inhibitors than cells grown in
normal (glucose) medium. As shown in Figure 3, MB strongly
diminished ATP levels in a concentration dependent fashion
(0.25, 0.5, and 2.5 μM) in Friedreich’s ataxia lymphocytes. MB
increased ATP levels slightly when used at 0.1 μM
concentration. In contrast, MV itself was less detrimental in
inhibiting ATP levels at higher concentrations. Similar results
to MB itself were observed for MB analogues, having a short to
medium length alkyl substituent chain (8−10 carbon atoms)
(1a−3a). Our earlier study34 suggested that compounds
having lipophilic side chains of 16-carbon atoms (5a) exhibit
lesser inhibition of mitochondrial respiratory chain complexes
and correspondingly permit more effective maintenance of
cellular ATP levels. Compound 5a clearly increased the ATP
level when employed at 0.1 and 0.25 μM concentrations,
although it suppressed ATP levels when used at 2.5 μM
concentration. On the other hand, MV analogues (1b−5b)
with similar substitution patterns of the lipophilic alkyl side
chain were also studied in this assay. The MV analogues having
a medium length alkyl side chain (8−10 carbon atoms, 1b−
3b) increased the ATP level when employed at 0.1 μM
concentration but suppressed the ATP level when used at
D
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Figure 3. Total ATP levels in FRDA lymphocytes following incubation with test compounds for 24 h in glucose free media (25 mM galactose) to
force the cell to employ OXPHOS. Results are expressed as percentage of total ATP relative to untreated control.
Figure 4. Effects of 1−5 on Δψm in FRDA lymphocytes using the ratiometric fluorescent marker JC-1. Flow cytometric determination of
depolarized Δψm using the ratiometric fluorescent probe JC-1. The actual percentage of cells with intact Δψm were extracted from a two-
dimensional color density dot blot. Results obtained were verified by running duplicates in two independent runs.
Figure 5. Cytotoxicity of phenothiazine analogues 1−5 toward cultured FRDA lymphocytes after incubation for 24 h in glucose-free (galactose)
media to force the cells to rely on mitochondrial ATP production. Flow cytometric determination of cell viability by dual fluorescence labeling
employed calcein-AM and ethidium homodimer-1 as live and dead stains, respectively. The actual percentage of live cells was extracted from a two-
dimensional color density dot blot. Results obtained were verified by running duplicates in two independent experiments.
higher concentrations. The MV analogue with a 12-carbon
atom alkyl side chain (4b) clearly increased the ATP level
when employed at 0.1, 0.25, and 0.5 μM concentrations but
suppressed the ATP level slightly when used at 2.5 μM
concentration. Compound 5b, having a 16-carbon atom alkyl
side chain, clearly increased the ATP level when employed at
0.1, 0.25, and 0.5 μM concentrations but had little effect on
ATP when employed at 2.5 μM concentration. While the
concentrations of the MB/MV analogues that had an effect on
ATP production are significantly higher than the concen-
trations of these compounds that block ferroptosis and are thus
not likely to be pertinent to the antiferroptotic mechanism(s),
E
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Figure 6. Effect of MB/MV analogues on AMPK and p-AMPK protein expression and their potential as AMPK-mediated inhibitors of ferroptosis.
(a) The protein expression levels of AMPK, p-AMPK, and β-actin (loading control) were determined using Western blot analysis in HepG2 cells
following treatment with or without MB/MV analogues. (b) Quantification of p-AMPK/AMPK protein expression. Experiments were performed in
triplicate and data are expressed as mean SEM (*p < 0.05 and **p < 0.01 as compared to control. Statistical analysis was performed using an
unpaired, two-tailed t test). (c) Antiferroptotic effects of MB/MV analogues in FRDA lymphocytes against RSL3-induced lipid peroxidation in the
presence and absence of AMPK inhibitor compound C (10 μM).
compounds that block ATP production are not likely to find
utility as therapeutic agents for Friedreich’s ataxia. Thus, the
data in Figure 3 support the potential therapeutic utility of
analogues such as 1−5.
carbon atom alkyl side chains, exhibited some cytotoxicity at
2.5 μM concentration. The modified methylene violet
analogues with longer alkyl side chains (4b−5b) were not
cytotoxic under any tested condition. From the results, it is
evident that cytotoxicity decreased with increasing lip-
ophilicity. MV and its analogues 1b−4b were less cytotoxic
than the corresponding MB analogues. Thus, the experiments
in Figures 3, 4, and 5 suggest that the lipophilic analogues of
MB and MV studied here have therapeutic potential no less
than that of the clinical agent methylene blue.
Recently, Pratt and his co-workers have studied both
phenoxazines and phenoxazine-N-oxyl as radical trapping
antioxidants.41,42 While neither the substitution patterns of
these compounds nor the experimental methods used to
measure their activities allow direct comparison with the
compounds studied here, both were potent inhibitors of RSL3-
induced ferroptosis in mouse embryonic fibroblasts, and both
exhibited better potency under the conditions employed than
Fer-1 and Lip-1. These observations reinforce the results
obtained here and suggest new alternatives to the structures
studied in the present report.
MB has been shown to utilize multiple molecular targets and
to have beneficial effects clinically.28−30 Recent studies
indicated that MB regulates the energy-sensing AMP-activated
protein kinase (AMPK).43,44 The energy metabolism of the
AMPK signaling pathway is also known to impact ferroptosis
regulation.43,44 Some groups studying ferroptosis have found
interesting but incompletely understood effects on the
regulatory function of AMPK. Song et al. reported that
AMPK-mediated BECN1 phosphorylation promotes ferropto-
sis.45 In comparison, Lee et al. reported that energy stress
activates AMPK that partly suppresses ferroptosis through
effects on lipid metabolism.46 To assess whether AMPK
activation by MB/MV analogues is involved in their
antiferroptotic activity, we first evaluated the effect of MB,
Mitochondrial inner membrane potential (Δψm) is another
key parameter that is essential for the bioenergetic perform-
ance of mitochondria. Accordingly, analogues 1−5 were tested
for their effects on inner mitochondrial membrane potential in
FRDA lymphocytes, using the JC-1 probe as described
previously.33,34 A similar trend was noted for the tested MB/
MV analogues (1−5) on mitochondrial membrane potential,
as they had been for total cellular ATP level (Figure 4). As
demonstrated in Figure 4, MB and MB analogues with short to
medium length side chains (8−12 carbon atoms) exhibited
depolarization of Δψm at 2.5 μM concentration (Figure 4)
while 5a and 5b having 16-carbon side chains exhibited no
effect at 2.5 μM concentration.
MB exhibits a hormetic dose−response, with opposite
effects at low and high doses; this has been attributed to its
unique auto-oxidizing properties.39 In this regard, MB and MB
analogues were tested for their cytotoxicity in FRDA cells
grown in glucose-free medium containing galactose. Under
these nutrient-sensitized conditions, cells rely largely on
OXPHOS to produce their ATP and become more sensitive
to mitochondrial respiratory chain inhibitors than cells grown
in a normal glucose medium.40 Measurement of cytotoxicity
was carried out as described previously.34 Methylene blue and
MB analogues 1a−4a, having 8−12 carbon atom alkyl side
chains, exhibited significant cytotoxicity at 2.5 μM concen-
tration (Figure 5). The hydrophilic properties of MB suggest
poor cell incorporation and/or their facile autoxidation in the
cytosol. More hydrophobic methylene blue analogue 5a having
a 16-carbon side chain lacked cytotoxicity at 0.25 or 2.5 μM
concentrations. MV itself was less cytotoxic than methylene
blue. Methylene violet and MV analogues 1b−3b, having 8−10
F
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MV, and analogues 3a and 3b on phosphorylated and
nonphosphorylated AMPK expression in HepG2 cultured
cells. We found that MB, MV, 3a, and 3b were able to induce a
significant increase in the p-AMPK/AMPK ratio, but only
when assayed at higher concentrations than those at which
they inhibited ferroptosis. (Figures 6a and b). We next sought
to clarify whether inhibition of ferroptosis was due to their
radical trapping antioxidant activity or to their ability to
stimulate phosphorylation of AMPK or both. We performed a
further experiment for these compounds in FRDA lympho-
cytes involving RSL3-induced lipid peroxidation in the
presence and absence of compound C (6-[4-(2-piperidin-1-
yl-ethoxy)phenyl]-3-pyridin-4-yl-pyrazolo[1,5-a]pyrimidine), a
pan inhibitor of AMPK phosphorylation. Figure 6c shows no
significant difference in the antiferroptotic activity of the tested
compounds in the presence or absence of compound C against
RSL3-induced lipid peroxidation at the tested concentrations.
The lipid peroxyl radical is a key molecule during the onset of
ferroptosis and the treatment with radical scavengers specific
for lipid peroxyl radicals represents a promising pharmacologic
approach to prevent ferroptosis. Phenothiazines were pre-
viously reported to show lipid peroxidation-preventing effects
of ferroptosis that were considered to be due to their
antioxidant functions.47−49 Again, when MB was acetylated
to eliminate its redox activity, it completely lost its ability to
suppress lipid peroxidation despite glutathione depletion in
RSL3-treated FRDA lymphocytes (Figure 6c, Table 1). These
findings support the interpretation that the MB/MV analogues
function primarily as lipid peroxyl radical scavengers,
contributing to the prevention of ferroptosis primarily by
suppressing lipid peroxidation.
In summary, given the favorable radical trapping antioxidant
activity of the MB/MV derivatives relative to Fer-1 and Lip-1,
the antiferroptotic activity of lipophilic MB/MV analogues was
investigated, as were other properties of the molecules essential
to their therapeutic utility, such as their effects on ATP
production. Ferroptosis was induced in cultured FRDA cells by
GPX4 inhibition with (1S,3R)-RSL3 or by the cystine/
glutamate antiporter (system xc− inhibitor erastin). The
lipophilic MV analogues were similar to, or had better potency
than, ferrostatin-1 and liproxstatin-1. Clearly, there is an
optimal side chain length for the MB/MV analogues. In
summary, we have identified potent new ferroptosis inhibitors
having structures distinct from those of ferrostatin and
liproxstatin. These may enable the identification and develop-
ment of improved ferroptosis inhibitors useful for the therapy
of several diseases. While not addressed in the current study, it
will certainly be necessary to define the nature of the cellular
uptake and distribution of new MB/MV derivatives, as well as
the critical cellular loci responsible for the effect(s) of such
compounds.
AUTHOR INFORMATION
Corresponding Authors
■
Omar M. Khdour − Biodesign Center for BioEnergetics, Arizona
State University, Tempe, Arizona 85287, United States;
Email: khdour@asu.edu
Sidney M. Hecht − Biodesign Center for BioEnergetics and
School of Molecular Sciences, Arizona State University, Tempe,
Arizona 85287, United States; orcid.org/0000-0002-5429-
2462; Email: sid.hecht@asu.edu
Authors
Jun Liu − Biodesign Center for BioEnergetics, Arizona State
University, Tempe, Arizona 85287, United States
Indrajit Bandyopadhyay − Biodesign Center for BioEnergetics
and School of Molecular Sciences, Arizona State University,
Tempe, Arizona 85287, United States
Lei Zheng − Biodesign Center for BioEnergetics, Arizona State
University, Tempe, Arizona 85287, United States
Complete contact information is available at:
https://pubs.acs.org/10.1021/acsmedchemlett.0c00293
Funding
This work was supported in part by a research grant from the
Friedreich’s Ataxia Research Alliance (FARA).
Notes
The authors declare no competing financial interest.
ABBREVIATIONS
■
FRDA, Friedreich’s Ataxia; MB, methylene blue; MV,
methylene violet; FXN, frataxin; ROS, reactive oxygen species;
GPX4, glutathione peroxidase 4; GSH, glutathione; FSP-1,
ferroptosis-suppressor-protein 1; Fer-1, ferrostatin; Lip-1,
liproxstatin-1; RSL-3, RAS-selective lethal 3; α-TOH, α-
tocopherol; OXPHOS, oxidative phosphorylation; AMPK,
AMP-activated protein kinase
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* Supporting Information
The Supporting Information is available free of charge at
https://pubs.acs.org/doi/10.1021/acsmedchemlett.0c00293.
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Details of the synthesis and characterization of new
compounds, as well as bioassay protocols (PDF)
NMR spectra of compounds 10−15, 1a−4a, and 1b−4b
(PDF)
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