Bioorganic & Medicinal Chemistry Letters
Small molecule inhibitors of anthrax edema factor
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Guan-Sheng Jiao , Seongjin Kim , Mahtab Moayeri , April Thai , Lynne Cregar-Hernandez ,
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Linda McKasson , Sean O’Malley , Stephen H. Leppla , Alan T. Johnson
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Hawaii Biotech, 650 Iwilei Road, Suite 204, Honolulu, HI 96817, USA
Laboratory of Parasitic Diseases, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, MD 20892, USA
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Article history:
Anthrax is a highly lethal disease caused by the Gram-(+) bacteria Bacillus anthracis. Edema toxin (ET) is a
major contributor to the pathogenesis of disease in humans exposed to B. anthracis. ET is a bipartite toxin
composed of two proteins secreted by the vegetative bacteria, edema factor (EF) and protective antigen
Received 31 August 2017
Revised 23 November 2017
Accepted 24 November 2017
Available online xxxx
(
PA). Our work towards identifying a small molecule inhibitor of anthrax edema factor is the subject of
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this letter. First we demonstrate that the small molecule probe 5 -Fluorosulfonylbenzoyl 5 -adenosine
FSBA) reacts irreversibly with EF and blocks enzymatic activity. We then show that the adenosine por-
(
Keywords:
Anthrax
Edema factor
Covalent inhibitor
tion of FSBA can be replaced to provide more drug-like molecules which are up to 1000-fold more potent
against EF relative to FSBA, display low cross reactivity when tested against a panel of kinases, and are
nanomolar inhibitors of EF in a cell-based assay of cAMP production.
Ó 2017 Elsevier Ltd. All rights reserved.
Anthrax results from infection by Bacillus anthracis and is highly
lethal unless quickly diagnosed and treated. B. anthracis makes two
bipartite toxins generated by the combination of three proteins
secreted by the bacteria. Edema Toxin (ET) composed of edema fac-
tor (EF), an adenylate cyclase,1 plus protective antigen (PA), and
Lethal Toxin (LT), made of lethal factor (LF), a zinc-dependent met-
EF is the cause of massive edema seen in some human patients
infected by B. anthracis. Recent ex vivo studies using a rat aortic
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ring model implicate ET as a contributor to shock in the host due
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to arterial relaxation. Due to their unique ability to evade normal
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clearance mechanisms of the host, both toxins can contribute to
mortality and morbidity of human patients long after the bacteria
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alloproteinase, plus PA, are considered major contributors to the
have been cleared from the blood stream by antibiotics.
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pathogenesis and death of humans exposed to B. anthracis.
Competitive
and allosteric
inhibitors of EF have been
At present, antibiotics and antibody therapies are available to
prevent the onset of disease following exposure; however, once a
patient is symptomatic these agents rapidly lose their effective-
ness. This is due to the release of ET and LT immediately following
reported. However, these compounds display low affinity, or are
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derivatives of ATP which exhibit poor drug-like properties mak-
ing them unlikely candidates for therapeutic intervention to treat
anthrax in humans. To address this unmet need, we present below
our initial studies towards identifying potent small molecule inhi-
bitors of anthrax EF.
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germination, and the rapid uptake of these toxins by cell surface
receptors which are ubiquitous throughout the host. Once inside
cells, EF and LF are no longer accessible to antibodies and initially
act to promote the infection by disarming the body’s immune sys-
The lack of an FDA approved small molecule drug to treat EF
intoxication results in part from the challenging nature of the
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tem. Both EF and LF target cell signal pathways resulting in abnor-
problem to be solved. In addition to the need for specificity, a sec-
mal cell function and cell death.6 Recent studies using mice
modified for expression of anthrax toxin receptors have demon-
strated that lethality from exposure to ET and LT results from each
ond consideration is the very high turnover rate (kcat ꢀ 1500 s
ꢁ1
)
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exhibited by this enzyme. A third issue is the very high intracel-
lular concentrations (ꢀ5 mM) of its substrate, ATP, which pre-
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toxin targeting specific tissues of the host. In the case of ET, hep-
sents a significant challenge for any competitive inhibitor.
atocytes appear to be the major target, whereas LT primarily
induces lethality through its effects on cardiomyocytes and vascu-
lar smooth muscle cells. Cardiovascular system damage has been
One solution to these problems is to employ a covalent edema
factor inhibitor (EFI) to irreversibly silence the enzyme. Covalent
inhibitors appear well suited for the treatment of acute diseases
and may be used at lower doses due to their higher efficiency
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linked to the intracellular activity of LF in the rabbit model while
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and lower need for optimized PK parameters.
This approach
also addresses the problem associated with active toxin remaining
after antibiotic treatment since any circulating and intracellular EF
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960-894X/Ó 2017 Elsevier Ltd. All rights reserved.
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