Available via license: CC BY 4.0
Content may be subject to copyright.
Deuterated Drugs and Biomarkers in the COVID-19 Pandemic
Ross D. Jansen-van Vuuren,*Luka Jedlovcnik, Janez Kosmrlj, Thomas E. Massey, and Volker Derdau
Cite This: ACS Omega 2022, 7, 41840−41858
Read Online
ACCESS Metrics & More Article Recommendations
ABSTRACT: Coronavirus disease 2019 (COVID-19) is a highly contagious
disease caused by the severe acute respiratory syndrome coronavirus 2 (SARS-
CoV-2). Initially identified in Wuhan (China) in December 2019, COVID-19
rapidly spread globally, resulting in the COVID-19 pandemic. Carriers of the
SARS-CoV-2 can experience symptoms ranging from mild to severe (or no
symptoms whatsoever). Although vaccination provides extra immunity toward
SARS-CoV-2, there has been an urgent need to develop treatments for COVID-
19 to alleviate symptoms for carriers of the disease. In seeking a potential
treatment, deuterated compounds have played a critical role either as
therapeutic agents or as internal MS standards for studying the pharmacological
properties of new drugs by quantifying the parent compounds and metabolites.
We have identified >70 examples of deuterium-labeled compounds associated
with treatment of COVID-19. Of these, we found 9 repurposed drugs and >20
novel drugs studied for potential therapeutic roles along with a total of 38
compounds (drugs, biomarkers, and lipids) explored as internal mass spectrometry standards. This review details the synthetic
pathways and modes of action of these compounds (if known), and a brief analysis of each study.
■INTRODUCTION
Drug Development and Discovery for COVID-19.
COVID-19 is primarily characterized as an acute respiratory
illness caused by a droplet-borne coronavirus, SARS-CoV-2,
also an RNA virus. By August 2022, the COVID-19 pandemic
had resulted in almost 600 million infections, ∼6.4 million
deaths, major disruption to global trade and travel, and closure
of local businesses. While most patients experience a mild to
moderate respiratory infection and fever, and can recover
without the need for special treatment, for some, COVID-19 is
more severe, leading to major respiratory shutdown and
multiple organ failure, requiring intensive care. Immunocom-
promised people are at greater risk of experiencing severe
COVID-19 symptoms or death, while their immune response
to vaccination is not as strong as in nonimmunocompromised
people.
While vaccination has oered the most eective way to
avoid experiencing a serious case of COVID-19, treatment is
still needed to reduce the symptoms and hasten the healing
process. Two major approaches that have been taken to
discover drugs that could be used to treat COVID-19 include
the repurposing of known drugs
1
and the development of
novel drugs.
2
Both types of drugs are proposed to act either by
disrupting a certain component of the life cycle of the
coronavirus
3
or as anti-inflammatories,
4
altering the body’s
response to the virus. Currently, several treatments are FDA
approved,
5
while much research (including clinical trials) is
underway to demonstrate the ecacy of treatments against
COVID-19.
6
In seeking a potential treatment for COVID-19,
deuterated drugs (compounds in which some hydrogens have
been exchanged for deuterium) have been explored either as
therapeutic agents or as internal MS standards for studying the
pharmacological properties of new drugs by quantifying the
parent compound and possible metabolites by liquid
chromatography/mass spectrometry (LC/MS) assays. To
date, no work has been carried out to review these deuterated
compounds. Thus, this paper oers a comprehensive review of
all deuterated compounds explored as internal standards,
potential treatments, or biomarkers during the development of
treatment for COVID-19 along with their synthetic pathways
and modes of action (where this is known).
Since the FDA approval of the first deuterated drug in
2017,
7,8
there has been a major surge of interest in the
development of new deuteration methodologies and the
preparation of novel deuterium-labeled compounds.
9
The
incorporation of deuterium has been found to overcome drug
limitations related to toxicity, bioavailability, and pharmacoki-
netics, mostly by altering the metabolic profile of the drug of
concern.
10,11
Also, the formation of deuterated compounds as
Received: July 2, 2022
Accepted: October 18, 2022
Published: November 13, 2022
Review
http://pubs.acs.org/journal/acsodf
© 2022 The Authors. Published by
American Chemical Society 41840
https://doi.org/10.1021/acsomega.2c04160
ACS Omega 2022, 7, 41840−41858
internal standards for analytical purposes is advantageous due
to the availability and economical value of deuterium over
other isotopes such as 15N or 18O.
By collating studies involving deuterated compounds related
to COVID-19, this review seeks to be an easy reference tool for
practitioners of isotope exchange chemistry as well as for
medicinal chemists, especially those involved in the develop-
ment of either COVID-19 treatments or methods to study the
ecacy of such therapies. By illuminating the approaches taken
in the synthesis and applications of deuterated drugs and
biomarkers, we hope to inspire new ideas for deuterium-
labeled compounds beyond what is in the literature.
Deuterated Compounds. Deuterium (D) is an isotope of
hydrogen (H), having the same proton number but double the
mass due to a neutron in the nucleus. Thus, exchanging H for
D results in a slight increase in the activation energy (EA)
needed for bond cleavage (1.4 kcal/mol) as well as a lower
reaction rate compared with C−H (C−D has a lower zero-
point energy), an eect known as the primary deuterium
kinetic isotope eect (DKIE) and expressed as kH/kD, the ratio
of the reaction rate constants of C−H versus C−D bond
cleavage.
12
Although the EA value seems negligible, especially
considering the nearly constant body temperature of 37−39
°C, it has been shown to alter the metabolic profiles of
compounds whose metabolic pathways are dependent on C−
H bond cleavage.
11
Examples include drugs metabolized by
cytochrome P450s or aldehyde oxidases, deuteration of which
can result in pharmaceutical compounds with improved
pharmacokinetics and reduced toxicity
13
but equal potency
to the parent drug.
11,14
However, some deuterated drugs
provide no improvement in terms of the metabolic process,
while others are prone to unexpected metabolic switching
resulting in the deuterated analogue having no pharmacoki-
netic advantages over the parent compound.
15
Since deuterated drugs are identical spatially and have the
same charge distribution as their nondeuterated analogues, in
most cases, both share similar physiochemical properties, e.g.,
lipophilicity,
16
and therefore interact comparably with cellular
components such as enzymes, ion channels, receptors, and
transporters.
7,17
However, a few studies have shown that
deuterium labeling can have a complex eect on intermolecular
interactions
18
and binding to enzymes,
19
while other studies,
particularly focused on histamine receptors, have questioned
the fact that labeling retains the same interactions with the
target compared to the protium-containing compound.
20
In this paper, we will review deuterated drugs that have been
explored as possible therapeutics for COVID-19 as well as
those which have been used as internal MS standards for
probing the properties of new or repurposed drugs being
explored as treatment options for COVID-19.
Figure 1. Three main advantages potentially provided by deuterated drugs: increased (A) safety, (B) tolerability, and (C) bioavailability. These are
achieved, respectively, by (A) “metabolic shunting”, resulting in reduced exposure to undesirable (toxic or reactive) metabolites, (B) reduced
systemic clearance, resulting in increased half-life, and (C) first-pass metabolism, resulting in higher bioavailability of the nonmetabolized drug.
AUC is area under the curve and represents drug exposure over time; Cmax is the maximum or peak concentration of a drug. Adapted with
permission from ref 24. Copyright 2014 The Pharmaceutical Society of Japan.
ACS Omega http://pubs.acs.org/journal/acsodf Review
https://doi.org/10.1021/acsomega.2c04160
ACS Omega 2022, 7, 41840−41858
41841
■DEUTERATED DRUGS AS THERAPEUTICS FOR
THE TREATMENT OF COVID-19
Some drugs that have demonstrated ecacy against SARS-
CoV-2 require relatively high dosages and/or result in patients
experiencing adverse eects. This is of particular concern since
patients suering from COVID-19 already have compromised
immune systems. For some COVID-19 antiviral and anti-
inflammatory drugs, toxicity is attributed to the parent
compound, while in some cases, reactive metabolites are
implicated. Furthermore, COVID-19 causes inflammation of
the liver, resulting in suppression of the hepatic cytochrome
P450 enzymes, causing reduced drug clearance, higher plasma
drug concentration, and severe toxicity in some cases,
triggering further deterioration in the condition of COVID-
19 patients.
21
The potential for toxicity of COVID-19 antivirals
is amplified by both the presence of a complex disease and the
fact that multiple drugs are often used concurrently.
22
For
example, cardiac dysfunction attributable to COVID-19 can be
exacerbated by some COVID-19 drugs.
23
Furthermore, drug−
drug interactions at the level of drug disposition, including
metabolism and transport, could increase plasma concen-
trations of certain drugs and their metabolites, further
increasing cardiac risk.
Deuteration presents an opportunity to improve three major
characteristics of drugs: (i) the safety of the drug via
“metabolic shunting” (the generation of less toxic metabolites),
(ii) higher drug tolerability, which means that the drug can be
administered in lower dosages and that it remains at a more
constant blood plasma concentration (rather than having
pronounced peaks and troughs), and (iii) increased drug
bioavailability (Figure 1).
24
This has invigorated research into
the deuteration of known drugs which have been removed
from clinical studies or circulation due to safety concerns,
resulting in the recent approval of the first deuterated drug for
commercial use by the U.S. Food and Drug Administration
(FDA). This drug is deutetrabenazine (1), the deuterated
version of tetrabenazine (2) (Figure 2), used for the treatment
of Huntington’s disease, an involuntary movement disorder.
25
In the case of 1, exchanging the six methoxy protiums with
deuteriums slightly alters the metabolism of the drug,
increasing its safety and tolerability by conferring upon it an
extended half-life and a more stable plasma concentration.
7
As
no head-to-head trial with tetrabenazine has ever been
performed, it is still not clear if there is a significant safety
advantage. However, the recommended daily dosage of
tetrabenazine is approximately double the recommended
starting dosage of deutetrabenazine.
26
Deuterated versions of various medicinal compounds are
now being explored as treatments; as of 2019, there were more
than 20 deuterated drugs in clinical trials, with 6 having
reached Phase III, while 200−300 patents had been filed for
deuterated medicinal compounds.
11
Several deuterated drugs
were explored as potential therapeutics for COVID-19. These
compounds are isotopologues (i.e., molecules that only dier
from the parent molecule in their isotopic composition) of
either repurposed drugs or completely new drugs. These
compounds are reviewed in the following section.
Repurposed Drugs. Deupirfenidone (LYT-100). Deupir-
fenidone or LYT-100 (3, CAS No. 1093951-85-9) is the
deuterated form of pirfenidone (4) (Figure 2), originally used
for the treatment of idiopathic pulmonary fibrosis, a severe
lung disease.
27
LYT-100, developed by PureTech Health
(based in Boston) and currently in Phase 2 clinical trials
(ClinicalTrials.gov Identifier NCT04652518), was already
under consideration to target lymphedema (lung inflamma-
tion) and lung fibrosis prior to the pandemic and was therefore
ideally suited to treat patients suering from long COVID-
19.
27,28
Metabolism of pirfenidone is carried out by
cytochrome P450 (CYP1A2) and involves oxidation of the
methyl group at C-5 of the pyridin-2(1H)-one ring of 4,
leading to the formation of the primary metabolite, 5-carboxy-
pirfenidone (5), which is inactive compared with 3or 4. Thus,
replacing hydrogen with deuterium on the methyl group ought
to inhibit the metabolism of the drug and enable less frequent
dosing, and this was shown to be the case through a 2021
Phase 1 clinical trial (ClinicalTrials.gov Identifier
NCT04243837).
29
When 3undergoes metabolism, the
secondary metabolites 5-hydroxymethylpirfenidone-d2(6)
and 4′-hydroxypirfenidone-d3(7) are formed in low
concentrations.
28
Deupirfenidone (pirfenidone-d3) (3) can be prepared via
many routes, mostly involving the use of alkyllithiums at
cryogenic temperatures.
30,31
More recently, however, three
routes have been reported which employ milder conditions
and have higher yields. First, 3can be prepared via visible light
(390 nm) driven, TBADT/thiol-catalyzed deuterium labeling
in 85% yield but with only 54% D incorporation.
32
Second, 3
can be prepared via Ni-catalyzed methylation with iodo-
methane-d3in >69% yield on a multigram scale (% D not
provided).
33
Finally, Falb et al. demonstrated that 3could be
Figure 2. Chemical structures of deutetrabenazine (1), tetrabenazine (2), deupirfenidone (3), and pirfenidone (4). Also shown are compound 5,
the primary metabolite of both 3and 4, and compounds 6and 7, the secondary metabolites of 3.
ACS Omega http://pubs.acs.org/journal/acsodf Review
https://doi.org/10.1021/acsomega.2c04160
ACS Omega 2022, 7, 41840−41858
41842
prepared on a multigram scale in 88% yield with >99% D
enrichment using Suzuki−Miyaura cross-coupling condi-
tions.
34
Initially the deuteromethyl group was installed using
expensive potassium methyl trifluoroborate (CD3BF3K).
Despite the fact that this route enabled the product to be
formed in 88% yield, however, the procedure was modified to
avoid the use of this chemical by employing greener and less
expensive CD3B(OH)2(Scheme 1). The reaction could be
performed by commencing with 8and proceeding either via:
(a) the “methylation-last” route (involving coupling of
methylboronic acid-d3with 9using Pd(OAc)2and the RuPhos
ligand) in 84% overall yield, or (b) the “methylation-first”
route (8→10→11→3).
Regrettably, for the latter, the percent yield for the Chan−
Lam conversion 11 →3was not provided. We can assume it is
similar to the yield obtained for the Chan−Lam coupling
reaction of the nondeuterated analogue of 11 with phenyl
boronic acid (70%)
34
giving an overall yield for the
“methylation first” route of 45%.
RNA-Dependent RNA Polymerase (RdRp) Inhibitors. The
enzyme RNA-dependent RNA polymerase (RdRp) is an
important therapeutic target in RNA virus-caused diseases,
including SARS-CoV-2. Nucleoside inhibitors (typically in
their nucleoside triphosphate form) act by binding to the
RdRp protein at the enzyme active site, therefore interfering
with the RNA synthesis step.
35,36
In this short subsection, we
describe deuterated RdRp inhibitors that have been
repurposed for the potential treatment of COVID-19.
Deuterated Oral Remdesivir Derivative VV116. Remdesivir
(12) (Figure 3), an intravenously administered nucleotide
prodrug, is currently approved for treatment of COVID-19 by
the FDA.
5
Two other analogs [molnupiravir (13) and AT-527
(14)], taken orally, are in phase II/III clinical studies for
COVID-19 [ClinicalTrials.gov identifiers NCT04405570
(molnupiravir) and NCT04709835 (AT-527)].
37
Xie et al.
have since developed a deuterated oral anti-SARS-CoV-2
nucleoside candidate, VV116 (15), also under clinical
evaluation (phase II/III) as a COVID-19 therapeutic agent
(ClinicalTrials.gov Identifier NCT05242042).
38
Compound
15, deuterated at C7 of the pyrrolotriazine ring, is a modified
version of GS-441524 (16), the parent nucleoside of
remdesivir (12), which inhibits the replication of SARS-CoV-
2 but mainly targets the liver, whereas COVID-19 is primarily a
lung disease.
Deuteration at this position is hypothesized to inhibit
enzymatic degradation of the ring (either by oxidation of the
double bond or by ring opening of the triazine).
38
In addition,
the inclusion of a tri-isobutyrate ester functionality in 15
improved the in vivo pharmacokinetics compared with that of
the parent nucleoside (16), while its formulation as the
Scheme 1. Suzuki-Coupling Approaches to the Synthesis of Deupirfenidone (3): (a) Methylation-Last Route and (b)
Methylation-First Route
Figure 3. Chemical structures of RdRp inhibitors 12−16.
ACS Omega http://pubs.acs.org/journal/acsodf Review
https://doi.org/10.1021/acsomega.2c04160
ACS Omega 2022, 7, 41840−41858
41843
hydrobromide salt gave it extra water solubility (and other
enhanced physical properties). Compared with other similar
derivatives of 16,15 showed the most favorable physicochem-
ical properties and had superior oral bioavailability, anti-SARS-
CoV-2 ecacy, and safety in mice, rats, and dogs in the
subsequent preclinical study. Furthermore, a recent study
demonstrated the safety and tolerability of 15 in healthy
Chinese patients.
39
VV116 (15) can be prepared in 15% overall yield in 5 steps
commencing with the commercially available substrate 17
(CAS No. 1355357-49-1) (Scheme 2).
38
Compound 17 is
initially iodinated to form intermediate 18 before deuterium is
introduced into 18 using a palladium catalyst complexed with
TMEDA and reduced using NaBD4to form 19 in 55% yield
(≥98% D incorporation). 19 could then be transformed into
15 via a further three steps: OH deprotection to 20
(monodeuterated analogue of 16), amine protection to form
21, and, finally, a one-pot acylation followed by amine
deprotection to 15.
In a subsequent study,
40
Zheng et al. describe a more
ecient approach to the synthesis of 19 which involves
reducing 18 with triethylamine, palladium, and deuterium gas
(D2) at 60 °C for 1 h. In this method, 19 was produced in 92%
yield with D incorporation of 99%. In addition, Zheng and co-
workers reported the synthesis of deuterated analogues of GS-
441524 (20 and 20a−d) along with a comparison of their
ecacy against SARS-CoV-2 in vitro against nondeuterated
GS-441524 (16) (Table 1). Essentially, all of the deuterated
GS-441524 analogs demonstrated similar antiviral activity to
GS-441524 against SARS-CoV-2. A further comparison of the
metabolic profiles and/or IC50 values of 16 with 20 and 20a−d
would add value to the study.
Deuterated Thymine Analogue. ACH-3422, CAS No.
798779-31-4 (abbreviated 22) [(Scheme 3a), the deuterated
analogue of PSI-7851 (23), is also an RNA-dependent RNA
polymerase and has been considered for the treatment of
COVID-19.
35,41
22 contains three deuteriums: one on the
pyrimidine and two on the ribose group side chain.
Substituting hydrogen for deuterium at these positions was
proposed to improve the safety profile of the parent drug 23 by
enabling a more stable drug concentration and reducing the
production of toxic metabolites. Indeed, in a separate study, 22
was well tolerated and did not induce any serious adverse
events in both healthy volunteers and hepatitis C patients.
41
Importantly, among COVID-19 patients, increasing the dose of
22 resulted in increased viral decline, and viral clearance was
achieved in 50% of patients after the administration of 700
mg/day over 2 weeks.
35
The preparation of 22 commences with the initial synthesis
of deuterated acetonide 26 by a combined reduction−
deprotection and H/D exchange at the α-C of acetonide
ester 25 (Scheme 3),
42
itself prepared from commercially
available 2′-C-methyl-uridine, 24 (CAS No. 31448-54-1).
43
Although an initial 1H NMR spectroscopic analysis of 26
indicated ∼85% deuterium incorporation at the 5-uracil
position, this was increased to >98% by filtration, removal of
EtOD, and addition of more D2O, followed by reheating of the
resulting mixture at 95 °C.
42
Subsequent deprotection of 26
with aqueous HCl provided nucleoside 27, which was then
reacted with 28 under Grignard conditions to form 22
Scheme 2. Synthetic Pathway to VV116 (15) Commencing with Nondeuterated Substrate 17
a
a
Adapted with permission from ref 38. Copyright 2021 Springer Nature.
Table 1. Inhibition of SARS-CoV-2 Replication and Cellular
Toxicity by Deuterated Remdesivir Analogues 20 and 20a−
d In Vitro (in Vero E6 cells) Relative to Nondeuterated
Remdesivir Nucleoside 16
a
compound R1R2R3R4R5EC50 (μM) CC50 (μM)
16 H H H H H 0.33 >100
20 D H H H H 0.24 >100
20a H D D H H 0.25 >100
20b D D D H H 0.23 >100
20c D D D D D 0.23 >100
20d H D D D D 0.31 >100
a
Adapted with permission from ref 40. Copyright 2022 Elsevier.
ACS Omega http://pubs.acs.org/journal/acsodf Review
https://doi.org/10.1021/acsomega.2c04160
ACS Omega 2022, 7, 41840−41858
41844
(regrettably, no yields were reported for the reaction
procedure).
42,43
Deuterated Dexamethasone. Dexamethasone (29), a
corticosteroid and anti-inflammatory agent, has demonstrated
ecacy as a treatment for COVID-19 patients.
44
Thus, the
deuterated analogue
45
is in demand�both as an internal MS
standard and to test as a therapeutic agent with improved
bioavailability and safety profile (compared with dexametha-
sone). As a glucocorticoid, long-term use of dexamethasone
has the potential to produce a wide range of undesirable
eects, many mediated by the glucocorticoid receptor which
regulates the expression of a vast array of genes. When used for
relatively short periods (up to 2 weeks), dexamethasone is not
usually expected to produce serious toxicities
46
although higher
doses can produce neurological eects, stomach ulcers,
autoimmune and cardiovascular events, and pancreatitis.
44,47
Thus, lowering the dosage while maintaining the necessary
bioavailability of the corticosteroid by regulating the
metabolism could oer a safer option for the use of
dexamethasone for COVID-19 as well as in treating other
conditions. In a similar fashion to the other drugs we have
discussed, deuteration could enable this by hindering the
metabolism of dexamethasone.
Furthermore, studies involving comparisons between dexa-
methasone and its deuterated analogues could provide valuable
information regarding the role of the metabolites in instigating
adverse side eects (not yet studied, as far we can ascertain),
which could remove any uncertainty regarding its role as a
therapeutic agent for COVID-19. The main routes of
metabolism of dexamethasone (29) involve hydroxylation at
C-6 (by CYP3A4 enzymes) and replacement of the 2-
hydroxyethan-1-one with a ketone group at C-17 (by
CYP17), resulting in the metabolites 30 and 31, respectively
(Scheme 4).
48
The synthesis of a deuterated version of 29 was first
reported in 1997 by Best et al., although the exact positions of
H/D exchange were not given.
45
More recently, Darshana et al.
reported the H/D exchange of dexamethasone at C-6 based on
the in situ spontaneous generation of deuterium chloride
(DCl) from a prenyl chloride (32) under mild conditions (rt,
48 h) in CD3OD (Scheme 5a).
49
The generated DCl induced
H/D exchange within the dexamethasone at the αand γ
positions next to the carbonyl groups of 29 via acid catalysis
chemistry, resulting in 74% deuterium incorporation in 33,
produced in 98% yield (Scheme 5b).
New Drugs. Deuterated Arachidonic Acid Ethyl Ester.
The ethyl ester of arachidonic acid (34) (Scheme 6) is a major
Scheme 3. (a) Chemical Structures of ACH-3422 (22) and the Parent Drug PSI-7851 (23); (b) Synthesis of ACH-3422 by
Initial Preparation of Deuterated Acetonide 26 Followed by a Double OH Deprotection to 27, which Then Reacts with 28 To
Form the Deuterated Thymine Analogue 22
Scheme 4. Metabolic Profile of Dexamethasone (29)
ACS Omega http://pubs.acs.org/journal/acsodf Review
https://doi.org/10.1021/acsomega.2c04160
ACS Omega 2022, 7, 41840−41858
41845
component of lipid bilayers and the key substrate for the
eicosanoid cascades. 34 is initially hydrolyzed to the acid form
by the enzyme phospholipase A2, prior to enzymatic oxidation,
e.g., by cytochrome P450 enzymes, and the metabolite
products induce inflammatory responses in nearly all tissues,
including lung tissues. Oxidation products of the acid form of
34 are elevated in COVID-19 patients.
50
Thus, one strategy to
interfere with the metabolism is by deuteration at the point of
oxidation (Scheme 6a). Molchanova et al. demonstrated that
deuteration at the bisallylic positions within 34 to form 35
(Scheme 6b) substantially decreases the overall rate of
oxidation when hydrogen abstraction is an initiating event.
51
The researchers also found that oral dosing with 35 resulted in
successful incorporation of 35 into various tissues and
significantly reduced E. coli lipopolysaccharide (LPS)-induced
adverse eects in the lung area. This work therefore suggests
novel therapeutic avenues for reducing lung damage during
COVID-19 infection.
The ethyl ester of arachidonic acid-d6(35) can be
synthesized from the naturally occurring, nondeuterated ethyl
ester of arachidonic acid (34) using a ruthenium catalyst (1
mol %) in quantitative yield,
52
as shown in Scheme 6. The
degree of deuteration was reportedly 32% and 25% at the
monoallylic and 95% at the bisallylic positions of 35,
respectively.
Deuteration at the monoallylic sites may not aect drug
potency since oxidation occurs predominantly at the bisallylic
sites.
53
Furthermore, one study suggests that the addition of
polyunsaturated fatty acids (PUFA) deuterated at the
monoallylic sites does not protect the cells from oxidation.
54
Nevertheless, further studies might show the ecacy or toxicity
of deuterium incorporation at the monoallylic sites. In
addition, PUFAs with only partially deuterated bisallylic
positions seem to be protected the same as that of PUFAs
with completely deuterated bisallylic positions. Moreover, from
the same study, we can conclude that inclusion of just a
fraction of deuterated PUFAs (20−50%) in the total pool of
PUFAs appears to preserve mitochondrial respiratory function
and confers cell protection. Thus, a quantitative study
involving samples of deuterated arachidonic acid to varying
degrees in preserving mitochondrial function might provide
clarification around this aspect.
Deuterated Broad-Spectrum Inhibitors of SARS-CoV-2
3CL Proteases. Dampalla et al.
55
developed the dipeptidyl
inhibitor GC376 (36) (Figure 4) in which the P1, P2, and P3
residues are: a lactam-containing glutamine surrogate, leucine,
and a benzyl acetate, respectively. P1, P2 and P3 refer to
fragments of the inhibitor that target the virus proteases of
SARS-CoV-2 by binding to the active site of MERS-CoV 3-
chymotrypsin-like protease (3CLpro), the protease that is
central to the replication of SARS-CoV-2 (generally known as
the coronavirus main protease, Mpro).
55
The potential to
achieve improved binding interactions was identified by
introducing dierent functionalities at the carbamate R groups
in the inhibitors. These are able to dock within an S4 pocket of
3CLpro surrounded by a set of primarily hydrophobic residues.
It was considered that deuterated variants of GC376 might
possess some superior properties as therapeutic agents
compared to the corresponding nondeuterated GC376 drug,
such as improved pharmacokinetics, lower toxicity, and higher
ecacy. Eleven deuterated variants of GC376 were therefore
studied by replacing hydrogen with deuterium at the
metabolically vulnerable sites of GC376 (the carbamate R
groups, the aromatic ring, and the benzylic carbon).
55
The
structures of the deuterated variants of GC376 are shown in
Scheme 7 (compounds 39a−cand 40a−h). They were
synthesized using a reaction sequence previously employed
Scheme 5. (a) Generation of DCl from Reaction of Prenyl
Chloride 32 with Methanol-d4Followed by (b) Acid-
Catalyzed H/D Exchange of Dexamethasone (29) at C-6
a
a
(a) Adapted with permission from ref 49. Copyright 2021 Royal
Society of Chemistry.
Scheme 6. (a) Hydrogen Abstraction of a Bisallylic
Hydrogen, Where the Key Step of PUFA Oxidation Is
Inhibited by Deuteration; (b) Synthesis of the Ethyl Ester of
Arachidonic Acid-d6(35) from the Nondeuterated
Analogue (34)
a
a
Adapted with permission from ref 51. Copyright 2022 MDPI.
Figure 4. Chemical structure of 3CLpro inhibitor GC376 (36). P1, P2
and P3 are the fragments of the inhibitor known to bind to the active
site of Mpro (the protease that is key to the replication of SARS-CoV-
2).
ACS Omega http://pubs.acs.org/journal/acsodf Review
https://doi.org/10.1021/acsomega.2c04160
ACS Omega 2022, 7, 41840−41858
41846
for the synthesis of nondeuterated analogues. Briefly,
deuterated benzyl alcohols 37a−c(purchased) were reacted
with L-leucine isocyanate methyl ester to yield carbamate
derivatives, which were then hydrolyzed to the corresponding
acids with lithium hydroxide (Scheme 7). The subsequent
coupling of the acid to the glutamine surrogate methyl ester 38
into dipeptides followed by lithium borohydride reduction and
oxidation with Dess−Martin periodinane reagent yielded
aldehydes 39a−c. The bisulfite adducts 40a−cwere generated
by the treatment of 39a−cwith sodium bisulfite. Further
treatment of 40a−cwith acetyl or n-pentyl anhydride resulted
in the corresponding esters 40d and 40e−g, respectively.
Reaction of 39a with benzyl isonitrile (benzyl isocyanide) with
subsequent Dess−Martin oxidation aorded compound 40h.
Regrettably, no % D values were provided by the authors for
the deuterated compounds shown in Scheme 7, although these
are likely to be the same as the commercially obtained alcohols
(37a−c).
Crystal structure investigations revealed that deuteration did
not alter the interactions between the deuterated GC376 (40a)
and the 3CLpro. However, the deuterated variants showed
enhanced activity, and this was attributed to tighter binding to
the target or improved physicochemical properties of the
drug.
55
The presence of the aldehyde group in 39a−cis
associated with toxicity; hence, the inclusion of deuterium was
also meant to reduce the toxicity of these derivatives.
The same authors prepared and evaluated another series of
carbamate derivatives of GC376, deuterated on the alcohol
side R commencing from the alcohol inputs 37d−l(Figure 5).
Synthesis of the inhibitors commencing from alcohols 37d−l
was via a separate process (not shown) involving initial
treatment of the alcohols with N,N′-disuccinimidyl carbonate
followed by coupling of the resulting mixed carbonate to a
Leu-Gln surrogate amino alcohol to form aldehyde (analogous
to 39a−c) and bisulfite (analogous to 40a−c) products.
56
Compounds 37j−lled to fluorinated and deuterated GC376
derivatives. All analogues were prepared to test whether
inclusion of fluorine or deuterium might improve the potency,
physicochemical parameters, and pharmacokinetics of the
inhibitors compared with the corresponding nondeuterated
inhibitor.
In addition, aldehyde 39d (analogous to 39a−c) and
bisulfite 40i (analogous to 40a−c) were prepared by following
this reaction sequence from azetidine alcohol 37m (Figures 5
Scheme 7. Synthesis of Deuterated GC376 Derivatives 39−40 As Described by Dampalla et al.
55,56
Figure 5. Chemical structures of deuterated alcohol substrates 37d−mused in the synthesis of GC376 derivatives 39 and 40.
ACS Omega http://pubs.acs.org/journal/acsodf Review
https://doi.org/10.1021/acsomega.2c04160
ACS Omega 2022, 7, 41840−41858
41847
and 6).
57
The azetidine cap, along with a series of spirocyclic
analogues, was investigated to potentially exploit the new
active site of the protease. The eect of deuteration on
pharmacological activity was investigated by determining the
IC50 values against SARS-CoV-2 3CLpro (0.33 μM for 39d and
0.34 μM for 40i) and comparing these with those of the
corresponding nondeuterated analogues (0.41 and 0.50 μM for
nondeuterated for 39d and 40i, respectively). The authors
anticipate that deuterated variants of similar inhibitors will
likely display improved pharmacokinetics in future studies.
Alcohols 37d−hwere purchased, while 37i−lwere obtained
by treatment of the commercially available precursor carboxylic
acid with carbonyl diimidazole followed by the addition of
NaBD4(general synthesis shown in Scheme 8). No % D values
were reported for any of the synthesized alcohols, but it could
be assumed that the purchased deuterated alcohols had high
(>98%) % D incorporation which would have been carried
through to the final products. An alternative synthesis of α,α-
dideuterio alcohols directly from feedstock carboxylic acids
using D2O as the D source and avoiding pyrophoric alkali
metal deuterides such as NaBD4was reported by Szostak et
al.
58
This reaction proceeds after the activation of Sm(II) with
a Lewis base and results in excellent levels of % D
incorporation (85−96 D2) across a wide range of substrates.
As has already been pointed out, Mpro inhibitors are
promising candidates for the treatment of COVID-19 because
Mpro plays a crucial role at the onset of viral replication.
Furthermore, Mpro is conserved among various variants of
concern. Thus, any interruption of its catalytic activity could
represent a relevant strategy for the development of
anticoronavirus drugs. However, the majority of Mpro inhibitors
belong to a class of compounds known as “peptidomimetics”
(synthetic molecules designed to mimic the structural domain
within a natural protein
59
); thus, they often possess poor
pharmacokinetic properties and oral bioavailability.
60,61
To
counter this, Quan et al. developed a series of orally available
Mpro inhibitors with potent in vivo antiviral activity against
emerging variants of SARS-CoV-2.
61
The general structure of
the inhibitor (41) is shown in Scheme 9a. The various
fragments (α-ketoamide, pyridine, R1, and R2) occupy the
main four pockets of Mpro. Due to two stereocenters in 41
(with a fixed S-configuration in 1-(4-fluorophenyl)ethan-1-yl
substituent R3), the molecules are mixtures of epimers in which
the (R)-epimers displayed much higher potency than the
corresponding (S)-epimers, while the most active (R)-epimers
rapidly convert to the less active epimer (S) in vivo, likely due
to the presence of an exchangeable hydrogen in the chiral
carbon center linking the two amides. Thus, to prevent or
reduce configuration conversion, the authors incorporated
deuterium at this position, forming deuterated Mpro inhibitors
with general structure 42 (in addition to nondeuterated
inhibitors 41). The deuterated inhibitors were prepared using
an Ugi 4-component reaction (Ugi-4CR). This involved the
fusion of (S)-2-hydroxypropanoic acid, nicotinaldehyde-
formyl-d1(43), an amine (R1-NH2), and an isocyanide (R2-
Figure 6. Chemical structures of azetidine-containing inhibitors 39d
and 40i.
Scheme 8. Generic Schematic Showing Preparation of
Deuterated Alcohols (37i−l) from Commercially Available
Carboxylic Acid Precursors
Scheme 9. (a) General Chemical Structure of Mpro Inhibitor 41 Explored by Quan et al.;
61
(b) Reaction To Form General
Deuterated Mpro Inhibitor, 42: (i) Formyl-Selective Deuteration of Nicotinaldeyde to 43, (ii) Classical One-Pot Ugi-4CR To
Form Diamine Derivative 44, and (iii) Dess−Martin Oxidation of 44 to 42; (c) Chemical Structures of 45 (Y180) and Its
Nondeuterated Analogue 46
ACS Omega http://pubs.acs.org/journal/acsodf Review
https://doi.org/10.1021/acsomega.2c04160
ACS Omega 2022, 7, 41840−41858
41848
CN) to generate diamine derivative 44, which was then
converted to the general inhibitor (42) by a Dess−Martin
oxidation [Scheme 9b(ii)]. Compound 43 can be generated
from the nondeuterated nicotinaldehyde by a repeated
reduction with NaBD4(to the deuterated pyridin-3-ylmetha-
nol) followed by Dess−Martin oxidation to the aldehyde three
times, sucient to generate compound 43 with D incorpo-
ration > 98% [Scheme 9b(i)]. We note that Dong et al.
62
reported an alternative single-step route to 43 in 82% yield
(94% D incorporation) that is formyl selective, uses readily
available and safe D2O, and involves combined hydrogen-atom
transfer photocatalysis and thiol catalysis. Similarly, Geng et
al.
63
reported an alternative single-step route to substituted
analogues of 43 using readily available and safe D2O under
mild reaction conditions with uniformly high (>95%) levels of
D incorporation.
The authors prepared 11 deuterated inhibitors along with
numerous nondeuterated inhibitors.
61
Although the generated
compounds showed similar potency to the nondeuterated
analogues, conversion from the (R)- to the (S)-epimer was
substantially reduced. Of the 11 isotopologues prepared, Y180
(45) (Scheme 9c) proved to be the most eective among the
tested inhibitors with the lowest rate of epimerization and an
IC50 of 8.1 nM against SARS-CoV-2 Mpro (compared with
∼13.3 μM for its nondeuterated analogue, 46). 45 protected
against wild-type SARS-CoV-2, B.1.1.7 (Alpha), B.1.617.1
(Kappa), and P.3 (Theta) with EC50 values of 11.4, 20.3, 34.4,
and 23.7 nM, respectively. Oral treatment with 45 displayed a
remarkable antiviral potency and substantially ameliorated the
virus-induced tissue damage in both the nose and the lung of
B.1.1.7-infected K18-human ACE2 (K18-hACE2) transgenic
mice. Furthermore, treatment of B.1.617.1-infected mice
(lethal infection model) with 45 improved their survival rate
from 0 to 44.4% (P= 0.0086). Importantly, 45 was also highly
eective against the B.1.1.529 (Omicron) variant both in vitro
and in vivo. This is a nice example where deuterium does not
directly aect the metabolism of the molecule but instead acts
to prevent epimerization, thus enabling the preparation of a
single, more ecient epimer. This approach is also known as
deuterium-enabled chiral switching (DECS) and is a powerful
tool to yield chirally pure drugs from chemically interconvert-
ing racemates, often resulting in therapeutic agents with
improved ecacy and stability and reduced toxicity.
11,64
■DEUTERATED DRUGS AS INTERNAL MS
STANDARDS
In general, internal MS standards are useful in applications in
which the amount of an analyte of interest within a mixture
(e.g., in urine, blood) varies or is reduced during a process, e.g.,
due to adsorption, but needs to be accurately quantified
throughout the procedure By including an isotope-labeled
standard of known concentration within the mixture, it is
possible to provide a measure of control throughout the
procedure by correcting for analyte losses, therefore ensuring
the accuracy and precision of reported concentrations.
65,66
Generally, most quantitative analytical methods rely on mass
spectrometry-related techniques such as LC/MS assays.
Isotopologues are the most practical to use as internal MS
standards as they usually coelute with the parent compound
chromatographically and ionize in the same manner as the
parent compound during mass spectrometry. However,
deuterated compounds may demonstrate unexpected results
such as dierent retention times for the analyte and deuterated
internal MS standard from the reversed-phase LC or dierent
extraction recovery and loss of deuterium due to H/D
exchange. This is especially true for standards containing
more than six deuterium atoms or with the label directly
neighboring a basic nitrogen atom.
67
For this reason, 13C-,
15N-, or 17O-labeled compounds may be more appropriate than
deuterium-labeled compounds.
65
On the other hand, H is
typically more abundant in individual compounds, and
deuterium is usually more cheaply and easily incorporated
(e.g., via late-stage deuteration
68
), so deuterated internal MS
standards are of great interest.
65,67
To successfully separate
compounds and prevent “cross talk”, the amount of
deuteration can be varied (M + 3 is a standard requirement
for hydrocarbons).
67
Sometimes naturally occurring isotopes
of the analyte also contribute significantly to the signal of the
Figure 7. Chemical structures of internal MS standards 47−51.
ACS Omega http://pubs.acs.org/journal/acsodf Review
https://doi.org/10.1021/acsomega.2c04160
ACS Omega 2022, 7, 41840−41858
41849
internal MS standard. This becomes more apparent in
isotopically rich compounds, such as those containing sulfur,
chlorine, or bromine, compounds with higher molecular
weight, and those at high analyte/internal MS standard
concentration ratios.
69
However, each case is dierent, and
the choice of which isotope label (or alternative internal MS
standard) to use requires judicious discernment. An important
requirement in preparing deuterated internal MS standards is a
very high level of deuterium incorporation at the positions of
enrichment within the standard, so that it does not cause
interference with the analyte. There are several commercially
available prediction software packages available, any of which
enable the user to find the sum formula of a compound to
calculate the necessary added mass units to generate a suitable
MS standard.
Several deuterated internal MS standards have been used
and prepared to quantify new or repurposed drugs and their
metabolites for the treatment of COVID-19.
Repurposed Drugs. First, Habler et al. developed and
validated a two-dimensional isotope-dilution liquid chroma-
tography tandem mass spectrometry (ID-LC-MS/MS) method
for the accurate and simultaneous quantification of drugs in
human serum, specifically for quantifying several repurposed
COVID-19 drugs simultaneously.
70
The work was performed
using stable deuterium-labeled analogues chloroquine-d4
phosphate (47), hydroxychloroquine-d4sulfate (48), ritona-
vir-d6(49), lopinavir-d8(50), and azithromycin-13C-d3(51) as
internal MS standards (Figure 7), all commercially sourced.
The dosage of repurposed COVID-19 therapeutics is typically
derived from in vitro-generated half-maximum eective
concentration (EC50) values for SARS-CoV-2 and pharmacol-
ogy-based pharmacokinetic models from other diseases and
clinical conditions. Because dosage regimens of repurposed
drugs cannot always be suitably translated from their original
purpose into appropriate drug exposure in COVID-19 patients
due to pathophysiological alterations, it can lead to possible
subtherapeutic or toxic concentrations without clinical benefit.
In addition, polytherapy can result in unreliable drug levels due
to interactions between dierent drugs. Therefore, therapeutic
drug monitoring is crucial. The developed assay was designed
to be an ecient method for the monitoring of these potential
drug candidates in COVID-19 patients and to increase
treatment ecacy and safety.
70
In a similar study, Sok et al.
71
developed and validated the
first ever LC-MS/MS method for simultaneous quantification
of azithromycin, hydroxychloroquine (HCQ) (both antima-
larial drugs and potential COVID-19 therapeutic agents), and
two metabolites of HCQ, desethyl-HCQ and bisdesethyl-
HCQ, in EDTA-treated human blood plasma. The study made
use of the commercially available internal MS standards
azithromycin-d5(52), hydroxychloroquine-d4(53) (the
neutralized version of 48), desethyl-hydroxychloroquine-d4
(54), and bisdesethylchloroquine-d4(55) (Figure 8), and is
suitable for clinical studies requiring a fast turnaround time and
small sample volume (the assay requires only 20 μL of
plasma). The method was developed to support clinical trials
and to assess the pharmacokinetics and pharmacodynamics of
these repurposed drugs in this new role.
71
The FDA approval of the antiviral drug remdesivir for the
treatment of COVID-19 in adult and pediatric patients 12
years or older requiring hospitalization led to the increased
need for a simple, sensitive, and selective assay to quantify drug
concentrations in clinical samples to study therapeutic dosing
and provide pharmacokinetic studies. Therefore, Nguyen et
Figure 8. Chemical structures of internal MS standards 52−56.
Figure 9. Chemical structures of ebselen (58) and its analogue (major metabolite of BS1801) 57, along with 59 (M2) and its deuterated analogue
60 (M2-d6).
ACS Omega http://pubs.acs.org/journal/acsodf Review
https://doi.org/10.1021/acsomega.2c04160
ACS Omega 2022, 7, 41840−41858
41850
al.
72
developed and validated a rapid and sensitive LC-MS/MS
assay for the quantification of remdesivir (compound 12) in
human plasma which made use of the commercially available
deuterium-labeled analog remdesivir-d5(56) (Figure 8) as the
internal MS standard. This method has proven to be the most
sensitive to date and is suitable for therapeutic dosing studies.
The precision, accuracy, and selectivity all met the FDA
Bioanalytical Guidelines.
Tian et al.
73a
reported the first-ever study carried out to
identify and quantify the major circulating metabolites of
BS1801 (57), an analogue of ebselen (58), found in the
hepatocytes of dierent species and human plasma (Figure 9).
Ebselen is one of the promising drugs for the treatment of
COVID-19.
74
To quantify the major BS1801 metabolite “M2”
(59), an accurate and fast LC-MS/MS method was established
which relied upon the use of a deuterated internal MS
standard.
To meet this need, the authors prepared a deuterated
internal MS standard (M2-d6)
73b
(60) by using a Grignard
reagent to introduce a deuterated methyl group on the
selenium atoms of 59 (Scheme 10). It was assumed that the
final product had >99% D incorporation since this was the
enrichment of the commercially obtained methyl-d3−magne-
sium iodide (CD3MgI) solution. The concept demonstrates
the utility of late-stage deuteration. The analytical method was
successfully used for the pharmacokinetic evaluation of BS1801
(57), demonstrating that this approach could provide a
reference for the pharmacokinetic analysis of other selenium-
containing drugs. The high D content of 60 is essential for the
use of deuterated internal standards in LC-MS/MS analysis.
Nevertheless, a greener alternative to traditional Grignard
solvents might be implemented for the synthesis of 60 in the
future.
75
Jansen-van Vuuren and Vohra
76
reported the development
of a simple synthetic route to baricitinib-d5(61), the
deuterated analog of baricitinib (62) (Figure 10). Baricitinib
is a therapeutic agent used to treat rheumatoid arthritis and, as
of May 11, 2022, approved by the FDA to treat COVID-19 in
hospitalized adults requiring supplemental oxygen or ven-
tilation.
77
Prior literature describing synthetic pathways to 61
involved the use of toxic reagents, so the authors developed an
alternative synthetic route, contingent on the initial integration
of deuterium into the ethanesulfonyl component through the
synthesis of ethanesulfonyl chloride-d5(63) from commercially
available ethanethiol-d5(>98% D) (Scheme 11). 63 was
immediately converted to the stable intermediate 64 in 94%
yield upon reaction with an easily prepared azetidinium salt. 64
could then be converted to the desired product (61) in an
additional two steps: reaction of 64 with commercially
available 65 to form stable and isolable intermediate 66,
followed by trimethylsilylethoxymethyl (SEM) deprotection.
The deuterated analog of baricitinib (61, 98% D), obtained in
an overall yield of 29%, may be used as an internal MS
standard in further studies.
Deuterated Biomarkers as Internal MS Standards.
Chhabra et al.
78
in preparing heparin/heparan sulfate (HS)
mimetics potentially targeted at a variety of diseases (including
SARS-CoV-2), developed a fully quantitative LC-MS/MS
assay for quantification of HS, also a biomarker for some
lysosomal storage diseases, namely, the family of mucopoly-
saccharidosis (MPS) disorders.
The disease causes the accumulation of undegraded HS
which is associated with multiple pathologies in the brain and
other organs. The published method proved to be ecient for
the determination of HS in the brain tissue of mice with MPS.
Given that quantification of HS in biological samples, e.g.,
urine or tissue, is complicated by its heterogeneity and high
molecular weight, acid-based methanolysis or butanolysis
resulting in desulfated disaccharide cleavage products which
are detectable by a LC-MS/MS assay is a suitable alternative
option. For this reason, the authors prepared a deuterium-
labeled version (67) of the major HS disaccharide butanolysis
product as an internal MS standard (Scheme 12). The
synthesis involves the initial saponification of 68 with NaOH
(aq) in a MeOH−chloroform solution.
79
The crude carbox-
ylate is then esterified under basic conditions with
commercially available 1-iodobutane-d9(≥98 atom % D) to
give the disaccharide 69 in 86% yield. Hydrogenolysis then
gave the deuterated disaccharide 67 in 57% yield (with the
same % D as the 1-iodobutane-d9). The method may also
prove useful in the study of HS mimetics intended for
pharmaceutical purposes, including for drugs targeting
COVID-19.
78,80
Deuterated Lipids as Internal MS Standards. Selected
bioactive lipids (BALs) and lipid mediators can initiate anti-
inflammatory activity, including during acute lung inflamma-
tion and injury. As such, BALs are pharmaceutical targets in
many inflammatory diseases, while higher levels of certain
BALs might signal cases of severe COVID-19. Archambault et
al.
81
used commercially available deuterium-labeled lipids and
lipid mediators (five examples (70−74) shown in Figure 11) as
internal/surrogate standards in the LC-MS/MS quantification
of certain BALs (eicosanoids and docosanoids) which
Scheme 10. Synthesis of 60 from 57 via Grignard Chemistry
a
a
Adapted with permission from ref 73. Copyright 2022 Elsevier.
Figure 10. Chemical structures of baricitinib-d5(61) and non-
deuterated baricitinib (62).
ACS Omega http://pubs.acs.org/journal/acsodf Review
https://doi.org/10.1021/acsomega.2c04160
ACS Omega 2022, 7, 41840−41858
41851
modulate lung inflammation in severe COVID-19 patients.
The goal was to find out if severe COVID-19 patients were
characterized by increased BALs modulating lung inflamma-
tion. A targeted lipidomic analysis of bronchoalveolar lavages
by tandem mass spectrometry on 25 healthy controls and 33
COVID-19 patients requiring mechanical ventilation was
performed; indeed, an increase in fatty acids and inflammatory
lipid mediators was observed. In the BALs of severe COVID-
19 patients, a predominance of eicosanoids such as
thromboxane B2 and prostaglandins was observed, which
were quantified with the use of deuterated internal MS
standards 70 (thromboxane B2-d4) and 71 (prostaglandin E2-
d4) (Figure 11). An increase was also observed in D-series
resolvins (pro-resolving mediators) and leukotrienes where
deuterated internal MS standards 72 (resolvin D2-d5), 73
(leukotriene C4-d5), and 74 (leukotriene B4-d4) came into use.
Similarly, Barberis et al.
82
used untargeted metabolomics
and lipidomics analysis of plasma from COVID-19 patients
Scheme 11. Synthesis of Deuterated Baricitinib (61) in a 29% Overall Yield, Starting from Commercially Available
Ethanethiol-d5
a
a
Adapted with permission from ref 76. Copyright 2022 John Wiley & Sons, Inc.
Scheme 12. Synthesis of Deuterated Dissacharide 67
a
a
Reagents and conditions: (a) (i) 5 M NaOH/MeOH/CHCl3/H2O, rt, 48 h; (ii) n-BuI-d9, KHCO3, DMF, rt, 24 h; (b) 20% Pd(OH)2/C, MeOH,
rt, 24 h. Adapted with permission from ref 79. Copyright 2019 American Chemical Society.
Figure 11. Chemical structures of commercially available deuterium-labeled lipids and lipid mediators 70−74.
ACS Omega http://pubs.acs.org/journal/acsodf Review
https://doi.org/10.1021/acsomega.2c04160
ACS Omega 2022, 7, 41840−41858
41852
and control groups to capture the host response to SARS-CoV-
2 infection. A deuterated standard mix (Splash Lipidomix;
https://avantilipids.com/product/330707) was used for LC-
MS/MS detection of dierent lipid classes which act as
potential COVID-19 biomarkers and therapeutic targets in
plasma. It was found that several circulating lipids,
triglycerides, and free fatty acids are correlated to the severity
of the disease. The study also provided further evidence for
considering phospholipase A2 (PLA2) activity as a potential
factor in the pathogenesis of COVID-19 and a possible
therapeutic target. A similar study
83
focuses on patients from
the Campania region, Italy.
This knowledge has the potential to assist in developing or
repurposing drugs which could be helpful at modulating the
observed lipidome to minimize the eects of pro-inflammatory
Table 2. List of Deuterated Drugs and Internal Standards Featured in This Review
code no. name of drug role played by deuterium ref
deuterated drugs as therapeutics: repurposed drugs
3deupirfenidone (LYT-100) inhibits the metabolism of the drug and enables less frequent dosing Schmidt 2021
27
Liu and Dong
2012
30
Chen et al.
2021
29
Chen et al.
2022
28
15 VV116 inhibits enzymatic degradation of the ring Xie et al.
2021
38
Qian et al.
2022
39
16, 20 GS-441524 deuterated analogues might improve antiviral activity of GS-441524 against SARS-CoV-2 Zheng et al.
2022
40
22 ACH-3422 improves the safety profile of the parent drug by enabling a more stable drug concentration and
reducing the production of toxic metabolites Gane et al.
2015
41
Tian et al.
2021
35
33 dexamethasone-d2might improve bioavailability and safety profile (hindered metabolism) Darshana et al.
2021
49
deuterated drugs as therapeutics: new drugs
35 arachidonic acid-d6decreases the overall rate of oxidation Molchanova et
al. 2022
51
Smarun et al.
2017
52
39, 40 GC376 deuterated analogues enhances activity due to tighter binding to the target or improves physicochemical properties of
the drug Dampalla et al.
2021
55−57
45 Y180 deuterium-enabled chiral switch Quan et al.
2022
61
deuterated drugs as internal MS standards: repurposed drugs
47−51 chloroquine-d4phosphate enables ID-LC-MS/MS quantification of repurposed COVID-19 drugs in human serum Habler et al.
2021
70
hydroxychloroquine-d4sulfate
ritonavir-d6
lopinavir-d8
azithromycin-13C-d3
52−55 azithromycin-d5enables LC-MS/MS quantification of repurposed drugs in EDTA-treated human blood plasma
to support clinical trials and assess the pharmacokinetics and pharmacodynamics of this
repurposed drug
Sok et al.
2021
71
hydroxychloroquine-d4
desethyl-hydroxychloroquine-d4
bisdesethylhydroxychloroquine-d4
56 remdesivir-d5enables LC-MS/MS quantification of remdesivir in human plasma Nguyen et al.
2021
72
60 M2-d6enables LC-MS/MS quantification of the major BS1801 metabolite “M2” Tian et al.
2022
73
61 baricitinib-d5could enable LC-MS/MS quantification of baricitinib Jansen-van
Vuuren et al.
2022
76
deuterated drugs as internal MS standards: biomarkers and lipids
67 deuterium-labeled HS disaccharide enables LC-MS/MS quantification of HS Ferro et al.
78−80
70−74 thromboxane B2-d4enables LC-MS/MS quantification of certain bioactive lipids Archambault et
al. 2021
81
prostaglandin E2-d4
resolvin D2-d5
leukotriene C4-d5
leukotriene B4-d4
ACS Omega http://pubs.acs.org/journal/acsodf Review
https://doi.org/10.1021/acsomega.2c04160
ACS Omega 2022, 7, 41840−41858
41853
lipids and enhance the eect of anti-inflammatory or pro-
resolving lipid mediators.
■CONCLUSION AND FUTURE OPPORTUNITIES
In conclusion, this review examines deuterated drugs which
have been featured during the COVID-19 pandemic, either as
potential therapeutic compounds or as internal MS standards
for studying the pharmacokinetic or metabolic properties of
new or repurposed COVID-19 drugs. Table 2 provides a
summary of all deuterated compounds included in this review.
For practitioners of HIE, there are many avenues worth
exploration.
First, the deuteration of repurposed or novel drugs being
studied as therapy options for COVID-19 whose safety profile
and/or bioavailability is poor or not yet be fully understood
may be of interest. For example, although remdesivir has been
approved for COVID-19 treatment, it can cause certain
adverse side eects;
84
thus, research into the safety profile of
remdesivir using deuterated analogues (e.g., remdesivir-d5,56)
could be of value.
Similarly, the synthesis of deuterated analogues of orally
administered COVID-19 antiviral agents which are being
considered for clinical trials, e.g., acriflavine,
85
or which have
advanced to late-stage trials, e.g., molnupiravir
(NCT04405570)
86
and Pfizer’s PF-07321332 (or paxlovid)
(NCT05011513),
87
would be useful as analytical standards
and/or for a deeper understanding of the metabolic profiles of
the nondeuterated versions.
Developing new synthetic methods to the drugs listed in
Table 2 which involve mid- or late-stage deuteration is a
welcome contribution to the field of HIE since this could
provide a greener synthetic route by decreasing the number of
steps and chemicals/resources needed.
Deuteration has been shown to stabilize drug enantiomers
and epimers. This approach holds much potential for enabling
the synthesis of pure enantiomers over racemic mixtures.
However, there has been limited exploration in this area
beyond basic research.
Arachidonic acid ethyl ester (34) metabolites are important
mediators in many physiological and pathophysiological
processes. In fact, many of the benefits and toxicities of both
glucocorticoids and nonsteroidal anti-inflammatory drugs are
due to blocking the production of beneficial and detrimental
metabolites of 34. Altering the metabolism of 34 via
deuteration at specific points in the chemical structure could
have a wide number of eects above and beyond what is
presented in the section Deuterated Arachidonic Acid Ethyl
Ester.
Overall, we hope that providing this reference tool and
highlighting new and interesting avenues for deuteration is of
value to isotope and medicinal chemists. We also anticipate
that exposing dierent strategies for drug development and
discovery would be beneficial in light of future global pandemic
situations.
■AUTHOR INFORMATION
Corresponding Author
Ross D. Jansen-van Vuuren −Faculty of Chemistry and
Chemical Technology, University of Ljubljana, Ljubljana
1000, Slovenia; Department of Chemistry, Queen’s
University, Kingston, Ontario K7L 3N6, Canada;
orcid.org/0000-0002-2919-6962;
Email: rossvanvuuren@gmail.com
Authors
Luka Jedlovc
nik −Faculty of Chemistry and Chemical
Technology, University of Ljubljana, Ljubljana 1000,
Slovenia
Janez Kos
mrlj −Faculty of Chemistry and Chemical
Technology, University of Ljubljana, Ljubljana 1000,
Slovenia; orcid.org/0000-0002-3533-0419
Thomas E. Massey −Department of Biomedical and
Molecular Sciences, School of Medicine, Queen’s University,
Kingston, Ontario K7L 3N6, Canada
Volker Derdau −Research &Development, Integrated Drug
Discovery, Isotope Chemistry, Sanofi-Aventis Deutschland
GmbH, Frankfurt/Main 65926, Germany; orcid.org/
0000-0002-3767-643X
Complete contact information is available at:
https://pubs.acs.org/10.1021/acsomega.2c04160
Notes
The authors declare the following competing financial
interest(s): V.D. is an employee of Sanofi and may hold
shares or options in the company. The other authors declare
no competing financial interest.
■ACKNOWLEDGMENTS
This project received funding from Queen’s University (Rapid
Response funding awarded to help confront COVID-19), The
Southeastern Ontario Academic Medical Organization
(SEAMO), and the European Union’s Horizon 2020 Research
and Innovation Programme under the Marie Skłodowska-
Curie grant agreement no. 945380. Financial support from the
Slovenian Research Agency (Research Core Funding grant P1-
0230) is also gratefully acknowledged.
■REFERENCES
(1) (a) Gordon, D. E.; Jang, G. M.; Bouhaddou, M.; Xu, J.;
Obernier, K.; White, K. M.; O’Meara, M. J.; Rezelj, V. V.; Guo, J. Z.;
Swaney, D. L.; Tummino, T. A.; Huttenhain, R.; Kaake, R. M.;
Richards, A. L.; Tutuncuoglu, B.; Foussard, H.; Batra, J.; Haas, K.;
Modak, M.; Kim, M.; Haas, P.; Polacco, B. J.; Braberg, H.; Fabius, J.
M.; Eckhardt, M.; Soucheray, M.; Bennett, M. J.; Cakir, M.;
McGregor, M. J.; Li, Q.; Meyer, B.; Roesch, F.; Vallet, T.; Mac
Kain, A.; Miorin, L.; Moreno, E.; Naing, Z. Z. C.; Zhou, Y.; Peng, S.;
Shi, Y.; Zhang, Z.; Shen, W.; Kirby, I. T.; Melnyk, J. E.; Chorba, J. S.;
Lou, K.; Dai, S. A.; Barrio-Hernandez, I.; Memon, D.; Hernandez-
Armenta, C.; Lyu, J.; Mathy, C. J. P.; Perica, T.; Pilla, K. B.; Ganesan,
S. J.; Saltzberg, D. J.; Rakesh, R.; Liu, X.; Rosenthal, S. B.; Calviello,
L.; Venkataramanan, S.; Liboy-Lugo, J.; Lin, Y.; Huang, X.-P.; Liu, Y.;
Wankowicz, S. A.; Bohn, M.; Safari, M.; Ugur, F. S.; Koh, C.; Savar, N.
S.; Tran, Q. D.; Shengjuler, D.; Fletcher, S. J.; O’Neal, M. C.; Cai, Y.;
Chang, J. C. J.; Broadhurst, D. J.; Klippsten, S.; Sharp, P. P.; Wenzell,
N. A.; Kuzuoglu-Ozturk, D.; Wang, H.-Y.; Trenker, R.; Young, J. M.;
Cavero, D. A.; Hiatt, J.; Roth, T. L.; Rathore, U.; Subramanian, A.;
Noack, J.; Hubert, M.; Stroud, R. M.; Frankel, A. D.; Rosenberg, O.
S.; Verba, K. A.; Agard, D. A.; Ott, M.; Emerman, M.; Jura, N.; von
Zastrow, M.; Verdin, E.; Ashworth, A.; Schwartz, O.; d’Enfert, C.;
Mukherjee, S.; Jacobson, M.; Malik, H. S.; Fujimori, D. G.; Ideker, T.;
Craik, C. S.; Floor, S. N.; Fraser, J. S.; Gross, J. D.; Sali, A.; Roth, B.
L.; Ruggero, D.; Taunton, J.; Kortemme, T.; Beltrao, P.; Vignuzzi, M.;
Garcia-Sastre, A.; Shokat, K. M.; Shoichet, B. K.; Krogan, N. J. A
SARS-CoV-2 protein interaction map reveals targets for drug
repurposing. Nature 2020,583 (7816), 459−468. (b) Guy, R. K.;
DiPaola, R. S.; Romanelli, F.; Dutch, R. E. Rapid repurposing of drugs
for COVID-19. Science 2020,368 (6493), 829−830. (c) El Bairi, K.;
Trapani, D.; Petrillo, A.; Le Page, C.; Zbakh, H.; Daniele, B.;
ACS Omega http://pubs.acs.org/journal/acsodf Review
https://doi.org/10.1021/acsomega.2c04160
ACS Omega 2022, 7, 41840−41858
41854
Belbaraka, R.; Curigliano, G.; Afqir, S. Repurposing anticancer drugs
for the management of COVID-19. Eur. J. Cancer 2020,141, 40−61.
(2) Sternberg, A.; McKee, D. L.; Naujokat, C. Novel drugs targeting
the SARS-CoV-2/COVID-19 machinery. Curr. Top. Med. Chem.
2020,20 (16), 1423−1433.
(3) V’Kovski, P.; Kratzel, A.; Steiner, S.; Stalder, H.; Thiel, V.
Coronavirus biology and replication: Implications for SARS-CoV-2.
Nat. Rev. Microbiol. 2021,19 (3), 155−170.
(4) Zhang, W.; Zhao, Y.; Zhang, F.; Wang, Q.; Li, T.; Liu, Z.; Wang,
J.; Qin, Y.; Zhang, X.; Yan, X.; Zeng, X.; Zhang, S. The use of anti-
inflammatory drugs in the treatment of people with severe
coronavirus disease 2019 (COVID-19): The perspectives of clinical
immunologists from China. Clin. Immunol. 2020,214, 108393.
(5) Know Your Treatment Options for COVID-19; https://www.
fda.gov/consumers/consumer-updates/know-your-treatment-options-
covid-19 (accessed 2022−08−25).
(6) (a) Ledford, H. Hundreds of COVID trials could provide a
deluge of new drugs. Nature 2022,603 (7899), 25−27. (b) Rahmah,
L.; Abarikwu, S. O.; Arero, A. G.; Essouma, M.; Jibril, A. T.; Fal, A.;
Flisiak, R.; Makuku, R.; Marquez, L.; Mohamed, K.; Ndow, L.;
Zarebska-Michaluk, D.; Rezaei, N.; Rzymski, P. Oral antiviral
treatments for COVID-19: Opportunities and challenges. Pharmacol.
Rep. 2022, 1−24. (c) Chavda, V. P.; Kapadia, C.; Soni, S.; Prajapati,
R.; Chauhan, S. C.; Yallapu, M. M.; Apostolopoulos, V. A global
picture: Therapeutic perspectives for COVID-19. Immunotherapy
2022,14 (5), 351−371. (d) Ho, W. S.; Zhang, R.; Tan, Y. L.; Chai, C.
L. L. COVID-19 and the promise of small molecule therapeutics: Are
there lessons to be learnt? Pharmacol. Res. 2022,179, 106201.
(7) Russak, E. M.; Bednarczyk, E. M. Impact of deuterium
substitution on the pharmacokinetics of pharmaceuticals. Ann.
Pharmacother. 2019,53 (2), 211−216.
(8) Schmidt, C. First deuterated drug approved. Nat. Biotechnol.
2017,35 (6), 493−494.
(9) (a) Kopf, S.; Bourriquen, F.; Li, W.; Neumann, H.; Junge, K.;
Beller, M. Recent developments for the deuterium and tritium
labeling of organic molecules. Chem. Rev. 2022,122 (6), 6634−6718.
(b) Li, N.; Li, Y.; Wu, X.; Zhu, C.; Xie, J. Radical deuteration. Chem.
Soc. Rev. 2022,51 (15), 6291−6306. (c) Ou, W.; Qiu, C.; Su, C.
Photo- and electro-catalytic deuteration of feedstock chemicals and
pharmaceuticals: A review. Chin. J. Catal. 2022,43 (4), 956−970.
(d) Atzrodt, J.; Derdau, V.; Fey, T.; Zimmermann, J. The renaissance
of H/D exchange. Angew. Chem., Int. Ed. 2007,46 (41), 7744−7765.
(e) Zhou, R.; Ma, L.; Yang, X.; Cao, J. Recent advances in visible-light
photocatalytic deuteration reactions. Org. Chem. Front. 2021,8(3),
426−444. (f) Rowbotham, J. S.; Ramirez, M. A.; Lenz, O.; Reeve, H.
A.; Vincent, K. A. Bringing biocatalytic deuteration into the toolbox of
asymmetric isotopic labelling techniques. Nat. Commun. 2020,11 (1),
1454.
(10) Atzrodt, J.; Derdau, V.; Kerr, W. J.; Reid, M. Deuterium- and
tritium-labelled compounds: Applications in the life sciences. Angew.
Chem., Int. Ed. 2018,57 (7), 1758−1784.
(11) Pirali, T.; Serafini, M.; Cargnin, S.; Genazzani, A. A.
Applications of deuterium in medicinal chemistry. J. Med. Chem.
2019,62 (11), 5276−5297.
(12) Harbeson, S. L.; Tung, R. D.; Macor, J. E. Deuterium in drug
discovery and development. Annu. Rep. Med. Chem. 2011,46, 403−
417.
(13) Zhan, Z.; Peng, X.; Sun, Y.; Ai, J.; Duan, W. Evaluation of
deuterium-labeled JNJ38877605: Pharmacokinetic, metabolic, and in
vivo antitumor profiles. Chem. Res. Toxicol. 2018,31 (11), 1213−
1218.
(14) (a) Sun, H.; Piotrowski, D. W.; Orr, S. T. M.; Warmus, J. S.;
Wolford, A. C.; Coffey, S. B.; Futatsugi, K.; Zhang, Y.; Vaz, A. D. N.
Deuterium isotope effects in drug pharmacokinetics II: Substrate-
dependence of the reaction mechanism influences outcome for
cytochrome P450 cleared drugs. PloS One 2018,13 (11), e0206279.
(b) Gant, T. G. Using deuterium in drug discovery: Leaving the label
in the drug. J. Med. Chem. 2014,57 (9), 3595−3611. (c) Knutson, D.
E.; Kodali, R.; Divovic, B.; Treven, M.; Stephen, M. R.; Zahn, N. M.;
Dobricic, V.; Huber, A. T.; Meirelles, M. A.; Verma, R. S.; Wimmer,
L.; Witzigmann, C.; Arnold, L. A.; Chiou, L.-C.; Ernst, M.;
Mihovilovic, M. D.; Savic, M. M.; Sieghart, W.; Cook, J. M. Design
and synthesis of novel deuterated ligands functionally selective for the
γ-aminobutyric acid type a receptor (gabaar) α6 subtype with
improved metabolic stability and enhanced bioavailability. J. Med.
Chem. 2018,61 (6), 2422−2446.
(15) Aprile, S.; Colombo, G.; Serafini, M.; Di Paola, R.; Pisati, F.;
Bhela, I. P.; Cuzzocrea, S.; Grosa, G.; Pirali, T. An unexpected
deuterium-induced metabolic switch in doxophylline. ACS Med.
Chem. Lett. 2022,13 (8), 1278−1285.
(16) Turowski, M.; Yamakawa, N.; Meller, J.; Kimata, K.; Ikegami,
T.; Hosoya, K.; Tanaka, N.; Thornton, E. R. Deuterium isotope
effects on hydrophobic interactions: The importance of dispersion
interactions in the hydrophobic phase. J. Am. Chem. Soc. 2003,125
(45), 13836−13849.
(17) Howland, R. H. Deuterated drugs. J. Psychosoc. Nurs. Ment.
Health Serv. 2015,53 (9), 13−16.
(18) (a) Kanao, E.; Kubo, T.; Naito, T.; Matsumoto, T.; Sano, T.;
Yan, M.; Otsuka, K. Isotope effects on hydrogen bonding and ch/
cd−πinteraction. J. Phys. Chem. C 2018,122 (26), 15026−15032.
(b) Wade, D. Deuterium isotope effects on noncovalent interactions
between molecules. Chem.-Biol. Interact. 1999,117 (3), 191−217.
(19) Swiderek, K.; Paneth, P. Binding isotope effects. Chem. Rev.
2013,113 (10), 7851−7879.
(20) (a) Krzan, M.; Vianello, R.; Marsavelski, A.; Repic, M.; Zaksek,
M.; Kotnik, K.; Fijan, E.; Mavri, J. The quantum nature of drug-
receptor interactions: deuteration changes binding affinities for
histamine receptor ligands. PloS One 2016,11 (5), e0154002.
(b) Krzan, M.; Keuschler, J.; Mavri, J.; Vianello, R. Relevance of
hydrogen bonds for the histamine h2 receptor-ligand interactions: A
lesson from deuteration. Biomolecules 2020,10 (2), 196.
(21) Deb, S.; Arrighi, S. Potential effects of COVID-19 on
cytochrome P450-mediated drug metabolism and disposition in
infected patients. Eur. J. Drug Metab. Pharmacokinet. 2021,46 (2),
185−203.
(22) Javorac, D.; Grahovac, L.; Manic, L.; Stojilkovic, N.;
Anđelkovic, M.; Bulat, Z.; Đukic- Cosic, D.; Curcic, M.;
Djordjevic, A. B. An overview of the safety assessment of medicines
currently used in the COVID-19 disease treatment. Food Chem.
Toxicol. 2020,144, 111639.
(23) Long, B.; Brady, W. J.; Koyfman, A.; Gottlieb, M.
Cardiovascular complications in COVID-19. Am. J. Emerg. Med.
2020,38 (7), 1504−1507.
(24) Harbeson, S. L.; Tung, R. D. Deuterium medicinal chemistry: a
new approach to drug discovery and development. MedChem. News
2014,24 (2), 8−22.
(25) Mullard, A. FDA approves first deuterated drug. Nat. Rev. Drug
Discovery 2017,16 (5), 305.
(26) Austedo Dosage; https://www.drugs.com/dosage/austedo.
html (accessed 2022−08−25).
(27) Schmidt, C. COVID-19 long haulers. Nat. Biotechnol. 2021,39
(8), 908−913.
(28) Chen, M. C.; Korth, C. C.; Harnett, M. D.; Elenko, E.; Lickliter,
J. D. A randomized phase 1 evaluation of deupirfenidone, a novel
deuterium-containing drug candidate for interstitial lung disease and
other inflammatory and fibrotic diseases. Clin. Pharmacol. Drug Dev.
2022,11 (2), 220−234.
(29) Chen, M.; Korth, C.; Harnett, M.; Lickliter, J.; Elenko, E. Phase
1 demonstrates LYT-100 (deupirfenidone) is dose-proportional and
well-tolerated when given twice-daily over multiple ascending doses
(MAD) and shows a minor food effect (FE). Eur. Respir. J. 2021,58
(65), PA469.
(30) Liu, F. J.; Dong, Y. Deuterated pirfenidone.
US20090131485A1, 2012.
(31) (a) Yin, Q.; Chen, Y.; Zhou, M.; Jiang, X.; Wu, J.; Sun, Y.
Synthesis and photophysical properties of deuteration of pirfenidone.
Spectrochim. Acta A Mol. Biomol. 2018,204, 88−98. (b) Shah, P.;
Saks, S. Substituted n-aryl pyridinones. WO2015112701A1, 2015.
ACS Omega http://pubs.acs.org/journal/acsodf Review
https://doi.org/10.1021/acsomega.2c04160
ACS Omega 2022, 7, 41840−41858
41855
(c) Zhang, C.; Sommers, A. Substituted n-aryl pyridinones.
WO2012122165, 2012. (d) Gant, T. G.; Sarshar, S. Preparation of
substituted N-aryl pyridinones as fibrotic inhibitors. WO2008157786,
2008.
(32) Kuang, Y.; Cao, H.; Tang, H.; Chew, J.; Chen, W.; Shi, X.; Wu,
J. Visible light driven deuteration of formyl C−H and hydridic
C(sp3)−H bonds in feedstock chemicals and pharmaceutical
molecules. Chem. Sci. 2020,11 (33), 8912−8918.
(33) Hu, L.; Liu, X.; Liao, X. Nickel-catalyzed methylation of aryl
halides with deuterated methyl iodide. Angew. Chem., Int. Ed. 2016,55
(33), 9743−9747.
(34) Falb, E.; Ulanenko, K.; Tor, A.; Gottesfeld, R.; Weitman, M.;
Afri, M.; Gottlieb, H.; Hassner, A. A highly efficient Suzuki−Miyaura
methylation of pyridines leading to the drug pirfenidone and its CD3
version (SD-560). Green Chem. 2017,19 (21), 5046−5053.
(35) Tian, L.; Qiang, T.; Liang, C.; Ren, X.; Jia, M.; Zhang, J.; Li, J.;
Wan, M.; YuWen, X.; Li, H.; Cao, W.; Liu, H. RNA-dependent RNA
polymerase (RdRp) inhibitors: The current landscape and repurpos-
ing for the COVID-19 pandemic. Eur. J. Med. Chem. 2021,213,
113201.
(36) Kokic, G.; Hillen, H. S.; Tegunov, D.; Dienemann, C.; Seitz, F.;
Schmitzova, J.; Farnung, L.; Siewert, A.; Höbartner, C.; Cramer, P.
Mechanism of SARS-CoV-2 polymerase stalling by remdesivir. Nat.
Commun. 2021,12 (1), 279.
(37) (a) Good, S. S.; Westover, J.; Jung, K. H.; Zhou, X.-J.; Moussa,
A.; La Colla, P.; Collu, G.; Canard, B.; Sommadossi, J.-P. AT-527, a
double prodrug of a guanosine nucleotide analog, is a potent inhibitor
of SARS-CoV-2 in vitro and a promising oral antiviral for treatment of
COVID-19. Antimicrob. Agents Chemother. 2021,65 (4), e02479.
(b) Fischer, W. A.; Eron, J. J.; Holman, W.; Cohen, M. S.; Fang, L.;
Szewczyk, L. J.; Sheahan, T. P.; Baric, R.; Mollan, K. R.; Wolfe, C. R.;
Duke, E. R.; Azizad, M. M.; Borroto-Esoda, K.; Wohl, D. A.; Coombs,
R. W.; James Loftis, A.; Alabanza, P.; Lipansky, F.; Painter, W. P. A
phase 2a clinical trial of molnupiravir in patients with COVID-19
shows accelerated SARS-CoV-2 RNA clearance and elimination of
infectious virus. Sci. Transl. Med. 2022,14 (628), eabl7430.
(38) Xie, Y.; Yin, W.; Zhang, Y.; Shang, W.; Wang, Z.; Luan, X.;
Tian, G.; Aisa, H. A.; Xu, Y.; Xiao, G.; Li, J.; Jiang, H.; Zhang, S.;
Zhang, L.; Xu, H. E.; Shen, J. Design and development of an oral
remdesivir derivative VV116 against SARS-CoV-2. Cell Res. 2021,31
(11), 1212−1214.
(39) Qian, H.; Wang, Y.; Zhang, M.; Xie, Y.; Wu, Q.; Liang, L.; Cao,
Y.; Duan, H.; Tian, G.; Ma, J.; Zhang, Z.; Li, N.; Jia, J.; Zhang, J.; Aisa,
H. A.; Shen, J.; Yu, C.; Jiang, H.; Zhang, W.; Wang, Z.; Liu, G. Safety,
tolerability, and pharmacokinetics of VV116, an oral nucleoside
analog against SARS-CoV-2, in Chinese healthy subjects. Acta
Pharmacol. Sin. 2022,DOI: 10.1038/s41401-022-00895-6.
(40) Zheng, W.; Hu, T.; Zhang, Y.; Wei, D.; Xie, Y.; Shen, J.
Synthesis and anti-SARS-CoV-2 activity of deuterated GS-441524
analogs. Tetrahedron Lett. 2022,104, 154012.
(41) Gane, E.; Schwabe, C.; Stedman, C.; Weilert, F.; Stuart, K.;
Cheng, W.; Yang, J.; Robison, H.; Hui, J.; Lahey, J.; Sorensen, R.;
Apelian, D.; Hindes, R. LP27: ACH-3422, A novel nucleotide prodrug
inhibitor of HCV NS5B polymerase. J. Hepatol. 2015,62 (2), S277.
(42) Phadke, A.; Hashimoto, A.; Ely, R. J. Pyrimidine nucleoside
phosphoramidate. WO2016044243, 2016.
(43) Deshpande, M.; Wiles, J. A.; Hashimoto, A.; Phadke, A.
Deuterated nucleoside prodrugs useful for treating hcv.
WO2014169280A3, 2014.
(44) Noreen, S.; Maqbool, I.; Madni, A. Dexamethasone:
Therapeutic potential, risks, and future projection during COVID-
19 pandemic. Eur. J. Pharmacol. 2021,894, 173854.
(45) Best, R.; Nelson, S. M.; Walker, B. R. Dexamethasone and 11-
dehydrodexamethasone as tools to investigate the isozymes of 11
beta-hydroxysteroid dehydrogenase in vitro and in vivo. J. Endocrinol.
1997,153 (1), 41−48.
(46) Viguerie, N.; Picard, F.; Hul, G.; Roussel, B.; Barbe, P.;
Iacovoni, J. S.; Valle, C.; Langin, D.; Saris, W. H. Multiple effects of a
short-term dexamethasone treatment in human skeletal muscle and
adipose tissue. Physiol. Genomics 2012,44 (2), 141−151.
(47) Yang, R.; Yu, Y. Glucocorticoids are double-edged sword in the
treatment of covid-19 and cancers. Int. J. Biol. Sci. 2021,17 (6),
1530−1537.
(48) Tomlinson, E. S.; Lewis, D. F. V.; Maggs, J. L.; Kroemer, H. K.;
Park, B. K.; Back, D. J. In vitro metabolism of dexamethasone (DEX)
in human liver and kidney: The involvement of CYP3a4 and CYP17
(17,20 LYASE) and molecular modelling studies. Biochem. Pharmacol.
1997,54 (5), 605−611.
(49) Darshana, D.; Sureram, S.; Mahidol, C.; Ruchirawat, S.;
Kittakoop, P. Spontaneous conversion of prenyl halides to acids:
Application in metal-free preparation of deuterated compounds under
mild conditions. Org. Biomol. Chem. 2021,19 (34), 7390−7402.
(50) Shoieb, S. M.; El-Ghiaty, M. A.; El-Kadi, A. O. S. Targeting
arachidonic acid−related metabolites in COVID-19 patients:
Potential use of drug-loaded nanoparticles. Emergent Mater. 2021,4
(1), 265−277.
(51) Molchanova, A. Y.; Rjabceva, S. N.; Melik-Kasumov, T. B.;
Pestov, N. B.; Angelova, P. R.; Shmanai, V. V.; Sharko, O. L.; Bekish,
A. V.; James, G.; Park, H. G.; Udalova, I. A.; Brenna, J. T.;
Shchepinov, M. S. Deuterated arachidonic acid ameliorates lip-
opolysaccharide-induced lung damage in mice. Antioxidants 2022,11
(4), 681.
(52) Smarun, A. V.; Petkovic, M.; Shchepinov, M. S.; Vidovic, D.
Site-specific deuteration of polyunsaturated alkenes. J. Org. Chem.
2017,82 (24), 13115−13120.
(53) Wang, D. H.; Park, H. G.; Wang, Z.; Lacombe, R. J. S.;
Shmanai, V. V.; Bekish, A. V.; Schmidt, K.; Shchepinov, M. S.;
Brenna, J. T. Toward quantitative sequencing of deuteration of
unsaturated hydrocarbon chains in fatty acids. Anal. Chem. 2021,93
(23), 8238−8247.
(54) Andreyev, A. Y.; Tsui, H. S.; Milne, G. L.; Shmanai, V. V.;
Bekish, A. V.; Fomich, M. A.; Pham, M. N.; Nong, Y.; Murphy, A. N.;
Clarke, C. F.; Shchepinov, M. S. Isotope-reinforced polyunsaturated
fatty acids protect mitochondria from oxidative stress. Free Radic. Biol.
Med. 2015,82, 63−72.
(55) Dampalla, C. S.; Zheng, J.; Perera, K. D.; Wong, L.-Y. R.;
Meyerholz, D. K.; Nguyen, H. N.; Kashipathy, M. M.; Battaile, K. P.;
Lovell, S.; Kim, Y.; Perlman, S.; Groutas, W. C.; Chang, K.-O.
Postinfection treatment with a protease inhibitor increases survival of
mice with a fatal SARS-CoV-2 infection. Proc. Natl. Acad. Sci. U.S.A.
2021,118 (29), e2101555118.
(56) (a) Dampalla, C. S.; Kim, Y.; Bickmeier, N.; Rathnayake, A. D.;
Nguyen, H. N.; Zheng, J.; Kashipathy, M. M.; Baird, M. A.; Battaile,
K. P.; Lovell, S.; Perlman, S.; Chang, K.-O.; Groutas, W. C. Structure-
guided design of conformationally constrained cyclohexane inhibitors
of severe acute respiratory syndrome coronavirus-2 3cl protease. J.
Med. Chem. 2021,64 (14), 10047−10058. (b) Dampalla, C. S.;
Rathnayake, A. D.; Perera, K. D.; Jesri, A.-R. M.; Nguyen, H. N.;
Miller, M. J.; Thurman, H. A.; Zheng, J.; Kashipathy, M. M.; Battaile,
K. P.; Lovell, S.; Perlman, S.; Kim, Y.; Groutas, W. C.; Chang, K.-O.
Structure-guided design of potent inhibitors of sars-cov- 2 3cl
protease: Structural, biochemical, and cell-based studies. J. Med. Chem.
2021,64 (24), 17846−17865.
(57) Dampalla, C. S.; Rathnayake, A. D.; Galasiti Kankanamalage, A.
C.; Kim, Y.; Perera, K. D.; Nguyen, H. N.; Miller, M. J.; Madden, T.
K.; Picard, H. R.; Thurman, H. A.; Kashipathy, M. M.; Liu, L.;
Battaile, K. P.; Lovell, S.; Chang, K.-O.; Groutas, W. C. Structure-
guided design of potent spirocyclic inhibitors of severe acute
respiratory syndrome coronavirus-2 3c-like protease. J. Med. Chem.
2022,65 (11), 7818−7832.
(58) Szostak, M.; Spain, M.; Procter, D. J. Selective synthesis of α,α-
dideuterio alcohols by the reduction of carboxylic acids using smi2
and d2o as deuterium source under set conditions. Org. Lett. 2014,16
(19), 5052−5055.
(59) Hou, J. Paracellular channel as drug target. In The Paracellular
Channel, 1st ed.; Academic Press, 2019; pp 175−199.
ACS Omega http://pubs.acs.org/journal/acsodf Review
https://doi.org/10.1021/acsomega.2c04160
ACS Omega 2022, 7, 41840−41858
41856
(60) Citarella, A.; Scala, A.; Piperno, A.; Micale, N. SARS-CoV-2
Mpro: A potential target for peptidomimetics and small-molecule
inhibitors. Biomolecules 2021,11 (4), 607.
(61) Quan, B.-X.; Shuai, H.; Xia, A.-J.; Hou, Y.; Zeng, R.; Liu, X.-L.;
Lin, G.-F.; Qiao, J.-X.; Li, W.-P.; Wang, F.-L.; Wang, K.; Zhou, R.-J.;
Yuen, T. T.-T.; Chen, M.-X.; Yoon, C.; Wu, M.; Zhang, S.-Y.; Huang,
C.; Wang, Y.-F.; Yang, W.; Tian, C.; Li, W.-M.; Wei, Y.-Q.; Yuen, K.-
Y.; Chan, J. F.-W.; Lei, J.; Chu, H.; Yang, S. An orally available Mpro
inhibitor is effective against wild-type SARS-CoV-2 and variants
including Omicron. Nat. Microbiol. 2022,7(5), 716−725.
(62) Dong, J.; Wang, X.; Wang, Z.; Song, H.; Liu, Y.; Wang, Q.
Formyl-selective deuteration of aldehydes with D2O via synergistic
organic and photoredox catalysis. Chem. Sci. 2020,11 (4), 1026−
1031.
(63) Geng, H.; Chen, X.; Gui, J.; Zhang, Y.; Shen, Z.; Qian, P.;
Chen, J.; Zhang, S.; Wang, W. Practical synthesis of C1 deuterated
aldehydes enabled by NHC catalysis. Nat. Catal. 2019,2(12), 1071−
1077.
(64) (a) DeWitt, S.; Czarnik, A. W.; Jacques, V. Deuterium-enabled
chiral switching (DECS) yields chirally pure drugs from chemically
interconverting racemates. ACS Med. Chem. Lett. 2020,11 (10),
1789−1792. (b) Maltais, F.; Jung, Y. C.; Chen, M.; Tanoury, J.; Perni,
R. B.; Mani, N.; Laitinen, L.; Huang, H.; Liao, S.; Gao, H.; Tsao, H.;
Block, E.; Ma, C.; Shawgo, R. S.; Town, C.; Brummel, C. L.; Howe,
D.; Pazhanisamy, S.; Raybuck, S.; Namchuk, M.; Bennani, Y. L. In
vitro and in vivo isotope effects with hepatitis c protease inhibitors:
Enhanced plasma exposure of deuterated telaprevir versus telaprevir
in rats. J. Med. Chem. 2009,52 (24), 7993−8001.
(65) Tan, A.; Awaiye, K. Use of internal standards in LC-MS
bioanalysis. In Handbook of LC-MS Bioanalysis; Li, W., Zhang, J., Tse,
F. L. S., Eds.; John Wiley & Sons, 2013; pp 217−227. DOI: 10.1002/
9781118671276.ch17.
(66) Higashi, T.; Ogawa, S. Isotope-coded ESI-enhancing
derivatization reagents for differential analysis, quantification and
profiling of metabolites in biological samples by LC/MS: A review. J.
Pharm. Biomed. 2016,130, 181−193.
(67) Stokvis, E.; Rosing, H.; Beijnen, J. H. Stable isotopically labeled
internal standards in quantitative bioanalysis using liquid chromatog-
raphy/mass spectrometry: Necessity or not? Rapid Commun. Mass
Spectrom. 2005,19 (3), 401−407.
(68) (a) Wu, D.-C.; Bai, J.-W.; Guo, L.; Hu, G.-Q.; Liu, K.-H.;
Sheng, F.-F.; Zhang, H.-H.; Sun, Z.-Y.; Shen, K.; Liu, X. A practical
and efficient method for late-stage deuteration of terminal alkynes
with silver salt as catalyst. Tetrahedron Lett. 2021,66, 152807.
(b) Uttry, A.; Mal, S.; van Gemmeren, M. Late-stage β-C(sp3)−H
deuteration of carboxylic acids. J. Am. Chem. Soc. 2021,143 (29),
10895−10901. (c) Levernier, E.; Tatoueix, K.; Garcia-Argote, S.;
Pfeifer, V.; Kiesling, R.; Gravel, E.; Feuillastre, S.; Pieters, G. Easy-to-
implement hydrogen isotope exchange for the labeling of n-
heterocycles, alkylkamines, benzylic scaffolds, and pharmaceuticals.
JACS Au 2022,2(4), 801−808. (d) Farizyan, M.; Mondal, A.; Mal,
S.; Deufel, F.; van Gemmeren, M. Palladium-catalyzed nondirected
late-stage c−h deuteration of arenes. J. Am. Chem. Soc. 2021,143
(40), 16370−16376. (e) Kallepalli, V. A.; Gore, K. A.; Shi, F.;
Sanchez, L.; Chotana, G. A.; Miller, S. L.; Maleczka, R. E.; Smith, M.
R. Harnessing C−H borylation/deborylation for selective deuteration,
synthesis of boronate esters, and late stage functionalization. J. Org.
Chem. 2015,80 (16), 8341−8353.
(69) Rule, G. S.; Clark, Z. D.; Yue, B.; Rockwood, A. L. Correction
for Isotopic Interferences between analyte and internal standard in
quantitative mass spectrometry by a nonlinear calibration function.
Anal. Chem. 2013,85 (8), 3879−3885.
(70) Habler, K.; Brugel, M.; Teupser, D.; Liebchen, U.; Scharf, C.;
Schönermarck, U.; Vogeser, M.; Paal, M. Simultaneous quantification
of seven repurposed COVID-19 drugs remdesivir (plus metabolite
GS-441524), chloroquine, hydroxychloroquine, lopinavir, ritonavir,
favipiravir and azithromycin by a two-dimensional isotope dilution
LC−MS/MS method in human serum. J. Pharm. Biomed. 2021,196,
113935.
(71) Sok, V.; Marzan, F.; Gingrich, D.; Aweeka, F.; Huang, L.
Development and validation of an LC-MS/MS method for
determination of hydroxychloroquine, its two metabolites, and
azithromycin in EDTA-treated human plasma. PloSs One 2021,16
(3), e0247356.
(72) Nguyen, R.; Goodell, J. C.; Shankarappa, P. S.; Zimmerman, S.;
Yin, T.; Peer, C. J.; Figg, W. D. Development and validation of a
simple, selective, and sensitive LC-MS/MS assay for the quantifica-
tion of remdesivir in human plasma. J. Chromatogr. B Analyt. Technol.
Biomed. Life Sci. 2021,1171, 122641.
(73) (a) Tian, Q.; Jiang, J.; Yin, H.; Ma, J.; Deng, G.; Zhou, J.;
Zhong, D. Quantification of the major circulating metabolite of
BS1801, an ebselen analog, in human plasma. J. Pharm. Biomed. 2022,
212, 114638. (b) Personal communication with Professor Dafang
Zhong, Chinese Academy of Sciences, Shanghai.
(74) Sun, L.-Y.; Chen, C.; Su, J.; Li, J.-Q.; Jiang, Z.; Gao, H.; Chigan,
J.-Z.; Ding, H.-H.; Zhai, L.; Yang, K.-W. Ebsulfur and ebselen as
highly potent scaffolds for the development of potential SARS-CoV-2
antivirals. Bioorg. Chem. 2021,112, 104889.
(75) Kadam, A.; Nguyen, M.; Kopach, M.; Richardson, P.; Gallou,
F.; Wan, Z.-K.; Zhang, W. Comparative performance evaluation and
systematic screening of solvents in a range of Grignard reactions.
Green Chem. 2013,15 (7), 1880−1888.
(76) Jansen-van Vuuren, R. D.; Vohra, R. Synthesis of [2H5]-
baricitinib via [2H5]ethanesulfonyl chloride. J. Labelled Comp.
Radiopharm. 2022,65 (6), 156−161.
(77) Kumari, P.; Pradhan, B.; Koromina, M.; Patrinos, G. P.; Steen,
K. V. Discovery of new drug indications for COVID-19: A drug
repurposing approach. PloS One 2022,17 (5), e0267095.
(78) Chhabra, M.; Doherty, G. G.; See, N. W.; Gandhi, N. S.; Ferro,
V. From cancer to COVID-19: A perspective on targeting heparan
sulfate-protein interactions. Chem. Rec. 2021,21 (11), 3087−3101.
(79) He, Q. Q.; Trim, P. J.; Lau, A. A.; King, B. M.; Hopwood, J. J.;
Hemsley, K. M.; Snel, M. F.; Ferro, V. Synthetic disaccharide
standards enable quantitative analysis of stored heparan sulfate in
MPS IIIA murine brain regions. ACS Chem. Neurosci. 2019,10 (8),
3847−3858.
(80) Guimond, S. E.; Mycroft-West, C. J.; Gandhi, N. S.; Tree, J. A.;
Le, T. T.; Spalluto, C. M.; Humbert, M. V.; Buttigieg, K. R.;
Coombes, N.; Elmore, M. J.; Wand, M.; Nyström, K.; Said, J.; Setoh,
Y. X.; Amarilla, A. A.; Modhiran, N.; Sng, J. D. J.; Chhabra, M.;
Young, P. R.; Rawle, D. J.; Lima, M. A.; Yates, E. A.; Karlsson, R.;
Miller, R. L.; Chen, Y.-H.; Bagdonaite, I.; Yang, Z.; Stewart, J.;
Nguyen, D.; Laidlaw, S.; Hammond, E.; Dredge, K.; Wilkinson, T. M.
A.; Watterson, D.; Khromykh, A. A.; Suhrbier, A.; Carroll, M. W.;
Trybala, E.; Bergström, T.; Ferro, V.; Skidmore, M. A.; Turnbull, J. E.
Synthetic heparan sulfate mimetic pixatimod (PG545) potently
inhibits SARS-CoV-2 by disrupting the spike−ACE2 interaction.
ACS Cent. Sci. 2022,8(5), 527−545.
(81) Archambault, A.; Zaid, Y.; Rakotoarivelo, V.; Turcotte, C.;
Doré, E.; Dubuc, I.; Martin, C.; Flamand, O.; Amar, Y.; Cheikh, A.;
Fares, H.; El Hassani, A.; Tijani, Y.; Coté, A.; Laviolette, M.; Boilard,
E.; Flamand, L.; Flamand, N. High levels of eicosanoids and
docosanoids in the lungs of intubated COVID-19 patients. FASEB
J. 2021,35 (6), e21666.
(82) Barberis, E.; Timo, S.; Amede, E.; Vanella, V. V.; Puricelli, C.;
Cappellano, G.; Raineri, D.; Cittone, M. G.; Rizzi, E.; Pedrinelli, A. R.;
Vassia, V.; Casciaro, F. G.; Priora, S.; Nerici, I.; Galbiati, A.; Hayden,
E.; Falasca, M.; Vaschetto, R.; Sainaghi, P. P.; Dianzani, U.; Rolla, R.;
Chiocchetti, A.; Baldanzi, G.; Marengo, E.; Manfredi, M. Large-scale
plasma analysis revealed new mechanisms and molecules associated
with the host response to SARS-CoV-2. Int. J. Mol. Sci. 2020,21 (22),
8623.
(83) Ciccarelli, M.; Merciai, F.; Carrizzo, A.; Sommella, E.; Di
Pietro, P.; Caponigro, V.; Salviati, E.; Musella, S.; Sarno, V. di;
Rusciano, M.; Toni, A. L.; Iesu, P.; Izzo, C.; Schettino, G.; Conti, V.;
Venturini, E.; Vitale, C.; Scarpati, G.; Bonadies, D.; Rispoli, A.;
Polverino, B.; Poto, S.; Pagliano, P.; Piazza, O.; Licastro, D.;
Vecchione, C.; Campiglia, P. Untargeted lipidomics reveals specific
ACS Omega http://pubs.acs.org/journal/acsodf Review
https://doi.org/10.1021/acsomega.2c04160
ACS Omega 2022, 7, 41840−41858
41857
lipid profiles in COVID-19 patients with different severity from
Campania region (Italy). J. Pharm. Biomed. 2022,217, 114827.
(84) (a) Davies, M.; Osborne, V.; Lane, S.; Roy, D.; Dhanda, S.;
Evans, A.; Shakir, S. Remdesivir in treatment of COVID-19: A
systematic benefit−risk assessment. Drug Saf. 2020,43 (7), 645−656.
(b) Fan, Q.; Zhang, B.; Ma, J.; Zhang, S. Safety profile of the antiviral
drug remdesivir: An update. Biomed. Pharmacother. 2020,130,
110532. (c) Wong, C. K. H.; Au, I. C. H.; Cheng, W. Y.; Man, K.
K. C.; Lau, K. T. K.; Mak, L. Y.; Lui, S. L.; Chung, M. S. H.; Xiong, X.;
Lau, E. H. Y.; Cowling, B. J. Remdesivir use and risks of acute kidney
injury and acute liver injury among patients hospitalised with
COVID-19: A self-controlled case series study. Aliment. Pharmacol.
Ther. 2022,56 (1), 121−130.
(85) Napolitano, V.; Dabrowska, A.; Schorpp, K.; Mourao, A.;
Barreto-Duran, E.; Benedyk, M.; Botwina, P.; Brandner, S.; Bostock,
M.; Chykunova, Y.; Czarna, A.; Dubin, G.; Fröhlich, T.; Hölscher, M.;
Jedrysik, M.; Matsuda, A.; Owczarek, K.; Pachota, M.; Plettenburg,
O.; Potempa, J.; Rothenaigner, I.; Schlauderer, F.; Slysz, K.;
Szczepanski, A.; Greve-Isdahl Mohn, K.; Blomberg, B.; Sattler, M.;
Hadian, K.; Popowicz, G. M.; Pyrc, K. Acriflavine, a clinically
approved drug, inhibits SARS-CoV-2 and other betacoronaviruses.
Cell Chem. Biol. 2022,29 (5), 774−784.
(86) Whitley, R. Molnupiravir �A step toward orally bioavailable
therapies for Covid-19. N. Engl. J. Med. 2022,386 (6), 592−593.
(87) Heskin, J.; Pallett, S. J. C.; Mughal, N.; Davies, G. W.; Moore,
L. S. P.; Rayment, M.; Jones, R. Caution required with use of
ritonavir-boosted PF-07321332 in COVID-19 management. Lancet
2022,399 (10319), 21−22.
ACS Omega http://pubs.acs.org/journal/acsodf Review
https://doi.org/10.1021/acsomega.2c04160
ACS Omega 2022, 7, 41840−41858
41858
