New insights into mechanisms of therapeutic Daniel

REVIEWS
New insights into mechanisms of therapeutic
effects of antimalarial agents in SLE
Daniel J. Wallace, Vineet S. Gudsoorkar, Michael H. Weisman and Swamy R. Venuturupalli
Abstract | Antimalarial agents have routinely been used for the treatment of systemic lupus erythematosus
(SLE) for over 50 years. These agents continue to enjoy success as the initial pharmacotherapy for SLE even
in the era of targeted therapies. Antimalarial agents have numerous biological effects that are responsible
for their immunomodulatory actions in SLE. Their inhibitory effect on Toll-like receptor-mediated activation of
the innate immune response is perhaps the most important discovery regarding their putative mechanism
of action, but some other, previously known properties, such as antithrombotic and antilipidaemic effects,
are now explained by new research. In the 1980s and 1990s, these antihyperlipidaemic and antithrombotic
effects were demonstrated in retrospective clinical studies, and over the past few years prospective studies
have confirmed those findings. Knowledge about the risk–benefit profile of antimalarial agents during
pregnancy and lactation has evolved, as has the concept of retinal toxicity. Antimalarial agents have unique
disease-modifying properties in SLE and newer iterations of this class of anti-inflammatory agents will have
a profound effect upon the treatment of autoimmune disease.
Wallace, D. J. et al. Nat. Rev. Rheumatol. 8, 522–533 (2012); published online 17 July 2012; doi:10.1038/nrrheum.2012.106
Introduction
As medicinal agents, antimalarial agents have been
prescribed for over 500 years, and their potential uses
are still being explored. Literary references to quinine
—the first effective treatment for malaria—date back to
the 16th century, although Payne reported its evidencebased use for cutaneous lupus for the first time in 1894.1
Quinacrine (also marketed as mepacrine or atabrine) was
patented in 1930, and was used by four million US and
allied soldiers during World War II. Studies in this cohort
of subjects yielded observational evidence that quinacrine ameliorated inflammatory arthritis and rash. This
discovery led to further research that marked the transition of antimalarial agents to the treatment of rheumatic
diseases. Chloroquine was first synthesized in 1943 and
hydroxychloroquine came on the market in 1955.1 This
Review presents new insights based on recent advances
in our understanding of the mechanisms of action of
antimalarial agents.
Pharmacology of antimalarial agents
Division of
Rheumatology, Cedars–
Sinai Medical Center,
8700 Beverly
Boulevard, Becker
B‑131, Los Angeles,
CA 90048, USA
(D. J. Wallace,
V. S. Gudsoorkar,
M. H. Weisman,
S. R. Venturupalli).
Chloroquine and hydroxychloroquine are weakly basic
4‑aminoquinoline compounds. Hydroxychloroquine
differs from chloroquine by a hydroxyl group attached
to a side chain. Quinacrine is an acridine compound
that differs from chloroquine by the presence of an
extra benzene ring (Figure 1).2 The clinically important
aspects of pharmacology and metabolism of antimalarial
agents are summarized in Box 1.3,4
Correspondence to:
D. J. Wallace
[email protected]
Competing interests
The authors declare no competing interests.
522 | SEPTEMBER 2012 | VOLUME 8
In general, chloroquine is 2–3 times more potent
than hydroxychloroquine,5 and the risk of retinopathy
is lower with the latter.6 For these reasons, hydroxy­
chloroquine is preferred over chloroquine in rheumatology practice. Commercially available tablet preparations
of hydroxychloroquine sulfate contain 155 mg base
equivalent in a 200 mg tablet, and tablets of chloroquine phosphate are available as 500 mg (300 mg base
equivalent) and 250 mg (150 mg base equivalent) doses.
Chloroquine hydrochloride is available in some countries as 100 mg tablets containing 80 mg base equivalent. Quinacrine is not commercially manufactured
in the USA but can be ordered from pharmacies that
compound it.
Blood concentrations of hydroxychloroquine can independently predict exacerbation in patients with SLE;7
therefore, measurements of hydroxychloroquine levels in
whole blood could help to identify ‘at-risk’ patients and
optimize treatment. Owing to hydroxychloroquine’s long
elimination half-life, routine measurements of serum
hydroxychloroquine concentration can also serve as a
marker of treatment adherence.8 Although these tests are
not yet routinely ordered by rheumatologists in the USA,
many laboratories around the world perform these tests
using high-performance liquid chromatography.
Mechanisms of action
Antimalarial agents exert their effects via multiple
molecu­lar pathways. Some well-known effects are listed
in Box 2,9 and the principal modes of action with new
insights into their mechanisms are discussed below.
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Interference with lysosome function
Chloroquine and hydroxychloroquine are weak bases
that have an affinity for the acidic lysosome (‘lysosomo­
tropic’ action).10 These agents alter the endolysosomal
pH and interfere with acidification of lysosomes.
In T cells, lysosomes perform the dual function of
degrada­tion of endocytosed material and participation
in apoptosis of antigen-presenting cells (APCs). These
functions, if impaired, can affect the cellular immune
response and APC function, and lead to several downstream effects such as inhibition of cytokine production, especially IL‑1, IL‑ 6 and TNF. IL‑6 is an important
inflammatory medi­ator that stimulates B‑cell differentiation leading to subsequent antibody response, and is
considered an important pathogenic mediator in SLE.
Our group has demonstrated that hydroxychloroquine
exerts a long-lasting suppressive effect on levels of IL‑6
possibly of macrophage–monocyte origin as opposed
to lymphocytic origin.11 This effect has been confirmed
independently by other studies.12,13 The exact nature
and reason for a preferential effect of hydroxychloroquine on IL‑6 is not clear. Possibly, in addition to inhibition of antigen presentation, antimalarial agents also
downregulate mRNA expression of cytokines at the
t­ranscriptional level.14
TLR-associated mechanisms
Perhaps the most important advance in our understanding of antimalarial agents is knowledge of their
antagonistic effect on the nucleic-acid sensing Toll-like
receptors (TLRs). The presence of antibodies against
nucleic acids is a hallmark of many autoimmune diseases, including SLE. Ineffective clearance of apoptotic
cellular material exposes intracellular contents such
as nucleic acids to the immune system and invokes an
autoimmune response, as evident from animal models
of DNase-deficient mice being prone to developing
antinuclear antibodies and SLE-like disease,15 and from
the observation of reduced activity of serum DNase‑I in
patients with active SLE.16
TLRs are type‑I transmembrane receptors that form
the early defense mechanism against foreign organisms.
These receptors are germline coded during the evolution
of species and are geared to recognize specific molecular
patterns associated with pathogenic species. Upon exposure to a pathogen, TLRs activate more-specific pathways
of the acquired immune response. Nucleic acid-sensing
TLRs (TLR3, TLR7, TLR8 and TLR9) are located in
the intracellular compartments to minimize accidental
exposure to self-nucleic material; these TLRs are activated only when foreign nuclear material is presented to
them by specialized intermediate molecules that facilitate the delivery of antigens to the intra­cellular compartment, such as Fcγ receptors on dendritic cells or B‑cell
receptors on the surface of B cells. Activated TLRs, via
adaptor molecules such as MyD88 (for TLR7 and TLR9)
and TRIF (for TLR3), stimulate the production of type I
interferons and proinflammatory cytokines (Figure 2).17
Plasmacytoid dendritic cells (pDCs) have a unique
ability to couple the signaling pathways of TLR7 and
Key points
■■ Antimalarial agents are the cornerstone agents in the clinical management
of systemic lupus erythematosus
■■ Toll-like receptor (TLR)-antagonism has emerged as an important mechanism
of action of antimalarial agents
■■ The antilipidaemic, photoprotective and antiproliferative effects of chloroquine,
hydroxychloroquine and quinacrine are in part explained by TLR antagonism
■■ Antimalarial agents also act by several additional molecular mechanisms, the
understanding of which continues to evolve
■■ Antimalarial agents are generally safe, effective and clinically useful in almost
all patients with systemic lupus erythematosus
■■ These drugs offer considerable promise for treating a variety of
immune-mediated as well as nonimmune diseases, and have exciting potential
N
Cl
NH
H3C
N
H3C
NH
H3C
Chloroquine
H3C
N
Cl
Hydroxychloroquine
N
H3C
H3C
OH
N
H3C
Quinacrine
H3C
NH
O
Cl
CH3
N
Figure 1 | Chemical structures of antimalarial agents.2
TLR9, which leads to the production of large quantities
of type I interferons and substantially increases transcription of type I interferon genes.18 IFN‑α has a crucial
role in the pathogenesis of SLE. IFN‑α levels are high in
patients with SLE and IFN genes have been known to be
upregulated in this population, an effect referred to as the
‘interferon signature’ of SLE. 19 pDCs are considered
the main source of IFN‑α. 20 The novel concept of
NETosis (Box 3) hypothesizes that neutrophils might
play an equally important part in interferon secretion
by stimulating pDCs.21 Importantly, TLRs are central to
both these interferon pathways. In pDCs, TLR activation
increases IFN‑α synthesis, as described above. In the case
of neutro­phils, evidence suggests that not only is pDC
activation by neutrophil extracellular traps (NETs) mediated via TLRs, but also the inhibition of TLR9 reduces
activation of pDCs by NETs.21
Evidence that antimalarial agents act by blocking the activation of TLRs comes from many sources.
Unmethylated CpG motifs are seen in bacterial DNA
whereas vertebrate DNA contains more of the methy­
lated fraction, 22 and unmethylated motifs evoke a
stronger immune response than their methylated
counterparts.22 Antimalarial agents have been shown
in animal studies and in vitro models to antagonize
immune stimulation by CpG-DNA (a ligand for TLR9)
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Box 1 | Pharmacology and metabolism of antimalarial agents
Absorption
Chloroquine, hydroxychloroqine and quinacrine all have very good bioavailability
after oral administration. Quinacrine can be given intralesionally. The drugs are
distributed extensively in tissues; they have an affinity for pigmented tissues and
are not retained in adipose tissue.
Metabolism
Hepatic metabolism by cytochrome P450 enzymes is the primary mode of
metabolism and results in S‑enantiomers and R‑enantiomers. Of the two types,
the S‑enantiomer of hydroxychloroquine has a shorter half-life and lower blood
levels. 21–47% of the dose is excreted without being metabolized.
Excretion
40–50% of the dose is excreted renally, and small amounts are excreted in
faeces, through the skin, and in breast milk. Antimalarial agents have a long
elimination half-life (approximately 40 days) but are detected in tissues for
prolonged periods (up to 5 years).103
Box 2 | Well-known effects of antimalarial agents9
■■
■■
■■
■■
■■
■■
■■
■■
■■
■■
■■
Inhibit phospholipase A2 and phospholipase C
Stabilize lysosome membranes
Decrease production of oestrogen
Hypoglycaemic
Quinidine-like cardiac actions
Antimicrobial effects
Block graft-versus-host reaction*
Induce apoptosis*
Antioxidant actions: block superoxide release*
Antiproliferative effects*
Dissolve circulating immune complexes*
*These effects occur at levels higher than those reached with
routine clinical use in rheumatology practice.
and inhibit CpG-DNA-induced synthesis of IL‑6 and
TNF.24,25 Additionally, chloro­quine inhibits immune
stimulation by small nuclear RNA (a ligand for TLR7)
and subsequent production of IFN‑α.26 Taken together,
this evidence suggests that antimalarial agents antagonize TLR-mediated immune responses and synthesis
of interferon and inflammatory cytokines (Box 4). The
exact nature of this antagonism is not clear and could be
either non­competitive (by pH alteration) or competitive
(by structural binding). In order to function, intra­cellular
TLRs require an acidic pH,27 probably because they are
proteolysed by acid-dependent proteases28 that function
normally within the acidic endoplasm of lysosomes.
Antimalarial agents, by altering the lysosomal pH, possibly prevent functional transformation of intracellular
TLRs and inhibit their activation. Research published
in 2011 proposed that TLR antagonism by antimalarial
agents might not be driven entirely by endosomal pH
alterations but by mechanical inhibition, whereby antimalarial agents structurally bind to nucleic acids and
mask their TLR-binding epitopes.29
Other actions relevant to SLE
In addition to the effects of antimalarial agents on TLRs,
these compounds have several additional actions that are
relevant to SLE (Figure 3). Recent discoveries regarding
the mechanisms of these actions, concentrating on those
made in the period 2000–2012, are discussed below.
524 | SEPTEMBER 2012 | VOLUME 8
Ultraviolet light absorption
A well-described action of antimalarial agents is ultra­­
violet light absorption, and exposure to ultraviolet light
is a proven risk factor for cutaneous lupus lesions. 30
Ultraviolet radiation can induce local inflammation and
cell injury to keratinocytes, leading to cell death and possibly TLR activation. Antimalarial agents might act by
negating this ultraviolet ­radiation-induced inflammation.
Chloroquine concentration in the epidermis is 5–15 times
higher than the dermis,31 probably owing to its affinity
for melanin. This high concentration facilitates local
anti-­inflammatory actions of antimalarial agents such as
inhibition of antigen presentation and cytokine synthesis.
Additionally, anti­malarial agents activate transcription of
the c‑Jun gene, which is postulated to be a component
of the early protective response to ultraviolet light.32
Anti-lipidaemic effects
Lipid-lowering actions of antimalarial agents in patients
with SLE were demonstrated by our group in the 1980s
and 1990s in retrospective studies.33 Antimalarial agents
have been suggested to inhibit hydrolysis of internalized
cholesterol esters through their lysosomotropic action.34
Chloro­quine upregulates transcription of LDL-receptor
genes and probably affects the activity of HMG-CoA
reductase, a rate limiting enzyme in lipid metabolism.35 Further­more, over the past decade it has become
increas­ingly apparent that TLRs play a major part in lipid
metabo­lism. TLR9-mediated stimulation of perilipin‑3
increases lipid accumulation inside macro­phages. 36
Other studies have shown that TLRs have an important
role in atherogenesis.37 These discoveries provide a possible mechanistic explanation for the anti-lipidaemic
effects of antimalarial agents.
Anti-angiogenic effects
Lesiak et al.38 demonstrated in vivo that chloroquine
inhibits angiogenesis by reducing expression of vascular
endothelial growth factor and by interfering with CD34
glycoprotein, thereby improving cutaneous lesions. TLRs
(specifically TLR2, TLR4, TLR7 and TLR9) strongly
upregulate expression of vascular endothelial growth
factor upon stimulation with unmethylated CpG motifs
in the presence of adenosine receptors which then
promote angiogenesis.39 Thus, although no evidence yet
suggests that antimalarial agents block angiogenesis by
antagonizing TLRs, this putative mechanism of action
remains a possibility.
Antithrombotic effects
Antiphospholipid syndrome frequently coexists with
SLE. In this condition, antiphospholipid antibodies
(aPL) disrupt the protective covering over the phospholipid bilayer of the cell membrane formed by the
natural anticoagulant annexin A5. This loss of protection
exposes the phospholipids to coagulation factors, leading
to inappropriate activation of the coagulation pathway.
Hydroxychloroquine abrogates the stimulatory effect
of aPL on platelet aggregation even in the presence of a
thrombin agonist,40 and reverses the antibody-mediated
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Antimalarial agent masks
TLR-binding epitopes
of nucleic acids
pDC
Fc receptor
Nucleic
acid bound
antibody
Lysosome
Endocytosis
3
ssRNA
2
Acid pH
activation
Antimalarial
agent
DNA
TLR7
TLR9
Endocytosis
MyD88
1
Inactive
TLR
Nucleic
acid bound
antibody
4
IRF5–IRF7
Nucleus
Vesicle
5
Positive
feedback
amplification
Positive
feedback
amplification
Golgi
ER
Positive feedback
amplification
(dendritic cell
maturation)
Anti-dsDNA
antibodies
6
IFN-α secretion
NETs
IFN-α/βR2
8
7
IFN-α/βR2
9
IFN-α/βR2
B cell
Antibody secretion
and immune complex
formation
T cell
PMN cell
Antigen presentation
IL-10 production
TH1 cells
TREG cells
NETosis
Figure 2 | Role of antimalarial agents in TLR-mediated innate immune pathways in SLE. TLRs exit Golgi complex in an
inactive state (1), and are cleaved and activated in endosomes by acid-dependent proteases (2), thus interacting with
nucleic acids presented to endosomal compartments by specialized receptors such as Fcγ receptors (3). Putative actions
of antimalarial agents involve blocking (2) or (3), or both. Upon interaction with the respective ligands, TLRs stimulate the
synthesis of type I interferon (4,5). The release of IFN‑α (6) has widespread effects on both the innate and adaptive
immune systems; most importantly, it stimulates gene expression of TLRs as well as feedback activation of more pDCs,
perpetuating a vicious cycle.104 IFN‑α promotes T‑cell survival, upregulation of the TH1 response, proliferation of CD8+ cells
and suppression of TREG cells (7). IFN‑α affects B cells by causing maturation of plasmablasts, immunoglobulin classswitching, and increased antibody secretion (8). IFN‑α also stimulates the development of memory B cells and induces
BAFF, a B‑cell maturation and survival factor.104 In the presence of immune complexes and interferons, PMNs undergo
NETosis and NETs in turn stimulate pDCs—possibly via TLRs (9). IFN‑α also promotes maturation of monocytes into
dendritic cells, which are more efficient in antigen processing and presentation. It upregulates expression of co-stimulatory
molecules and HLA, and stimulates synthesis of IL‑10105 and IL‑12 by dendritic cells.106 Abbreviations: dsDNA, doublestranded DNA; ER, endoplasmic reticulum; IRF, interferon regulatory factor; NET, neutrophil extracellular trap; pDC,
plasmacytoid dendritic cell; PMN, polymorphonuclear cell; SLE, systemic lupus erythematosus; ssRNA, single-stranded
RNA; TH1 cell, type 1 T helper cell; TLR, Toll-like receptor; TREG cell, regulatory T cell.
disruption of the annexin A5 shield.41 These observations
provide a new rationale for the use of antimalarial agents
as preventive agents in aPL-positive patients.
MMP–TIMP modulation
Matrix metalloproteinases (MMPs) are a group of
enzymes involved in extracellular matrix remodeling.
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Box 3 | NETosis
NETs are chromatin structures containing antimicrobial
peptides formed by neutrophils and are capable of
trapping microorganisms, thus constituting a form
of innate immunity. A process of neutrophil cell
death, distinct from apoptosis and necrosis, results
in release of these NETs and is hence referred to as
NETosis. As these NETs are rich in antigenic structures,
including self-nucleic acids, it has been suggested
that activation of plasmacytoid dendritic cells by
NETs is a putative pathogenic mechanism in systemic
lupus erythematosus, and NET formation has been
implicated in progression of endothelial damage and
renal disease.107
Abbreviation: NET, neutrophil extracellular trap.
Box 4 | TLR-dependent effects in SLE
■■ TLR3, TLR7 and TLR9 are nucleic acid-sensing TLRs
that have been implicated in the pathogenesis of SLE
■■ By activating the immune system, TLRs upregulate
production of inflammatory cytokines including IFN‑α
■■ Activation of TLRs is a controlled step requiring an
acidic pH
■■ Antimalarial agents antagonize TLR activation possibly
by altering pH or via competitive inhibition
■■ TLR antagonism results in inhibition of IFN‑α
expression and thereby prevents activation of multiple
IFN‑α-mediated pathways and resultant widespread
inflammatory damage
Abbreviations: SLE, systemic lupus erythematosus; TLR,
Toll-like receptor.
These enzymes are secreted by monocytes, macrophages,
neutrophils, fibroblasts, endothelial cells and various
tumour cells. Tissue inhibitors of metalloproteinases
(TIMPs) are counter-regulatory enzymes that inhibit
MMPs and maintain homeostasis of the extra­cellular
matrix. An imbalance between MMPs and TIMPs has
been suggested to have a pathogenic role in auto­immune
diseases. 42 Quinacrine and chloroquine have been
shown to inhibit MMPs in vitro and in vivo, respectively.
Stuhlmeier et al.43 demonstrated that quinacrine, but not
chloroquine, downregulates the transcription of mRNA
thereby inhibiting the synthesis of MMP‑1, MMP‑2 and
MMP‑8. Lesiak et al.44 found that blood levels of MMP‑9
and TIMP‑1 were significantly elevated in 25 patients
with SLE in comparison with 25 healthy individuals;
following 3 months of treatment with chloro­quine,
MMP‑9 levels were significantly reduced and TIMP
levels were increased in the patients with SLE, suggesting that chloro­quine modulated MMP–TIMP inter­
actions. Studies have suggested that activation of TLR9
(and TLR4) promotes MMP production.45 Chloroquine
possibly acts through a TLR-mediated pathway whereas
quinacrine acts at the mRNA transcription level to
achieve similar effects on MMP–TIMP modulation.
Other mechanistic effects of quinacrine
Similar to other, weakly basic antimalarial agents, quinacrine also shows tropism for lysosomes.46 Additionally,
526 | SEPTEMBER 2012 | VOLUME 8
quinacrine exerts an inhibitory effect on B‑cell-activating
factor (also known as BLyS and TNF ligand super­family,
member 13b)—a survival factor for B cells and an important therapeutic target in SLE.47 Quinacrine stabilizes the
cell membrane and inhibits the enzymatic activity of cytosolic phospholipase A2, thereby inhibiting the arachido­
nic acid pathway and eventually resulting in inhibition of
inflammatory processes such as chemotaxis, cell adhesion
and platelet aggregation.48
Clinical effects in SLE
Antimalarial agents have both disease-modifying effects
and benefits for specific outcome measures in SLE
(Box 5). The effects of these agents on various disease
outcomes are discussed in the following sections.
Effects on SLE onset, progression and survival
In 1991, the Canadian Hydroxychloroquine Study Group
demonstrated that discontinuation of hydroxychloro­
quine increased the relative risk of a clinical flare by 2.5
times over a 6‑month period, compared with maintenance
of hydroxychloroquine therapy. Also, the risk of severe
disease exacerbation was 6.1 times higher for patients on
placebo as compared with those who continued hydroxychloroquine.49 Several subsequent clinical trials over
the past decade have confirmed the clinical benefits of
hydroxychloroquine in SLE patients (Table 1).50–55
In 2002, Molad et al.50 reported that, in a cohort of
151 patients with SLE, hydroxychloroquine use corre­
lated negatively with Systemic Lupus International
Col­labor­ating Clinics–American College of Rheumato­
logy (SLICC–ACR) damage index score, and positively
with damage-free survival measured over a period of
3.5 years.
In the LUMINA (Lupus in Minorities, Nature versus
Nurture) cohort, hydroxychloroquine use was associated
with reduced accrual of new damage. Patients with no evidence of prior damage benefited the most,51 which seems
to suggest that early initiation of hydroxychloroquine
potentially maximizes its benefits. Hydroxychloroquine
also improves overall survival: in 608 patients, 5% of
deaths occurred in patients using hydroxychloroquine as
compared with 17% in patients not receiving hydroxychloroquine.52 Ruiz-Irastroza et al.53 also reported a survival benefit associated with use of antimalarial agents in
232 patients with SLE from Spain.
In 2007, a retrospective chart review of 130 US military personnel demonstrated that hydroxychloroquine
use in patients before diagnosis of SLE was associated
with a delay in the onset of SLE from the time of appearance of initial symptoms (median time 1.08 years), as
compared with no use of hydroxychloroquine (median
time 0.29 years). The same paper reported that hydroxy­
chloroquine use before SLE diagnosis was associated with
lesser likelihood of developing proteinuria, leucopenia
or lymphopenia.54
In the longitudinal GLADEL (Grupo Latino Ameri­
cano de Estudio del Lupus Eritematoso) cohort of 1,480
patients with SLE, antimalarial agents were determined
to have a beneficial effect on survival, possibly in a
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TLR-independent mechanisms of antimalarial therapy
UV protection
Antilipidaemic effects
■ Local anti■ Act at the lipid
inflammatory
receptor level to
effects and
regulate enzyme
upregulation of
activity and possibly
the protective
also through TLRs
c-Jun-encoding gene ■ Reduce LDL, VLDL
■ Control of photoand cholesterol,
sensitivity and
and increase
cutaneous lupus
HDL levels
Antiangiogenic effects
■ Reduce epidermal
expression of VEGF
■ In vitro antiproliferative and
apoptotic effects
on ECs
■ Possible mode
of action in
discoid lupus
MMP−TIMP
modulation
PLA2
inhibition
BAFF
inhibition
■ Inhibit
expression
of MMP-1,
MMP-2, MMP-8,
MMP-9
■ Regulate ECM
homeostasis
■ Inhibit excess
ECM breakdown
■ Cell membrane
stabilization
■ Inhibit
arachidonic acid
pathway and
downstream
synthesis of
inflammatory
mediators
■ Reduce
maturation
and survival
of B cells,
including
autoreactive
B cells
Antithrombotic effects
■ Inhibit platelet aggregation
■ Block interaction between
platelets and coagulation
factors
■ Reduce thrombotic events
in patients with SLE
■ Possible role in primary
thromboprophylaxis in APS
and SLE
Figure 3 | Summary of TLR-independent mechanisms of antimalarial agents. Antimalarial agents have several actions that are relevant to SLE
and independent of their effects on TLRs. Abbreviations: APS, antiphospholipid syndrome; BAFF, B-cell activating factor; EC, endothelial cell;
ECM, extracellular matrix; MMP, matrix metalloproteinase; PLA2, phospholipase A2; SLE, systemic lupus erythematosus; TIMP, tissue inhibitor of
metalloproteinases; TLR, Toll-like receptor; UV, ultraviolet; VEGF, vascular endothelial growth factor.
time-dependent manner.55 In other words, patients who
used antimalarial agents for a longer time had lower
mortality than patients who used them for a shorter time.
Thus, early initiation of antimalarial agents in symptomatic patients delays the onset and progression of SLE and
improves overall survival.
Effects on specific outcome measures
In addition to its benefits for survival, hydroxychloroquine has been shown to improve morbidity in lupus
by affecting specific outcome measures such as integument damage, thrombosis, lipid profile, glycaemic status,
infections and renal failure (summarized in Table 2).
Data from 580 SLE patients in the LUMINA cohort suggests that hydroxychloroquine use is associated with
less integu­ment damage. The cumulative probability of
develop­ing integument damage at 5 years was 5% for
patients on hydroxychloroquine as opposed to 24%
for those not taking hydroxychloroquine.56
Since our initial report in 1987,57 numerous studies
have seemingly confirmed that hydroxychloroquine
is thrombo­protective in SLE.53,58–62 The lipid-lowering
proper­t ies of hydroxychloroquine, chloroquine and
quinacrine have been demonstrated in retrospective as
well as blinded, prospective study settings.33,63–65 Data
from the Baltimore lupus cohort demonstrated that
the mean blood glucose level in hydroxychloroquine
users was significantly lower than patients not taking
hydroxychloroquine.66
Hydroxychloroquine use is also protective against
renal damage 67,68 and major infections in SLE. 69
Additionally, evidence suggests that antimalarial agents
may prevent bone mass loss, subclinical atherosclerosis
and development of cancer in SLE (moderate-to-low
level evidence).70
Special considerations in SLE
Although antimalarial agents have been in widespread
clinical use for decades, they are often used suboptimally
in SLE.71 In this section, we present an update on the
clinical use of these agents along with ‘clinical pearls’ we
have employed over the years in using these agents.
Box 5 | Summary of effects of antimalarial agents in SLE
■■ Hydroxychloroquine has sustained beneficial
effects on overall survival, disease-free survival
and damage accrual
■■ Hydroxychloroquine delays onset of SLE and reduces
the number of and severity of clinical flares
■■ Early use of hydroxychloroquine maximizes these benefits
■■ Antimalarial agents improve survival in a time-dependent
manner
■■ Use of hydroxychloroquine protects against thrombosis,
even in aPL-positive patients
■■ All three antimalarial agents have lipid-lowering
properties that are also apparent in patients taking
corticosteroids
■■ Antimalarial agents have beneficial effects on glycaemic
status in patients with SLE, and this benefit possibly
increases with duration of use
■■ Antimalarial agents have a protective effect against
renal damage and major infections in SLE
Abbreviations: aPL, antiphospholipid antibody; SLE, systemic
lupus erythematosus.
Antimalarial agents for cutaneous lupus
Hydroxychloroquine, chloroquine and quinacrine have all
been mainstays in the treatment of cutaneous lupus and
are considered first-line systemic agents for the treatment
of widespread skin manifestations. Hydroxy­chloroquine is
most often used as the initial agent, and more than 50% of
the patients respond to therapy with hydroxychloroquine
alone.72 If hydroxy­chloroquine monotherapy fails, the
addition of quinacrine could be beneficial. If the response
is still inadequate, we recommend a combination of
chloroquine and quinacrine. A combination of hydroxy­
chloroquine and quinacrine improves Cutaneous Lupus
Erythematosus Disease Area and Severity Index (CLASI)
score, enhances response rate in non­responders and
possib­ly reduces mean daily dose of steroids.73,74
Antimalarial agents and smoking
Tobacco smoking is thought to reduce the efficacy of
anti­malarial agents in lupus. This interference is dose-­
dependent (that is, the number of cigarettes smoked per
day is inversely proportional to the degree of clinical
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Table 1 | Beneficial clinical effects of antimalarial agents for patients with SLE
Reference
Study design
Population
Findings
Canadian
Hydroxychloroquine
Study Group (1991)49
Double-blind,
randomized,
placebo-controlled
47 patients with clinically stable
SLE assigned to continue
hydroxychloroquine therapy (n = 25)
or to receive placebo (n = 22)
Discontinuation of hydroxychloroquine
increased the relative risk of a clinical flare
Risk of severe disease exacerbation was 6.1
times higher for patients on placebo as
compared with those who continued
hydroxychloroquine
Molad et al. (2002)50
Cohort
151 patients with SLE
Hydroxychloroquine use correlated negatively
with damage index score, and positively with
damage-free survival
Fessler et al. (2005)51
Observational
cohort (LUMINA)
518 patients with SLE
Hydroxychloroquine use associated with
reduced accrual of new damage
Patients with no evidence of prior damage
benefited the most
Alarcon et al. (2007)52
Case–control study
within the LUMINA
cohort
608 patients with SLE (of whom
61 were deceased)
Hydroxychloroquine use increases overall
survival (5% of hydroxychloroquine users died
vs 17% of non-users)
Ruiz-Irastorza et al.
(2006)53
Observational
prospective cohort
232 patients with SLE from Spain
Survival benefit associated with use of
antimalarial agents
James et al. (2007)54
Retrospective
chart review
130 US military personnel
Use of hydroxychloroquine before diagnosis
associated with delayed onset of SLE, and
with lesser likelihood of developing proteinuria,
leucopenia or lymphopaenia
Shinjo et al. (2010)55
Longitudinal cohort
(GLADEL)
1,480 patients with SLE
Beneficial effect of antimalarial agents on
survival, with greater benefit associated with
longer duration of use
Abbreviation: SLE, systemic lupus erythematosus.
res­ponse to antimalarial therapy) and this effect is especially pronounced in cutaneous lupus.75,76 Conversely,
patients who quit smoking seem to respond better to treat­
ment with antimalarial agents.77 The exact mechanism by
which smoking interferes with the efficacy of anti­malarial
agents is not well understood. Tobacco smoke is a potent
inducer of cytochrome p450 enzymes (Box 1);78 one hypo­
thesis is that it interferes with metabo­lism of antimalarial
agents. However, Leroux et al.78 did not find any correlation between blood concentrations of the metabolites of
hydroxychloroquine and smoking habits in 223 patients,
which suggests that the interference might involve other,
indirect pathways. Other possible pathways common to
the mechanisms of antimalarial agents include MMP–
TIMP modulation via IL‑10 and TNF,79,80 and induction
of cytokines such as IL‑6 by tobacco smoke.81
On the other hand, a few reports found no associ­
ation between smoking and response to antimalarial
agents.82,83 Also, smoking is independently associated
with cutaneous lupus,84 thus confounding the picture.
Smoking is also associated with skin lesions of lupus that
are refractory to conventional therapies not limited to
antimalarial agents,85 suggesting the possibility that the
interference caused by smoking is not restricted to antimalarial agents. However, this perception was challenged
in 2012 by a prospective study in patients with cutaneous
lupus, which reported that smokers, surprisingly, had a
better response when treated with antimalarial agents
alone.86 Clearly, more prospective clinical studies free
of confounders and biases, as well as better biological
models, are required to understand the exact relationship
between smoking and antimalarial agents. Regardless, we
528 | SEPTEMBER 2012 | VOLUME 8
advise that all patients, irrespective of their therapeutic
regimen, should be encouraged to quit smoking.
Antimalarial agents in pregnancy and lactation
Whilst we recommend a detailed conversation with each
individual patient about the potential risks of any therapeutic agent in pregnancy, several studies have assessed the
effect of hydroxychloroquine on pregnancies in women
with SLE. In case series published in the 1980s and 1990s,
Parke87,88 suggested that hydroxychloroquine was safe to
use in pregnant patients with SLE. Buchanan89 reported
that exposure to hydroxychloroquine during pregnancy
did not have any terato­genic effects. Data regarding 257
pregnancies from the Hopkins cohort showed no fetal
abnormalities directly attributable to hydroxychloroquine; this study also reported that stopping hydroxychloroquine during or just before preg­nancy resulted
in increased disease activity.90 Hydroxychloroquine use
during pregnancy by patients positive for anti-Ro/SSA or
anti-La/SSB antibodies is associated with reduced risk of
developing the cardiac manifestations of neonatal lupus.91
Thus, hydroxychloroquine use during pregnancy results
in favourable maternal outcomes and there has been no
evidence of visual or auditory abnormalities in the foetus,
either congenital or developmental.92
Hydroxychloroquine is secreted in breastmilk.
Hydroxy­c hloroquine ingestion by breastfed infants
corres­­ponds to 0.06–0.20 mg/kg per day after adjustment
for body weight. Compared with the adult therapeutic
dose of 6.5 mg/kg per day, the amount transferred can be
con­sidered a low dose unlikely to cause any significant
toxic effects.93
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Table 2 | Beneficial clinical effects of antimalarial agents on specific SLE outcomes
Reference
Study design
Population
Findings
Ho et al.
(2005)58
Observational
cohort (LUMINA)
442 patients with SLE and known
aPL status
Hydroxychloroquine use protective against
thrombosis in univariate analysis (OR 0.536)
Ruiz-Irastroza
et al. (2006)53
Observational
prospective cohort
232 patients with SLE
Antimalarial use protective against thrombosis
(HR 0.28)
Sisó et al.
(2008)59
Cohort
206 patients with lupus nephritis,
56 of whom previously used
antimalarial agents
Previous hydroxychloroquine use protective against
thrombosis (5% in hydroxychloroquine users vs 17%
in non-users)
Kaiser et al.
(2009)60
Cohort
1,930 patients with SLE
Hydroxychloroquine use thromboprotective after
propensity analysis (OR 0.62)
Tektonidou
et al. (2009)61
Longitudinal
case–control
144 aPL positive patients with SLE
matched with 144 aPL-negative
patients with SLE
Hydroxychloroquine use protective against
thrombosis in both groups (aPL positive patients:
HR per month 0.99; aPL negative patients: HR per
month 0.98)
Jung et al.
(2010)62
Nested case–
control
162 patients with SLE, 54 of whom
had a history of thrombotic event
Ever-use of antimalarial agents thromboprotective
(OR 0.32)
Overall 68% reduction in thrombotic events in users
of antimalarial drugs
Wallace et al.
(1990)33
Retrospective
155 patients with SLE or RA
subdivided according to exposure to
hydroxychloroquine and/or steroids
Addition of hydroxychloroquine to steroids reduced
levels of LDL cholesterol and triglyceride by 15%
compared with steroids alone
Petri et al.
(1994)63
Longitudinal
cohort (Hopkins)
264 patients with SLE
Hydroxychloroquine use associated with an
8.94mg% reduction in serum total cholesterol level
Kavanaugh
et al. (1997)64
Double blind,
prospective
17 patients with SLE
Hydroxychloroquine associated with a mean
decrease of 11.6 mg/dl in serum total cholesterol
level
Borba et al.
(2001)65
Case–control
60 patients with SLE (subdivided
according to exposure to
hydroxychloroquine and steroids)
and 30 healthy controls
Chloroquine use associated with elevated level of
HDL cholesterol as compared with no therapy, and
with lower level of VLDL cholesterol in the group
taking chloroquine plus steroids
Pons-Estel
et al. (2009)67
Prospective study
in a longitudinal
cohort (LUMINA)
203 patients with lupus nephritis
without renal damage at baseline
Hydroxychloroquine use associated with lower
frequency of WHO class IV glomerulonephritis, lower
disease activity and lower steroid requirement
Hydroxychloroquine protective against renal
damage occurrence in full (HR 0.12) and reduced
(HR 0.29) models after adjustment for confounding
factors
Pons-Estel
et al. (2012)68
Nested case–
control study
within the GLADEL
cohort
265 patients with SLE and renal
disease and 530 controls without
renal disease
Use of antimalarial agents negatively associated
with risk of development of renal disease (OR 0.39)
Protective effect persisted after adjustment for
confounding factors (OR 0.38)
Longitudinal
cohort (LUMINA)
580 patients with SLE
Hydroxychloroquine use associated with longer time
to integument damage (HR 0.23)
Cumulative probability of developing integumental
damage at 5 years lower in patients taking
hydroxychloroquine than those not taking the drug
(5% vs 24%)
Longitudinal
cohort
71 patients with SLE who had serial
cohort visits both on and off
hydroxychloroquine
Mean blood glucose level whilst taking
hydroxychloroquine was 84.9 ± 15.2 mg/dl,
significantly lower than level whilst off
hydroxychloroquine (89.0 ± 21.5 mg/dl)
Nested case–
control within a
prospective cohort
86 patients with SLE and major
infections and 166 controls (without
major infections)
After logistic regression, treatment with antimalarial
agents had an independent protective effect against
major infections in SLE (OR 0.06)
Thrombosis*
Lipid profile*
Renal damage
Integument damage
Pons-Estel
et al. (2010)56
Glycemic status
Petri (1996)66
Major infections
Ruiz-Irastorza
et al. (2009)69
*Studies included are evidence-based level A or B studies with various study designs. Abbreviations: aPL, antiphospholipid antibody; RA, rheumatoid arthritis;
SLE, systemic lupus erythematosus.
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Box 6 | Non-ophthalmologic toxic effects
■■ Common*: nausea, vomiting, pruritus, maculopapular
rash, skin & mucosal pigmentation, insomnia,
nightmares, nervousness
■■ Less common‡: tinnitus, vestibular changes,
neuropathy, psychosis, seizures
■■ Rare: leucopenia, anaemia, porphyria, depigmentation
of hair, hair loss, anorexia, diarrhoea, liver dysfunction
*Frequency 1–20% with chloroquine use, 1–10% with
hydroxychloroquine use. ‡Frequency <5% with chloroquine use,
<1% with hydroxychloroquine use.
Box 7 | New insights into ophthalmologic toxicity
■■ Retinopathy can be missed in early stages and might
be irreversible later
■■ Baseline screening at the beginning of treatment
should be followed by annual screening after 5 years
of treatment
■■ Pre-existing macular disease should be considered a
contraindication for antimalarial therapy; if antimalarial
agents must be used, annual screening should start
from the onset of treatment
■■ Special consideration should be given to elderly
patients and those with kidney or liver disease,
entailing annual screening from the onset of treatment;
short-statured and obese patients also deserve
careful monitoring
■■ Recognize the possibility of ‘outlier’ cases, in which
retinopathy occurs even within the safe limits of
antimalarial treatment; patients should be made
aware of this possibility and provided with risk–benefit
counselling
■■ Quinacrine can be used relatively safely in the
presence of pre-existing retinopathy
Thus, from the available evidence, we opine that
hydroxychloroquine is generally safe for use during pregnancy and lactation and should be continued throughout
this period since its withdrawal may be associated with
worsening of the disease and adverse outcome of the preg­
nancy. Hydroxychloroquine might also reduce the risk of
heart block associated with neonatal lupus.
Update on the safety of antimalarial agents
Antimalarial agents are generally considered relatively safe and non-toxic as compared with other
­disease-modifying agents for SLE. Non-ophthalmologic
adverse effects of antimalarial agents are listed in Box 6.
Advances since 2000 have increased our understanding
of the ocular toxicity of antimalarial agents, and these
are discussed in detail below and summarized in Box 7.
Ocular adverse effects of antimalarial agents include
keratopathy in the form of corneal deposits, bull’s eye
maculopathy, cycloplegia and posterior cataracts. The
incidence rate of keratopathy is more than 50% (up to
90%) with chloroquine, 10% with hydroxychloroquine
and approximately 5% with quinacrine.94 Keratopathy is
almost always completely reversible without any residual
corneal damage and by itself is not an indication for stopping treatment. Retinopathy is seen in 10% of patients
receiving chloroquine and in approximately 1% of those
530 | SEPTEMBER 2012 | VOLUME 8
receiving hydroxychloroquine after 7 years;94 it is not a
reported consequence of quinacrine use.
The adverse effect profile of quinacrine is generally
similar to the other two antimalarial agents, but some
important differences exist. First, quinacrine does not
cause retinopathy or haemolysis in patients deficient in
­g lucose‑6-phosphate dehydrogenase. Second, quinacrine use has been associated with aplastic anaemia, but
infrequently in patients receiving modern, lower doses
of quinacrine, thus making aplastic anaemia a very
rare event.95
The causal association between use of antimalarial
agents and retinopathy has been recognized for many
years, but the precise incidence of retinopathy has been
a topic of debate. Although it is argued that retinopathy
is a rare adverse effect of antimalarial therapy, the sheer
number of patients undergoing long-term treatment
with these agents makes it an important issue. Much has
been speculated about the mechanism of retinal toxicity of antimalarial agents—theories put forward include
melanin binding, phototropic effect and alteration of
retinal pigment metabolism—but no theories have been
conclusively proven. The clinical picture of chronic eye
toxicity is bull’s eye maculopathy, which often occurs
bilaterally. The patient might have excellent visual acuity
in the initial stages despite underlying foveal changes.
These changes are almost always reversible if monitoring is performed in accordance with updated guidelines
on ophthalmologic monitoring for patients on anti­
malarial agents published by the American Academy of
Ophthalmology in 2011.96
The cumulative dose of antimalarial agents has
recently been recognized as an independent risk factor
for retinopathy,96 a consideration that is prominent in
the American Academy of Ophthalmology guidelines.96
Advanced age, impaired liver and/or renal function
and pre-existing macular disease have been identified
as additional risk factors.96 Reflecting advances since
publication of the previous guidelines in 2002, the new
recommendations call for the use of more-advanced
and objective tests for eye monitoring. In addition to
dilated eye examination, the guidelines recommend
automated visual field testing, multifocal electro­
retinogram, spectral domain coherence tomography
and fundus autofluorescence; by contrast, use of Amsler
grid testing, fluorescein angiography and fundus photo­
graphy are no longer recom­mended. A cumulative dose
of 1,000 g hydroxychloroquine is considered the typical
safety limit above which caution is advised. This limit
is reached within 7 years for patients receiving the
usual hydroxy­chloroquine dose of 400 mg per day. The
guidelines advise that dosing should be adjusted according to ideal (lean) body weight and not by actual body
weight because antimalarial agents are not retained in
adipose tissue; this adjustment is particularly important
con­sidering the global rise in the incidence of obesity
and the weight gain associated with steroids. Finally, it
should be noted that patients with kidney or liver disease
and elderly patients carry an increased risk of earlier
develop­ment of retinopathy.96
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Table 3 | Community-based use of antimalarial agents in patients with lupus: authors’ perspective
Agent
Administration
Monitoring
Hydroxychloroquine
Once‑a-day dose of 5 mg/kg can be considered optimal in most cases, but can
be used at 7 mg/kg per day for up to 3 months
This dose should be reduced back to 5 mg/kg or less for maintenance therapy
This regimen gives a response rate of up to 80% for non-organ-threatening
disease and cutaneous lupus
In our experience, anti-inflammatory and photoprotective effects are seen by
6–12 weeks
Monitor CBC and liver and
kidney functions every
3–4 months for the duration
of use
Perform eye examinations
as per AAO guidelines96
Chloroquine
Can be used for refractory cutaneous disease at 500 mg per day for
30–60 days followed by 250 mg per day
Transition to hydroxychloroquine is advised once a response is seen;
however, if transition is not possible, tapering should be achieved by reducing
the frequency of administration by one day every week, keeping the dose
(mg per day) the same
Monitor CBC and perform
comprehensive metabolic
panel every 3 months
Perform eye examinations
as per AAO guidelines96
Quinacrine
Add-on treatment for refractory skin disease (synergistic with
hydroxychloroquine or chloroquine) or as monotherapy for patients with
contraindications to these two drugs (for example, those with glucose‑6phosphate dehydrogenase deficiency or pre-existing retinopathy)
Starting dose 50 mg per day and then can be adjusted to 25–100 mg per day
according to the response observed
Should be stopped if no response is seen by 6 weeks or if cytopaenia or
lichen planus develops
Monitor CBC and perform
comprehensive metabolic
panel every month for the
first 3 months and every
3 months thereafter
Eye examination is not
required
Abbreviation: AAO, American Academy of Ophthalmology.
How to use antimalarial agents in SLE
As antimalarial agents have disease-modifying proper­ties
and can affect inflammatory activity as well as favourably
influence quality of life, morbidity and prognosis, most
patients with SLE should be treated with one of these
agents. Table 3 summarizes how we use anti­malarial
agents in our patients with SLE.
Potential translational insights
TLRs have been implicated in a variety of rheumatologic
disorders, and both laboratory and clinical research
seem to show a promising role for antimalarial agents
in the treatment of these diseases.97 Attempts are being
made to synthesize small molecule TLR antagonists that
can be administered orally or subcutaneously. 98 The
enantio­specific metabolism of antimalarial agents provides an opportunity to develop a pure formation of the
S‑enantiomer, which could be less toxic, especially to the
retina, than current formulations owing to its preferential metabolism and elimination (Box 1). Topical preparations of hydroxychloroquine could be useful in treating
cutaneous lupus.99 It has been suggested that patients
with genotypes associated with expression of low levels of
IL‑10 and high levels of TNF respond best to anti­malarial
therapy.100 As additional data from genome-wide association studies becomes available, this area of individualized
tailoring of medications on the basis of genotype will
become more important. Quinacrine has also generated
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Author contributions
D. J. Wallace, V. S. Gudsoorkar and S. R. Venuturupalli
researched the data for the article. All authors
provided a substantial contribution to discussions of
the content and contributed equally to writing the
article and to review and/or editing of the manuscript
before submission.
VOLUME 8 | SEPTEMBER 2012 | 533
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