3. Clinical evidence of immunomodulatory and anti-inflammatory
agents
Since the beginning of the COVID-19 pandemic, the determining role of
inflammatory cytokines in the worsening of clinical conditions has been
identified. Indeed, one of the reasons underlying the occurrence of
serious symptoms (figure 1) associated with the clinical progression of
SARS-CoV-2 infection is represented by the increase in levels of
pro-inflammatory cytokines, mainly interleukins, granulocyte-colony
stimulating factor and TNF-α. These molecules are responsible for the
so-called cytokine storm that, in turn, induces ARDS, organ failure and
sepsis (Guo et al., 2020b). Many drugs, acting through different
mechanisms, are able to block or reduce the effects mediated by the
cytokines storm (figure 2).
Tocilizumab was one of the most studied drugs able to reduce
the hyperinflammatory state associated with COVID-19. It is a monoclonal
antibody (figure 2), authorized for the treatment of many diseases,
including rheumatoid arthritis, which is active against the
interleukin-6 receptor, one of the key mediators of the inflammatory
process (Scott, 2017). At the beginning of the pandemic, the drug was
tested in China to reduce lung complications in 20 patients with severe
SARS-CoV-2 infection, being associated with a reduction of oxygen
requirement, resolution of CT lesions, normalization of lymphocyte
count, reduction of C-reactive protein levels and hospital discharge (Xu
et al., 2020). Few months ago, many trials evaluating the effects of
tocilizumab started in Italy, after the approval of the Italian Medicine
Agency and the Spallanzani Ethics Committee. However, despite the
encouraging preliminary results, the availability of new evidence from
other ongoing clinical trials has marked a change of course. Indeed, the
study carried out by the Local Health Unit-IRCCS of Reggio Emilia (which
evaluated the efficacy of tocilizumab at an early stage in patients with
COVID-19 pneumonia who did not require invasive or semi-invasive
mechanical ventilation procedures invasive) was earlier stopped, after
the enrolment of 126 patients. The reason for the early conclusion lies
in the absence of proof of benefit in treated patients either in terms
of worsening (access to the ICU) or survival (Italian Medicine Agency,
2020a). Similarly, the results of the COVACTA trial, who was ongoing
when we published our first paper (Scavone et al., 2020), did not reveal
a benefit of tocilizumab over placebo in patients with severe COVID-19
pneumonia. Indeed, as reported in a press document released by the
pharmaceutical company, no difference in clinical status between
patients treated with tocilizumab and placebo were found (primary
endpoint). In addition, no differences between groups were found
regarding to the percentage of patients who died by week four and in
ventilator-free days (Roche, 2020). Despite the negative results of
these trials, tocilizumab was associated with positive outcome in other
clinical studies, both interventional and observational. For instance,
the single arm trial TOVICID-19 evaluated the effects of tocilizumab in
hospitalized patients with SARS-CoV-2 infection, oxygen saturation at
rest in ambient air ≤ 93% or required oxygen support or mechanical
ventilation either non-invasive or invasive. One hundred and eighty
patients received a dose of 8 mg/kg up to a maximum of 800 mg per dose
and a second administration 12 h after the first one if respiratory
function did not recover. The results revealed that the treatment with
tocilizumab seems to reduce lethality at 30 days, although its impact at
14 days seems less relevant (Perrone et al., 2020). Lewis TC et al.
reported the results of an observational cohort study of patients
hospitalized at 3 hospitals within the NYU Langone Health system in New
York who received tocilizumab 400-mg IV once in addition to standard of
care or standard of care alone. Out of 3,580 patients, 497 were treated
with tocilizumab. The propensity score analysis showed that a lower
number of death occurred in tocilizumab-treated patients and that
tocilizumab was associated with improved survival compared to controls
(p>0.001). No difference were found between groups in term
of adjusted time to hospital discharge, while patients treated with
tocilizumab had longer adjusted ICU length of stay and a higher adjusted
infection rate (both p>0.001) than controls (Lewis et al.,
2020). Tleyjeh IM et al. carried out a systematic review and
meta-analysis of 24 studies, including 5 randomized controlled trials
and 19 observational studies (enrolling 1,325 and 10,021 patients,
respectively). The results of this study demonstrated that tocilizumab
is able to reduce the risk of mechanical ventilation in hospitalized
COVID-19 patients but not the short-term mortality, at least according
to the results of randomized controlled trials. On the contrary, data
from observational studies suggested an association between tocilizumab
and lower mortality. Authors suggested that many reasons could explain
differences found for the effect on the mortality, including the
enrolment of different patient populations, a different inflammatory
status, and timing/dosing of tocilizumab. Furthermore, in order to be
sufficiently powered to detect a difference in mortality between
tocilizumab and control groups, the sample size of clinical trials
should have been substantially higher. In terms of safety profile, no
higher risk of infections or any other adverse events was found with
tocilizumab use (Tleyjeh et al., 2020). Given the evidence currently
available on the effects of tocilizumab in COVID-19 patients no firm
conclusion can be drawn. We still have to wait for the results of other
studies, such those from the RECOVERY trial, which is currently
evaluating the efficacy of tocilizumab and other drugs, including
low-dose dexamethasone, colchicine, convalescent plasma, REGN-COV2 and
aspirin (University of Oxford, 2020b). Furthermore, biochemical
parameters like serum IL6 levels and TH17 cells may contribute to select
subgroups of COVID-19 patients that would specifically benefit from the
treatment with Tocilizumab, according to the view of a personalized
therapy in such patients (Cacciapuoti et al., 2020).
Conflicting results were also obtained for sarilumab (figure
2), a drug belonging to the same drug class of tocilizumab. In our first
document (Scavone et al., 2020), we stated that three randomized
controlled trials were underway. Nowadays, the results of one trial
investigating the effects of sarilumab in severely or critically ill
patients hospitalized with COVID-19 did not meet its primary and
secondary endpoints (Sanofi, 2020b). Other studies, mainly
observational, that investigated the effects of sarilumab were carried
out on a very limited number of patients. For instance, Benucci M et al.
evaluated the efficacy of sarilumab in 8 patients (mean age: 62 years)
hospitalized at the San Giovanni di Dio Hospital (Florence, Italy). The
drug was administered in a dose of 400 mg in combination with
hydroxychloroquine, azithromycin, darunavir, cobicistat and enoxaparin.
Seven patients had an improvement of the Horovitz index (oxygenation
expressed by an increased SpO2/FiO2 ratio). A progressive reduction in
the serum amyloid A and CRP inflammation parameters was observed and
patients were discharge within 14 days of hospitalization (Benucci et
al., 2020). Another study, carried out at the Fondazione Policlinico
Universitario A. Gemelli-IRCCS (Italy), evaluated the effects of
sarilumab in 53 patients (median age: 66 years) with severe COVID-19.
Almost 70% of patients were also treated with darunavir/ritonavir, 24%
with lopinavir/ritonavir, while the remaining patients did not receive
antiviral treatment. Hydroxychloroquine, heparin and azithromycin were
administered in 94,3%, 74,9% and 54,7% of patients, respectively.
Thirty-nine patients were treated in medical wards (the majority with a
single infusion), while 14 in ICU (the majority with two infusions).
Among patients who received the drug in the medical wards, at 19 days
median follow-up, almost 90% significantly improved. Among patients who
received sarilumab in ICU, almost 36% were still alive at the last
follow-up. The overall mortality rate was 5,7% (Gremese et al., 2020).
A further retrospective study was carried out at the Azienda Ospedaliera
dei Colli – Cotugno Hospital (South of Italy) on 15 patients (12
treated with a single dose and 3 with two infusions). All patients
received hydroxychloroquine, lopinavir/ritonavir and heparin, while 11
patients were also treated with methylprednisolone. Improvements in
respiratory parameters were observed in 10 patients after sarilumab
administration. Five patients who received sarilumab died (Montesarchio
et al., 2020). Overall, the majority of clinical studies carried out on
sarilumab were observational and limited by a low number of patients. In
addition, many other drugs were co-administered. Therefore, it is not
easy to fully appreciate the positive effect deriving from sarilumab
administration. Thus, also for this drug further clinical data are
strongly needed.Chloroquine and hydroxychloroquine (figure 2) are
noteworthy too. These are antimalarial drugs that have also been used
for the treatment of different rheumatic diseases (Mascolo et al.,
2018). Both drugs show many pharmacological effects, including the
stabilization of the lysosomal membranes, the inhibition of
prostaglandin synthesis, polymorphonuclear chemotaxis and phagocytosis
and a possible interference with the production of interleukin 1 by
monocytes and inhibition of the release of superoxide by neutrophils
(Italian Medicine Agency). These drugs have demonstrated to be effective
in inhibiting the replication of SARS-CoV-2 in experimental models
(figure 2) (Italian Medicine Agency) and have been increasingly tested
for COVID-19 in many clinical studies. Specifically, many studies, which
are currently ongoing, are evaluating the effects of hydroxychloroquine
as prophylactic agent (Lother et al., 2020; Niriella et al., 2020;
Tirupakuzhi Vijayaraghavan et al., 2020). At the moment, only one
randomized, double-blind, placebo-controlled study on the prophylactic
role of hydroxychloroquine was concluded. The trial enrolled 821
subjects who had had family or occupational exposure with a subject with
COVID-19. According to the study results, there was no difference in the
incidence of new onset of COVID-19 between participants treated with
hydroxychloroquine (11.8%) and those who received placebo (14.3%).
Adverse events were more commonly observed with hydroxychloroquine than
with placebo, although no serious adverse reactions were reported
(Boulware et al., 2020). Many clinical trials evaluated the efficacy and
safety of hydroxychloroquine, alone or in combination with azithromycin,
as therapeutic agent. Gautret P et al. carried out a single-arm
interventional study, which evaluated the effects of hydroxychloroquine
(plus azithromycin) in 36 hospitalized patients with COVID-19. Authors
reported that a significant reduction in viral load was found on day 6
in patients treated with hydroxychloroquine compared to controls. The
addition of azithromycin to hydroxychloroquine therapy significantly
contributed to the reduction of viral load (Gautret et al., 2020).
Another study enrolled 11 hospitalized patients treated with
hydroxychloroquine and azithromycin. After 5-6 days of starting
treatment, the nasopharyngeal swab gave a positive result in 8/10
patients. These virological results are in contrast with those reported
by Gautret et al. and raise doubts about the effectiveness of the drugs
combination (Molina et al., 2020). A further study found no difference
in viral load reduction and clinical outcomes between hydroxychloroquine
and standard of care (CHEN et al., 2020). In addition, the results of
the randomized clinical study by Tang et al., which was carried out on
150 hospitalized patients with COVID-19 (148 with mild to moderate
disease and 2 with severe disease), did not show any difference on the
viral clearance at 28 days. The only positive result came from a post
hoc analysis in a subgroup of patients who did not take antivirals in
which a greater effect on symptoms was observed (Tang et al., 2020).
Similarly, the results of a randomized, double-blind, placebo-controlled
clinical trial (carried out in 491 outpatients at an early stage of
COVID-19) found no differences in symptom severity over 14 days between
hydroxychloroquine and placebo (relative difference 12%; p=0.117).
(Skipper et al., 2020). Huang et al. conducted a randomized study that
showed no significant differences up to 14 days, in terms of viral
clearance and improvement on CT scan between chloroquine and
lopinavir/ritonavir, but on day 14, 100% of patients treated with
chloroquine were discharged from the hospital, compared with 50% in the
lopinavir / ritonavir group. However, it should be underlined that
patients in the chloroquine group were younger and they had started the
treatment earlier (Huang et al., 2020). Lastly, the preliminary results
of the SOLIDARITY study seem to suggest that hydroxychloroquine, the
combination lopinavir / ritonavir and IFN-based regimens have little or
no effect on mortality at 28 days or on hospital course (World Health
Organization, 2020c). Alongside with data on the efficacy profile of
chloroquine and hydroxychloroquine, many data on the safety profile of
both drugs were collected as well. For example, a recent warning
discouraged the friendly use of chloroquine in the COVID-19 disease due
to the possible side effects (like a severe haemolytic crisis) in G6PD
deficient males and in unaware carriers of the disease, that are
estimated to be about 400 million worldwide (Capoluongo et al., 2020).
Based on these, the main regulatory agencies have issued many
recommendations for clinicians. For instance, the EMA drew the attention
on risks of serious adverse reactions (including heart rhythm
disturbances) (European Medicine Agency, 2020a), highlighting the need
for prescribers to closely monitor patients treated with both drugs
(European Medicine Agency, 2020b). In addition, the European agency
recommended the use of these medicines only in clinical trials or in
national emergency management programs in hospitalized patients under
close monitoring (European Medicine agency, 2020). The FDA has also
highlighted the need for patients’ monitoring when treated with
hydroxychloroquine, especially regarding to the risk of severe changes
in heart rhythm (tachycardia, atrial fibrillation and torsades de
pointes), observed in patients treated with these drugs often in
combination with azithromycin and other drugs able to prolong the QT (US
Food and Drug Administration, 2020a). The American agency also
recommended the use of chloroquine and hydroxychloroquine only in the
context of clinical trials and revoked, in June 2020, the previously
granted Emergency Use Authorization (US Food and Drug Administration,
2020a). In May 2020, the World Health Organization (WHO) suspended the
hydroxychloroquine treatment arm in the multicentre clinical trial
SOLIDARITY (World Health Organization, 2020e). In June 2020, the
hydroxychloroquine treatment arm was reopened for enrolments, only to be
permanently closed in the same month (Italian Medicine Agency, 2020b).
Lastly, on November 2020, the PRAC of the EMA recommended updating the
product information for chloroquine or hydroxychloroquine due to the
risk of psychiatric disorders and suicidal behaviour (European Medicine
Agency, 2020d). In conclusion, data related to the efficacy and safety
profile of antimalarial drugs are quite conflicting. We should wait for
the results of clinical trials currently ongoing in COVID-19 patients
(www.clinicaltrials.gov).
At the time of the publication of our first paper (Scavone et al.,
2020), the evidence on the efficacy of corticosteroids in
patients with COVID-19 was quite conflicting. Nowadays, we know that
these drugs, especially dexamethasone and methylprednisolone, could be
considered one of the mainstays in the treatment of COVID-19, especially
for patients with severe disease, to prevent its worsening and
complications and to resolve its serious symptoms. As reported in figure
2, corticosteroids are associated with multiple mechanisms, including
the increase in the gene transcription of anti-inflammatory cytokines,
the decrease in gene transcription of pro-inflammatory cytokines,
chemokines, and adhesion molecules and a reduction in inflammatory
response. All these effects seem to be responsible of the counteraction
of the cytokine storm caused by SARS-CoV-2 infection. First and
foremost, the preliminary results of the RECOVERY trial, which were
recently published on the NEJM, provided the most significant evidence
supporting the use of dexamethasone. In this controlled, open-label
trial, patients were randomized to receive oral or intravenous
dexamethasone 6 mg/day for up to 10 days (n=2,104) or the usual care
alone (n=4,321). Within 28 days after randomization, 22.9% of patients
treated with dexamethasone and 25.7% of patients in the usual care
group died (age-adjusted rate ratio, 0.83; 95% confidence interval
[CI], 0.75 to 0.93; P<0.001). According to the level of
respiratory support, the incidence of death was lower in dexamethasone
group among patients receiving invasive mechanical ventilation or oxygen
without invasive mechanical ventilation (Horby et al., 2020a). These
results, together with those obtained from other randomized clinical
studies (Angus et al., 2020; Dequin et al., 2020; Jeronimo et al., 2020;
Tomazini et al., 2020), showed that dexamethasone could be considered a
treatment option in hospitalized subjects with severe COVID-19 infection
who require oxygen therapy, whether or not they have mechanical
ventilation (invasive or not invasive). Based on the preliminary results
of RECOVERY trial, on September 2020 the CHMP of the EMA concluded that
dexamethasone can be considered a treatment option for patients who
require oxygen therapy and the EMA is endorsing the use of this drug in
adults and adolescents (from 12 years of age and weighing at least 40
kg) who require supplemental oxygen therapy at a dose of 6 milligrams
once a day for up to 10 days (European Medicine Agency, 2020c). Recently
the WHO REACT Working Group carried out a meta-analysis of seven
randomized clinical trials that evaluated the efficacy of
corticosteroids (dexamethasone, hydrocortisone, or methylprednisolone)
in 1,703 critically ill patients with COVID-19 on all-cause mortality at
28 days and the occurrence of serious adverse events. The results showed
that, compared with usual care or placebo, the OR for the association
with mortality was 0.64 (p<0.001) for dexamethasone, 0.69
(p=0.13) for hydrocortisone and 0.91 (p=0.87) for methylprednisolone.
Sixty-four serious adverse events were observed among patients
randomized to corticosteroids and eighty serious adverse events among
patients randomized to usual care or placebo. Authors concluded that the
administration of systemic corticosteroids, compared with usual care or
placebo, was associated with lower 28-day all-cause mortality (Sterne et
al., 2020). Lastly, several clinical trials are currently underway or in
development to evaluate corticosteroids for the treatment of COVID-19
(www.clinicaltrials.gov).
Similarly to corticosteroids, nonsteroidal anti-inflammatory
drugs (NSAIDs) (figure 2) and paracetamol represent valuable
therapeutic options in patients with COVID-19, especially for
outpatients at early stage (not severe form), to treat fever and muscle
pain. At the beginning of the outbreak, the role of NSAIDs was misjudged
due to some concerns that arose from few studies (Capuano et al.,
2020b). Specifically, in 2009 a group of researchers from the University
of Rochester highlighted that NSAIDs might lower host defence following
infection or vaccination (Bancos et al., 2009). On April 2019 the French
regulatory agency reported the results of a survey carried out by the
regional pharmacovigilance centres of Tours and Marseille in which
serious infectious complications occurred in patients receiving NSAIDs,
mainly ibuprofen and ketoprofen, used in the treatment of fever and pain
(ANSM, 2019). On March 2020, the French Minister of Health recommended
the use of paracetamol instead of ibuprofen or oral cortisone for the
treatment of fever in patients with COVID-19. This statement was
supported also by the French Authorities, which announced that NSAIDs
might worsen clinical conditions of patients with COVID-19 (Le monde,
2020; Ministère de la Santé, 2020). Recently few clinical data have
become available, mainly from observational studies, regarding to the
use of NSAIDs in patients with COVID-19. For instance, Rinott E et al.
carried out a retrospective cohort study on 403 patients with COVID-19.
The authors evaluated the association between the use of ibuprofen,
administered from a week before diagnosis of COVID-19 throughout the
disease, and the mortality and the need for respiratory support. Of the
entire cohort, 87 patients received ibuprofen and 316 did not. No
significant difference was found between groups in terms of mortality
and respiratory support. The comparison of ibuprofen users with
paracetamol users did not reveal any difference in mortality rates or
the need for respiratory support (Rinott et al., 2020). A prospective
cohort study enrolled patients with COVID-19, who were interviewed about
NSAID use and infection outcomes (such as death, admission, severity,
time to clinical improvement, oxygen requirement and length of stay)
through a telephone questionnaire. According to study’s results, neither
the acute use of ibuprofen nor chronic NSAID use were associated with a
greater risk of mortality compared to non-use (adjusted HR: 0.632
[95% CI 0.073-5.441; P = 0.6758 and adjusted HR 0.492 [95% CI
0.178-1.362; P = 0.1721], respectively) (Abu Esba et al., 2020).
Lastly, a further multicentre, observational study, which collected data
related to 1222 patients with COVID-19 admitted to eight UK hospitals
(of whom 54 were routinely prescribed NSAIDs prior to admission), found
no evidence that routine NSAID use was associated with higher COVID-19
mortality (Bruce et al., 2020).
Other drugs able to reduce the hyperinflammation are currently under
evaluation in patients with COVID-19, including baricitinib,
ruxolitinib, aviptadil and eculizumab. For instance, the effects ofbaricitinib , a Janus kinase (JAK) inhibitor authorized for the
treatment of rheumatoid arthritis and able to suppress the cytokine
storm and to interrupt the passage and intracellular assembly of
SARS-CoV-2 into the target cells (Zhang et al., 2020b), were evaluated
in 12 patients with mild-moderate COVID-19 pneumonia. At week 1 and week
2, the drug demonstrated to improve the clinical and laboratory
parameters and none of the patients required ICU support. Even though
encouraging, this study enrolled a limited number of patients, had no
randomization and no control group (Cantini et al., 2020a). The same
research group carried out an observational, retrospective, multicentre
study in hospitalized patients with moderate COVID-19 pneumonia to
compare the 2-week effectiveness and safety of baricitinib and
lopinavir/ritonavir (n=113) vs. hydroxychloroquine and
lopinavir/ritonavir (n=78). The results showed that the fatality rate
was significantly lower among patients treated with baricitinib (0% vs.
6.4%; p= 0.010). Similarly, ICU admission was significantly lower among
patients treated with baricitinib (0.88% vs. 17.9%) (Cantini et al.,
2020b). The drug was also used in a patient with severe COVID-19 with
insufficient response to lopinavir/ritonavir, hydroxychloroquine,
azithromycin and sarilumab. Authors reported that after the
administration of baricitinib, the patient experienced a prompt
resolution of the respiratory function and improvement in the
radiological findings (Cingolani et al., 2020). Based on data currently
available, no firm conclusion can be drawn on the effects of baricitinib
in COVID-19 patients. In addition, due to a possible increased risk of
herpes zoster and simplex infections, a group of Italian researchers
suggested that the use of baricitinib should be considered with extreme
caution (Favalli et al., 2020). Ruxolitinib is a Janus kinase
(JAK) 1 and JAK2 inhibitor authorized for the treatment of
myeloproliferative malignancies (El Bairi et al., 2020). The effects of
the drug were evaluated in the RUXCOVID study, a phase III, multicentre,
randomised and placebo-controlled trial, but its results are still not
available. Furthermore, the drug is currently evaluated in the
RUXCOVID-DEVENT trial (NCT04377620) in COVID-19 patients with ARDS
requiring mechanical ventilation.Aviptadil is a synthetic vasoactive intestinal polypeptide
(VIP), which increases the activity of adenosine cyclase (Leuchte et
al., 2008). By acting as a weak pulmonary vasodilator, the drug is able
to improve the oxygenation in patients with chronic lung disease
(Ferrara et al., 2005). Currently, many clinical studies are evaluating
the effects of aviptadil as therapeutic or prophylactic agent. For
instance, clinical trials NCT04311697 and NCT04360096 are investigating
the effects of aviptadil on acute lung injury, ARDS, and dyspnoea
associated with COVID-19, while the phase I study NCT04536350 will
evaluate the prophylactic effect of aviptadil for COVID-19 in 80
participants (at this moment all studies are still recruiting
participants)
(www.clinicaltrials.gov).
Lastly, two recent studies highlighted the benefits deriving from the
treatment with eculizumab (Annane et al., 2020; Laurence et
al., 2020). This is a monoclonal antibody approved for the treatment of
atypical haemolytic uremic syndrome, refractory generalized myasthenia
gravis and neuromyelitis spectrum disorders. The drug is able to inhibit
the terminal portion of the complement cascade involved in the
inflammatory response (Gralinski et al., 2018). The study carried out by
Laurence J et al. reported the experience of 3 critical COVID-19
patients who experienced a marked decline in D-dimers and neutrophil
counts after eculizumab treatment (Laurence et al., 2020). The results
of the proof-of-concept study carried out by Annane D et al. showed
that, in a population of 80 patients with severe COVID-19 admitted to
ICU, the estimated survival was 82.9% with eculizumab and 62.2%
without eculizumab (log-rank test, P = 0.04). Based also on the positive
results on rapid decrease in lactate, blood urea nitrogen, bilirubin
levels and a rapid increase in platelet count and prothrombin time
observed with eculizumab, authors suggested that the drug could improve
survival and reduce hypoxia in patients with severe COVID-19 (Annane et
al., 2020). In conclusion, many immunomodulatory and anti-inflammatory
drugs have been tested in patients with COVID-19, but today the evidence
is quite conflicting for most of them. In addition, many of the
published studies are observational or suffer from many limitations,
including the lack of a sample size calculation or control groups and
the use of surrogate endpoints (viral load instead of mortality rate).
At present, the highest number of concluded clinical studies was found
for tocilizumab, hydroxychloroquine and corticosteroids. Among these
drugs, only the use of corticosteroids seems to be supported by robust
evidence, while data related to the efficacy and safety of tocilizumab
and hydroxychloroquine are quite conflicting. Therefore, further data
from randomized controlled trial or well-designed observational studies
are strongly needed.