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\begin{document}

\title{Combination Therapy of High-dose Intravenous Anakinra and Baricitinib in
Patients with Critical COVID-19: Promising Results from Retrospective
Observational Study}



\author[1]{Murat Bektas}%
\author[1]{Mustafa Ay}%
\author[1]{Muhammed Hamdi Uyar}%
\author[2]{Muhammed İkbal Kılıç}%
\affil[1]{Aksaray Universitesi Tip Fakultesi}%
\affil[2]{Aksaray Universitesi Dahili Tip Bilimleri Bolumu}%


\vspace{-1em}



  
  \date{May 24, 2024}


\begingroup
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\selectlanguage{english}
\begin{abstract}
\textbf{Background:} The leading cause in COVID-19 patients is
development of cytokine storm .~ ~ \textbf{Material and Methods:} This
study was conducted in a tertiary center with diagnosis of COVID-19
patients. Clinical and laboratory features of patients were obtained
from their medical cards and hospital software system and evaluated by
retrospectively.~~~ ~ ~ ~ ~ ~ ~ ~ ~ \textbf{Statistical analysis}: In
our study, 21.0 version (IBM, Armonk, NY, USA) of the SPSS (Statistical
Package for the Social Sciences) program was used for statistical
analysis of data. Descriptive statistics, discrete and continuous
numerical variables were expressed as mean, \selectlanguage{ngerman}± standard deviation or
median and interquartile range (IQR). Categorical variables were
expressed as number of cases and (\%). Cross table statistics were used
to compare categorical variables (Chi-Square, Fisher exact test).
Normally distributed parametric data were compared with Student's t-test
and Paired t-test; non-parametric data that did not meet normal
distribution were compared with Mann Whitney U and Kruskal Wallis tests.
Multiple intergroup comparisons were made by Post Hoc Tukey analysis.
Kaplan-Meier and log-rank methods were used for survival analysis.
Multivariable analysis was performed by using logistic regression.
Correlation analysis was performed with Pearson or Spearman method
according to normality distribution.~ p\textless{}0.05 value was
considered statistically significant. ~ \textbf{Results:} Data of 15
patients in combination group and 43 patients in control group were
evaluated and included into the study. Of these patients 73.3\% was male
in combination arm and 72.1\% in control group (p=0.9). Demographic
findings and frequency of comorbidities were similar between two groups
(table 1). Overall mortality was 46.7\% (n=7) in combination arm and
69.8\% (n=30) in control group although it was not statistically
significant (p=0.1). Similarly, need of intubation was also lower in
combination arm (46.7\%) compared to control group (69.8\%), it was not
significantly different (p=0.1). ICU admission was significantly lower
in combination (46.7\%, n=7) arm than control group (76.7\%, n=33)
(p=0.03, Odds ratio {[}OR{]}:4.7). Development of severe infection
(20\%, n=3 vs 25\%, n=9/36), pulmonary embolism (6.7\%, n=1 vs 0),
myocardial infarction (6.7\%, n=1 vs 2.6\%, n=1/38) and pneumothorax
(13.3\%, n=2 vs 2.6\%, n=1/38) were not different between two groups
(p=0.7, p=0.3, p=0.5 and p=0.2). In multivariable analysis only cHIS
score was associated with high mortality (p=0.018, OR:2.8, {[}95\%
confidence interval: 1.2-6.6{]}). In survival analysis, mortality rate
was significantly lower in combination arm than control group
(Log-Rank:p=0.04;figure 1). In conclusion, combination therapy of
high-dose anakinra and baricitinib may be an adequate treatment option
in patients with COVID-19 who had critical disease and no additional
safety signal.%
\end{abstract}\selectlanguage{ngerman}%



\sloppy


\textbf{Short communication}

\textbf{Title Page:}



\textbf{Combination Therapy of High-dose Intravenous Anakinra and
Baricitinib in Patients with Critical COVID-19: Promising Results from
Retrospective Observational Study}

\textbf{Running Head:}

\textbf{Combination of Anakinra and Baricitinib in COVID-19}

1. Murat Bekta\selectlanguage{polish}ş, \selectlanguage{english}MD

Division of Rheumatology, Department of Internal Medicine, Aksaray
Training and Research Hospital, Aksaray, Istanbul Ayd\selectlanguage{polish}ı\selectlanguage{english}n University,
Turkey

Postal Address: Yeni Sanayi Mahallesi, 68200 Merkez/AKSARAY, TURKEY

E-post: murat.b88@hotmail.com

ORCID ID: 0000-0002-188-3837

2-Mustafa Ay, MD

Aksaray University, Aksaray Training and Research Hospital, Aksaray,
Turkey

Postal Address: Yeni Sanayi Mahallesi, 68200 Merkez/AKSARAY, TURKEY

E-post: mustafaay666@gmail.com

3. Muhammed Hamdi Uyar, MD

Aksaray University, Aksaray Training and Research Hospital, Aksaray,
Turkey

Postal Address: Yeni Sanayi Mahallesi, 68200 Merkez/AKSARAY, TURKEY

E-post: hamdiuyar68@gmail.com

4. Muhammed \selectlanguage{polish}İ\selectlanguage{english}kbal K\selectlanguage{polish}ı\selectlanguage{english}l\selectlanguage{polish}ı\selectlanguage{ngerman}ç, MD

Department of Internal Medicine, Aksaray Training and Research Hospital,
Aksaray, Turkey

Postal Address: Yeni Sanayi Mahallesi, 68200 Merkez/AKSARAY, TURKEY

E-post: dr.muhammedkilic@gmail.com

\emph{Corresponding author}\textbf{:}

Murat Bekta\selectlanguage{polish}ş, \selectlanguage{english}MD

Division of Rheumatology, Department of Internal Medicine,

Aksaray Training and Research Hospital, Aksaray,

Istanbul Ayd\selectlanguage{polish}ı\selectlanguage{english}n University, Turkey

Postal Address: Yeni Sanayi Mahallesi, 68200 Merkez/AKSARAY, TURKEY

E-post: murat.b88@hotmail.com

ORCID ID: 0000-0002-188-383

\textbf{DECLARATIONS}

\textbf{Funding:} No specific funding was received from any bodies in
the public, commercial or not-for-profit sectors to carry out work
described in this article.

\textbf{Conflicts of Interest:} The authors declare no conflicts of
interest.

\textbf{Availability of data and material:} The Dataset of the study is
available from the corresponding author on reasonable request.

\textbf{Code availability:} Not applicable.

\textbf{Author contributions:} MB designed and planned the study, MB,
MA, MHU, and M\selectlanguage{polish}İ\selectlanguage{english}K collected the data, M\selectlanguage{polish}İ\selectlanguage{english}K and MB carried out the data
evaluation and basic analyses, all authors contributed to the follow-up
of the patients and interpretation of results. MB wrote the first draft,
and all authors provided critical feedback for the last version.

\textbf{Ethics approval} : Individual written patient consent and local
ethic committee approval was obtained for this study (date/number:
24.02.2022, 2022/04-09).

\textbf{Word count:} 647 words in main text without references

\textbf{Abstract word count} : Not applicable

\textbf{Number of tables} : One

\textbf{Number of figures:} One

\textbf{References:} 8

\textbf{Acknowledgement:} No

\textbf{Key words:} Anakinra, Baricitinib, COVID-19, cytokine storm

\textbf{Background}

Coronavirus disease 2019 (COVID-19) is an emerging infectious disease
caused by SARS-Cov-2. The leading cause in COVID-19 patients is
development of cytokine storm (1). Several cytokines such as IL-1,
TNF-\selectlanguage{greek}α, \selectlanguage{english}IL-6, and interferon (IFN) have important role into the
development of COVID-19 associated cytokine storm (1). Several targeted
therapies such as anakinra, tocilizumab, and janus kinase (JAK)
inhibitors were found to be effective in patients with COVID-19
associated cytokine storm (2) (3) (4). However, there is limited
evidence with combination of targeted therapies in patients with
COVID-19.

\textbf{Material and Methods}

\textbf{Patients and data:} This study was conducted in a tertiary
center with diagnosis of COVID-19 patients. Study population consisted
of patients with positive polymerase chain reaction and computer
tomography findings compatible with COVID-19 as well as critical illness
requiring high flow nasal oxygen supplement, or its equivalent defined
according to National Institute of Health (NIH) COVID-19 disease
severity scale (5). Clinical and laboratory features of patients were
obtained from their medical cards and hospital software system and
evaluated by retrospectively. All laboratory results were recorded at
patients' admission and inflammatory parameters such as C-reactive
protein (CRP), ferritin, d-dimer, lactate dehydrogenase (LDH) also on
the highest levels and last day of hospitalization. Hyperinflammatory
state of the patients were evaluated and scored according to COVID-19
hyperinflammatory syndrome score (cHIS score) (6).

\textbf{Treatment protocol and outcome:} All patients received
background glucocorticoid treatment with methylprednisolone 80 mg/day
intravenously or equivalent concomitant anticoagulant prophylaxis with
enoxaparin 0.4 mg/day subcutaneously. Anakinra was initiated in patients
who did not respond to two days of corticosteroid treatment or
concomitantly in high risky patients with 400 mg/day intravenously in
two divided dose and gradually increased to 1600 mg/day if necessary.
The dose adjustment was performed by daily for each patient according to
current clinical and laboratory findings by the same rheumatologist
(MB). In combination group, baricitinib was started in 8 mg/day in two
divided dose (4 mg/day in chronic renal failure) in addition to steroid
and anakinra in patients with unresponsive to anakinra 1600 mg/day at
least three days, thereafter dose reduction was performed according to
clinical and laboratory results of the patients. Drug tapering was
performed in anakinra doses initially when clinical and laboratory
response achieved thereafter treatment was stopped when the patients
were oxygen free and/or had normal inflammatory parameters such as CRP
and ferritin. Treatment failure was defined as no reduction in oxygen
support and/or inflammatory parameters as well as need for intensive
care unit (ICU) admission and/or invasive mechanical ventilation and/or
development of mortality. Age, gender, disease severity (NIH score 4,
critically ill patients) and inflammatory burden according to cHIS score
matched patients who receiving steroid and anakinra defined as control
group. Steroid and anakinra treatment protocol were similar between two
groups. Individual patient consent and ethical committee approval were
obtained for this study (date/number: 24.02.2022, 2022/04-09).

\textbf{Statistical analysis} : In our study, 21.0 version (IBM, Armonk,
NY, USA) of the SPSS (Statistical Package for the Social Sciences)
program was used for statistical analysis of data. Descriptive
statistics, discrete and continuous numerical variables were expressed
as mean, \selectlanguage{ngerman}± standard deviation or median and interquartile range (IQR).
Categorical variables were expressed as number of cases and (\%). Cross
table statistics were used to compare categorical variables (Chi-Square,
Fisher exact test). Normally distributed parametric data were compared
with Student's t-test and Paired t-test; non-parametric data that did
not meet normal distribution were compared with Mann Whitney U and
Kruskal Wallis tests. Multiple intergroup comparisons were made by Post
Hoc Tukey analysis.\selectlanguage{english} \selectlanguage{english}Kaplan-Meier and log-rank methods were used for
survival analysis. Multivariable analysis was performed by using
logistic regression. Correlation analysis was performed with Pearson or
Spearman method according to normality distribution.~ p\textless{}0.05
value was considered statistically significant.

\textbf{Results}

Data of 15 patients in combination group and 43 patients in control
group were evaluated and included into the study. Of these patients
73.3\% was male in combination arm and 72.1\% in control group (p=0.9).
Demographic findings and frequency of comorbidities were similar between
two groups (table 1). Overall mortality was 46.7\% (n=7) in combination
arm and 69.8\% (n=30) in control group although it was not statistically
significant (p=0.1). Similarly, need of intubation was also lower in
combination arm (46.7\%) compared to control group (69.8\%), it was not
significantly different (p=0.1). ICU admission was significantly lower
in combination (46.7\%, n=7) arm than control group (76.7\%, n=33)
(p=0.03, Odds ratio {[}OR{]}:4.7). Development of severe infection
(20\%, n=3 vs 25\%, n=9/36), pulmonary embolism (6.7\%, n=1 vs 0),
myocardial infarction (6.7\%, n=1 vs 2.6\%, n=1/38) and pneumothorax
(13.3\%, n=2 vs 2.6\%, n=1/38) were not different between two groups
(p=0.7, p=0.3, p=0.5 and p=0.2). In multivariable analysis only cHIS
score was associated with high mortality (p=0.018, OR:2.8, {[}95\%
confidence interval: 1.2-6.6{]}). In survival analysis, mortality rate
was significantly lower in combination arm than control group
(Log-Rank:p=0.04;figure 1).

\textbf{Discussion}

Baricitinib, is a selective JAK1 inhibitor, decreases production of
several proinflammatory cytokines including IL-2, IL-6, IL-7, TNF-\selectlanguage{greek}α, \selectlanguage{english}and
type 1 IFNs (7). Although baricitinib decreases many cytokine levels as
mentioned above it does not affect to IL-1 levels. High-dose intravenous
anakinra treatment was found to be safe and effective in patients with
severe COVID-19 in our previous study (8). In the current study we added
baricitinib to the extremely ill COVID-19 patients who had refractory to
both steroids and high-dose anakinra. The rationale of the current
protocol is more effective suppression of cytokine production in
patients had extremely higher mortality. Our study emphasizes no
additional safety signal including severe infection, thrombosis, and
cardiovascular events with combination group. Furthermore, combination
therapy tended to reduce mortality in patients with critical COVID-19 in
our study. We emphasize that in this study, combination therapy was
applied only to critically ill patients and did not include milder
disease therefore this finding may not be generalized to milder cases.

In conclusion, combination therapy of high-dose anakinra and baricitinib
may be an adequate treatment option in patients with COVID-19 who had
critical disease and no additional safety signal.

\textbf{Table 1} : Clinical and laboratory features and outcomes of
study participants\selectlanguage{english}
\begin{longtable}[]{@{}llll@{}}
\toprule
Variables & Combination group (n=15) & Control group (n=43) & p value
(OR)\tabularnewline
\midrule
\endhead
Age, years, mean\selectlanguage{ngerman}±SD & 63.3\selectlanguage{ngerman}±9 & 65.7\selectlanguage{ngerman}±15 & 0.6\tabularnewline
Gender, male, n (\%) & 11 (73.3) & 31 (72.1) & 0.9\tabularnewline
Duration of hospitalization (days), median (IQR) & 19 (11) & 15 (14) &
0.02\tabularnewline
Admission to ICU interval (days), median (IQR) & 14.5 (22) & 6 (7) &
\textless{}0.001\tabularnewline
Comorbidities, n (\%) & & &\tabularnewline
Diabetes mellitus & 6 (40) & 8/41 (19.5) & 0.2\tabularnewline
Hypertension & 7 (46.7) & 19/40 (47.5) & 1\tabularnewline
Coronary heart disease & 3 (20) & 6/40 (15) & 0.7\tabularnewline
Heart failure & 1 (6.7) & 4/40 (10) & 0.7\tabularnewline
Chronic renal failure & 3 (20) & 9 (21) & 1\tabularnewline
Chronic obstructive pulmonary disease & 3 (20) & 9/41 (22) &
0.9\tabularnewline
Malignancy & 1 (6.7) & 6/42 (14.3) & 0.7\tabularnewline
Rheumatic disease & 2 (13.3) & 3/40 (7.5) & 0.6\tabularnewline
Immunosuppressive drug usage & 3 (20) & 6 (14) & 0.7\tabularnewline
cHIS Score, mean\selectlanguage{ngerman}±SD & 4\selectlanguage{ngerman}±1 & 4\selectlanguage{ngerman}±0.9 & 0.9\tabularnewline
Laboratory results, median (IQR) & & &\tabularnewline
NLR & 6.7 (9.2) & 9 (10) & 0.5\tabularnewline
CRP (mg/L) & & &\tabularnewline
Admission & 161 (153) & 146 (141) & 0.6\tabularnewline
Max & 163 (173) & 173 (138) & 0.8\tabularnewline
Last & 30 (218) & 67 (153) & 0.7\tabularnewline
Ferritin (pg/mL) & & &\tabularnewline
Admission & 334 (588) & 492 (575) & 0.4\tabularnewline
Max & 727 (1410) & 1032 (1961) & 0.3\tabularnewline
Last & 554 (1528) & 531 (1449) & 0.7\tabularnewline
D-dimer (pg/mL) & & &\tabularnewline
Admission & 0.89 (0.65) & 1.4 (1.1) & 0.006\tabularnewline
Max & 23 (17) & 12.3 (20) & 0.2\tabularnewline
Last & 8.4 (14) & 5 (16.7) & 0.9\tabularnewline
LDH (U/L) & & &\tabularnewline
Admission & 348 (328) & 447 (274) & 0.5\tabularnewline
Max & 676 (376) & 728 (466) & 0.6\tabularnewline
Last & 486 (396) & 548 (507) & 0.5\tabularnewline
Outcomes, n (\%) & & &\tabularnewline
Severe infection & 3 (20) & 9/36 (25) & 0.7\tabularnewline
Pneumothorax & 2 (13.3) & 1/38 (2.6) & 0.2\tabularnewline
Pulmonary thromboembolism & 1 (6.7) & 0 & 0.3\tabularnewline
Myocardial infarction & 1 (6.7) & 1/38 (2.6) & 0.5\tabularnewline
ICU admission & 7 (46.7) & 33 (76.7) & 0.03 (4.7)\tabularnewline
Intubation & 7 (46.7) & 30 (69.8) & 0.1\tabularnewline
Mortality & 7 (46.7) & 30 (69.8) & 0.1\tabularnewline
\bottomrule
\end{longtable}

SD: standard deviation, IQR: Interquartile range, ICU: Intensive care
unit, NLR: Neutrophile lymphocyte ratio

\textbf{Figure 1} : Comparison of mortality rate between patients with
combination and control arm (Kaplan-Meier analysis)\selectlanguage{english}
\begin{figure}[H]
\begin{center}
\includegraphics[width=0.70\columnwidth]{figures/image1/image1}

\end{center}
\end{figure}

Log-Rank: p=0.04

\textbf{References}

1. Ye Q, Wang B, Mao J. The pathogenesis and treatment of the `Cytokine
Storm' in COVID-19. The Journal of infection. 2020;80(6):607-13.

2. Tocilizumab in patients admitted to hospital with COVID-19
(RECOVERY): a randomised, controlled, open-label, platform trial. Lancet
(London, England). 2021;397(10285):1637-45.

3. Kyriazopoulou E, Poulakou G, Milionis H, Metallidis S, Adamis G,
Tsiakos K, et al. Early treatment of COVID-19 with anakinra guided by
soluble urokinase plasminogen receptor plasma levels: a double-blind,
randomized controlled phase 3 trial. Nature medicine.
2021;27(10):1752-60.

4. Baricitinib in patients admitted to hospital with COVID-19
(RECOVERY): a randomised, controlled, open-label, platform trial and
updated meta-analysis. Lancet (London, England). 2022;400(10349):359-68.

5. COVID-19 Treatment Guidelines Panel. Coronavirus Disease 2019
(COVID-19) Treatment Guidelines. 2022 {[}Available
from:\emph{https://www.covid19treatmentguidelines.nih.gov/}. .

6. Webb BJ, Peltan ID, Jensen P, Hoda D, Hunter B, Silver A, et al.
Clinical criteria for COVID-19-associated hyperinflammatory syndrome: a
cohort study. The Lancet Rheumatology. 2020;2(12):e754-e63.

7. Assadiasl S, Fatahi Y, Mosharmovahed B, Mohebbi B, Nicknam MH.
Baricitinib: From Rheumatoid Arthritis to COVID-19. Journal of clinical
pharmacology. 2021;61(10):1274-85.

8. Bekta\selectlanguage{polish}ş \selectlanguage{english}M, Y\selectlanguage{ngerman}üce S, Ay M, Uyar MH, Önder ME, K\selectlanguage{polish}ı\selectlanguage{english}l\selectlanguage{polish}ı\selectlanguage{ngerman}ç M. High-dose
intravenous anakinra treatment is safe and effective in severe and
critical COVID-19 patients: a propensity score-matched study in a single
center. Inflammopharmacology. 2023;31(2):787-97.

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