Scientists employ the knowledge of pathological processes and the immune responses to SARS-CoV-2 to develop clinical therapeutic strategies. The pathogenesis of COVID-19 involves viral replication and over-exuberant inflammatory reactions due to immune dysregulation [2]. Consequently, a cytokine storm syndrome is triggered, releasing large amounts of cytokines and immune cells that infiltrate and destroy organs causing lung lesions, respiratory dysfunction, multiple organ damage, and death [3]. The JAK/STAT signaling pathway, which forms the main regulatory cell signaling pathways, controls the activation of cytokines. Therefore, inhibiting JAK kinase activity and thus preventing STAT activation prevent several cellular responses [4], providing an attractive therapeutic approach for COVID-19 treatment.

The FDA approved Ruxolitinib, a potent JAK1 and JAK2 inhibitor, for Myelofibrosis [5], Polycythemia Vera [6], Hemophagocytic Lymphohistiocytosis [7], and Acute Graft-Versus-Host Disease [8], to reduce the high level of cytokine release associated with these disorders. Preclinical studies have demonstrated that Ruxolitinib could potentially reduce pro-inflammatory cytokine production, including interferon-g (IFN-g) and interleukins, such as IL-2, IL-6, IL-12, and IL-23 [9]. Due to Ruxolitinib’s anti-inflammatory and immunomo- dulatory properties, it is currently being assessed as a potentially curative option for severe COVID-19.

Acute Respiratory Distress Syndrome (ARDS), characterized by refractory hypoxemia, is a major complication in severe cases of COVID-19, affecting 20%-41% of hospitalized patients [10]. It can result in respiratory failure and death. Therefore, whether inhibition of Janus kinases could also reverse severe COVID infection and its associated ARDS became a major point of interest and area of research of several recently published clinical trials and studies on Ruxolitinib (Table 1). A trial conducted by Cao et al. described the results of a prospective, multicenter, single- blinded randomized phase II trial involving 43 patients with severe COVID-19, out of which 22 patients received 5 mg Ruxolitinib twice daily (b.i.d), and 21 were given a placebo based on the current standard of care. Numerically, patients treated with Ruxolitinib showed no significant chest computed tomography improvement (p = 0.495) and faster clinical recovery by day 14, with no mortality in the Ruxolitinib group and no new safety signals recorded [11].

La Rosée et al. [12] conducted a retrospective analysis of patients with severe COVID-19, stratifying them using COVID Inflammation Score (CIS), which involved chest X-ray, laboratory markers of inflammation, and coagulation profile as parameters to identify patients who would benefit from Ruxolitinib treatment. Out of 105 patients, 14 patients with CIS ≥ 10 out of 16 points received Ruxolitinib with a median dose of 7.5 mg b.i.d, resulting in 86% improvement in the clinical course, as indicated by a significant reduction in CIS by day 7 with sustained clinical improvement in 78% without Ruxolitinib-induced toxicity [12].

Likewise, Capochiani et al. [13] treated 18 critically ill patients with COVID-19 related progressive ARDS with 20 mg Ruxolitinib b.i.d for two days followed by a two-step dose de-escalation performed at 10 mg and 5 mg over two weeks. Following this treatment, 16 patients showed notable improvement in respiratory response within 48 hours and did not require mechanical ventilation, and by the end of 2 weeks, they had regained complete respiratory function with no fatalities [13]. Their study provided evidence that a short-term high dose initiating schedule can result in a faster and clinically more relevant response.

Age and associated comorbidities emerged as major risk factors for severe complications and deaths in individuals with COVID-19 [14]. A prospective observational study by Vannucchi et al. (2020) [15] described the compassionate use of Ruxolitinib in 34 patients with severe COVID-19 not requiring mechanical ventilation. The study included patients of advanced age (median age, 80.5 years) and high-risk comorbidities. They observed clinical improvements in 85.3% of patients after treatment with a median dose of 20 mg/day with a median exposure time of 13 days and an overall survival rate of 94.1% by 28 days [15]. Successful treatment with Ruxolitinib was also observed in a case of an elderly patient from a hospital in Germany who had severe COVID-19 associated ARDS requiring invasive ventilation [16].

A recent case report described a hematopoietic stem cell transplant recipient with concomitant chronic Graft Versus Host Disease (cGVHD) suffering from severe COVID-19 who showed clinical improvement after Ruxolitinib treatment and exhibited good tolerance [9]. A similar result was achieved in a 55-year-old man with myelofibrosis and significant comorbidities who developed COVID-19 and recovered without the need for mechanical ventilation [17]. Likewise, Rojas and Sarmiento from Chile reported 2 cases of severe COVID- 19, one with a history of acute lymphoblastic leukemia and the other with multiple myeloma, who responded well to Ruxolitinib 10 mg b.i.d, with repeat chest CT scan showing near-complete regression of the pulmonary infiltrates within a week and inflammatory markers normalizing within two weeks [18, 19].

Table 1
Published clinical trials on Ruxolitinib.

References	Date	Ruxolitinib Dose	Type of Observation	Median Age	Result	Mean Exposure (In Days)	Number of Patients
Cao et al. [11]	26/5/2020	5 mg b.i.d	Prospective, single-blind, randomized controlled phase II trial	63	No statistical difference was observed, but ruxolitinib recipients had a numerically faster clinical improvement with favorable side effect profile.	21-28	20
F.La Rosée et al. [12]	9/6/2020	Started dose of 7.5 mg b.i.d with stepwise dose increase (15mg-0-7.5 mg; 15mg- 0-15mg) at days 3, 5, or 7.	Retrospective analysis	66	Reduction of COVID Inflammation Score (CIS) in 86% of patients with sustained clinical improvement upon treatment with Ruxolitinib.	9	14
Capochiani et al. [13]	4/8/2020	20 mg b.i.d for the first 48 h and subsequent two-step de-escalation at 10 mg b.i.d and 5 mg b.i.d for a maximum of 14 days of treatment.	Multicenter retrospective cohort study	62.5	Evident clinical improvement within 48 hours and Overall Response Rate (ORR) was 89% in severe COVID-19 with ARDS patients by day 14.	14	18
M. Vannucchi et al. [15]	5/8/2020	Starting dose of 5 mg b.i.d; if no improvement, escalating to 10 mg b.i.d after 24–48h; further escalation to 25 mg OD after 48 h.	Prospective Observational Study	80.5	Compassionate use of ruxolitinib was safe and associated with improvement of pulmonary function and overall survival of 94.1%.	13	34

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Table 1
Published clinical trials on Ruxolitinib.

Although the previous studies achieved satisfactory outcomes, few reports showed unexpected results. In a recent case report, two elderly patients with SARS-CoV-2 infection developed severe drug-induced cutaneous reactions and a rapid drop in hematocrit values after compassionate use of Ruxolitinib [20]. Cao et al. reported adverse events in two patients, where one developed transient hypertension while the other developed lymphopenia that improved without interrupting the drug [11]. Negative effects, such as polyomavirus-related fatal encephalopathy [21], meningitis [22], and the emergence of opportunistic infections [2], have also been observed.

Recent research has suggested that dysregulation of the JAK-STAT pathway, particularly IL-6 blockade, is associated with viral persistence, as IL-6 contributes to host defense against infections required for viral clearance [23]. Notably, concerns have been raised about Ruxolitinib unfavorably slowing down viral clearance and production of the SARS- CoV-2 antibody due to its unknown mechanism of SARS-CoV-2 viral clearance. Nevertheless, Cao et al. reported that specific SARS-CoV-2 antibody production, virus clearance, and lymphocyte recovery were similar in both the treatment and control groups [11].

Preliminary research studies and case reports have demonstrated encouraging results that call for further controlled trials to clarify the role of Ruxolitinib in the treatment of COVID-19. Several questions remain to be addressed, including whether Ruxolitinib treatment shows efficacy in severely affected patients with ARDS on invasive ventilation (NCT04362137) and whether treatment with low dose Ruxolitinib (5 mg bid), when compared to placebo, is effective in preventing moderately affected patients from progression to severe/critical disease stages (NCT04362137). Moreover, it is unclear whether laboratory or combined clinical inflammation scores can be used to define optimal timing for initiating medical treatment, as proposed by a currently active trial using the newly developed COVID Inflammation Score (CIS) that is now prospectively being evaluated by the RuxCoFlam trial (NCT04338958). Five registered clinical trials (Table 2) are currently investigating the efficacy and/or safety of Ruxolitinib for COVID-19 (NCT04362137, NCT04414098, NCT04331665).

Table 2
Registered clinical trials to investigate the use of Ruxolitinib in COVID-19.

Registration Number	Official Title	Dose	Study Type & Study Phase	Status
NCT04362137	Phase 3 Randomized, Double-blind, Placebo-controlled Multi-center Study to Assess the Efficacy and Safety of Ruxolitinib in Patients With COVID-19 Associated Cytokine Storm (RUXCOVID)	Ruxolitinib 5mg tablets b.i.d in the experimental group and matching-image placebo in the comparator group for 14 days with possible extension of treatment to 28 days.	Interventional Phase III	Completed October 17, 2020
NCT04338958	A Phase-II Clinical Trial for First Line Treatment of Stage II/III Covid-19 Patients to Treat Hyperinflammation	2x10mg Ruxolitinib with defined response adapted dose escalation up to 2x20mg for a duration of 7 days.	Interventional Phase II	Recruiting August 31, 2021
NCT04359290	Ruxolitinib for Treatment of Covid-19 Induced Lung Injury ARDS A Single-arm, Open-label, Proof of Concept Study	Ruxolitinib will be administered p.o. or by gavage feeding starting with 2x10mg b.i.d at day 1 and can be increased up to 2x15mg b.i.d from day 2 to day 28 (max) (depending on platelet counts and renal function).	Interventional Phase II	Active, not recruiting July 2021
NCT04414098	Safety and Efficacy Study of Ruxolitinib in the Treatment of Severe Acute Respiratory Syndrome Due to SARS-COV-2	Ruxolitinib 5mg orally every 12 hours during 14 days with follow-up time of 45 days.	Interventional Phase II	Not yet recruiting September 15, 2020
NCT04348071	Safety and Efficacy of Ruxolitinib for COVID-19	Participants will receive 10mg ruxolitinib b.i.d.	Interventional Phase II and Phase III	Not yet recruiting October 2021
NCT04361903	Ruxolitinib for the Treatment of Acute Respiratory Distress Syndrome in Patients With COVID-19 Infection	Ruxolitinib oral tablet dosage of at least 2x20mg once daily in the first 48 hours	Observational Cohort	Not yet recruiting May 31, 2020
NCT04331665	A Single Arm Open-label Clinical Study to Investigate the Efficacy and Safety of Ruxolitinib for the Treatment of COVID-19 Pneumonia	Participants will receive ruxolitinib at 10mg, b.i.d, for 14 days, followed by 5mg, b.i.d, for 2 days and 5mg, once daily, for 1 day.	Interventional	Not yet recruiting January 31, 2021
NCT04355793	Expanded Access Program of Ruxolitinib for the Emergency Treatment of Cytokine Storm From COVID-19 Infection	Ruxolitinib starting dose level 5 mg orally, b.i.d.	Expanded Access	Not Available
NCT04377620	A Phase 3, Randomized, Double-Blind, Placebo-Controlled, Multicenter Study to Assess the Efficacy and Safety of Ruxolitinib in Participants With COVID-19-Associated ARDS Who Require Mechanical Ventilation (RUXCOVID-DEVENT)	Arm 1- Placebo + Standard of Care (SoC) Arm 2- Ruxolitinib 5mg + Standard of Care (SoC) Arm 3- Ruxolitinib 15mg + Standard of Care (SoC) Matching Placebo and Ruxolitinib will be administered b.i.d 12 hours apart	Interventional Phase III	Recruiting April 30, 2021
NCT04334044	Treatment of Severe Acute Respiratory Syndrome Caused by COVID-19 With Ruxolitinib	Ruxolitinib 5mg tablet b.i.d	Interventional Phase I and Phase II	Recruiting June 1, 2020
NCT04337359	Ruxolitinib Managed Access Program (MAP) for Patients Diagnosed With Severe/Very Severe COVID-19 Illness	Ruxolitinib 5mg tablet once daily	Expanded Access	Not Available

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Table 2
Registered clinical trials to investigate the use of Ruxolitinib in COVID-19.