Study protocol for assessment of the efficacy of calcium dobesilate versus placebo on SARS-CoV-2 viral load in outpatients with COVID-19 (CADOVID study): a randomised, placebo-controlled, double-blind, monocentric phase II trial

Abstract

Introduction

SARS-CoV-2 mainly infects respiratory endothelial cells, which is facilitated through its spike protein binding to heparan sulphate. Calcium dobesilate (CaD) is a well-established, widely available vasoactive and angioprotective drug interacting with heparan sulphate, with the potential to interfere with the uptake of SARS-CoV-2 by epithelial cells. The CADOVID trial aims to evaluate the efficacy and safety of CaD in reducing the SARS-CoV-2 viral load in non-hospitalised adult patients diagnosed with COVID-19, confirmed by a positive SARS-CoV-2 PCR, including its efficacy to reduce the impact of persistent COVID-19 symptoms.

Methods and analysis

This is a randomised, placebo-controlled, double-blind, monocentric phase II trial. Enrolment began in July 2022. A total of 74 adult patients will be randomly allocated to the CaD arm or the placebo group with a 1:1 ratio, respectively. Participants in the intervention arm will receive two capsules of CaD 500 mg two times per day and the placebo arm will receive two matching capsules of mannitol 312.5 mg two times per day, with a treatment period of 7 days for both arms, followed by a 77-day observational period without treatment administration. Participants will be asked to complete secured online questionnaires using their personal smartphone or other electronic device. These include a COVID-19 questionnaire (assessing symptoms, temperature measurement, reporting of concomitant medication and adverse events), a COVID-19 persistent symptoms’ questionnaire and the Short Form 12-Item (SF-12) survey. SARS-CoV-2 PCR testing will be performed on nasopharyngeal swabs collected on days 1, 4, 8 and 21. The primary endpoint is the reduction from baseline of SARS-CoV-2 viral load determined by RT-PCR at day 4.

Ethics and dissemination

This trial has received approval by the Geneva Regional Research Ethics Committee (2022-00613) and Swissmedic (701339). Dissemination of results will be through presentations at scientific conferences and publication in scientific journals.

Trial registration number

NCT05305508; Clinicaltrials.gov; Swiss National Clinical Portal Registry (SNCTP 000004938).

STRENGTHS AND LIMITATIONS OF THIS STUDY.

• This randomised, placebo-controlled, double-blind, monocentric phase II trial is designed to evaluate SARS-CoV-2 viral load reduction at an early time point (day 4) and at days 8 and 21.

• The study design allows to considerably reduce bias in the assessments of treatment efficacy.

• Participant recruitment into the study will depend on the COVID-19 testing rate. With the current decrease in the number of COVID-19 tests performed due to national testing policies, the number of potential participants is reduced proportionally and will affect the duration of study recruitment.

• Patients at a higher risk of disease progression and with an indication for treatment by commercialised therapies (eg, nirmatrelvir/ritonavir) are not proposed participation in the study, thus restricting the study population.

Introduction

The COVID-19 pandemic caused by the SARS-CoV-2 virus resulted in huge worldwide morbidity and mortality, including some long-lasting symptoms known as ‘post-COVID-19 condition’.¹ The viral attachment of SARS-CoV-2 and infection of host cells have been shown to be dependent on heparan sulphate binding to angiotensin-converting enzyme 2.^{2 3} Calcium dobesilate (CaD) is known to interfere with heparan sulphate binding of vascular endothelial growth factor (VEGF).⁴ Preclinical results using lentiviral pseudotyped particles demonstrated that CaD interferes with the binding of SARS-CoV-2 spike protein to heparan sulphate and significantly reduced its uptake in cultured endothelial cells.⁵ By contrast, CaD effects on cell infection were not mediated by cell toxicity or the inhibition of endocytosis.

Although SARS-CoV-2 infects pulmonary epithelial cells, it may also infect other cell types, particularly endothelial, thus affecting multiple organs, apart from the lungs.⁶ Indeed, patients with chronic comorbidities with an impaired endothelium (eg, obesity, hypertension, diabetes, cardiovascular/cerebrovascular disease, chronic respiratory disease, malignancy) were shown to be at a particularly higher risk of severe or fatal COVID-19 disease as SARS-CoV-2 infection may aggravate the underlying endothelial dysfunction.^7–10 This latter hypothesis provides a rationale for assessing COVID-19 therapies that stabilise the endothelium.

CaD is a well-established vasoactive and angioprotective drug improving endothelial dysfunction^{11 12} with a good safety tolerability profile.^{13 14} Based on clinical data and real-world evidence, CaD was shown to result in relatively few adverse events, many of which are benign and reversible on treatment discontinuation.¹⁵ Its use is indicated for the treatment of microangiopathies, chronic venous insufficiency and haemorrhoidal disease^{15 16} as it strengthens vessel integrity and improves blood flow. CaD has been proposed to play a potential key role in COVID-19 pathophysiology through the inhibition of VEGF/fibroblast growth factor pathways,^{4 17–21} which are highlighted as potential targets for COVID-19 infection and its effects.^17–22 Thus, the use of CaD as a potent inhibitor of these pathways may potentially contribute to reduce SARS-CoV-2 cell infection and subsequent inflammation as well as to protect capillary barrier permeability and alleviate thrombosis and perivascular oedema.¹⁷

To determine whether CaD acts in the tissues or sites of interest, we conducted a phase I study to collect pharmacokinetic data. We assessed the bioavailability of CaD in the nasal tissue and saliva after oral intake in humans requiring this treatment. The study showed a CaD presence in nasal mucosa and saliva, thus demonstrating that the drug is present in the tissues where it would need to exert its potential anti-SARS-CoV-2 activity (unpublished data, protocol CCER-2020-03050). As the first contact with SARS-CoV-2 mostly occurs in the nasal mucosae,^23–25 CaD would be able to exert its potential antiviral activity on the first stages of infection.

Therefore, we hypothesised that oral CaD treatment in the early symptomatic phase may represent a safe COVID-19 treatment. Compared with other tested treatments, such as seawater which only acts locally,²³ CaD has the potential of acting both in the initial stage of infection through its antiviral activity, and in the later stages on endothelial inflammation/dysfunction, to prevent severe effects of SARS-CoV-2 infection and subsequent long-term sequelae.

The CADOVID study will evaluate the efficacy and safety of CaD in reducing SARS-CoV-2 viral load (VL) in non-hospitalised adult patients diagnosed with COVID-19, documented with a positive SARS-CoV-2 PCR, and the occurrence of COVID-19 symptoms. The efficacy of CaD to reduce the impact of persistent COVID-19 symptoms will also be evaluated.

Methods

Study design

This phase II, randomised, double-blind, placebo-controlled, monocentric trial will be conducted to assess the efficacy and safety of CaD compared with placebo in reducing disease duration in adult patients diagnosed with COVID-19. As there are currently no oral treatments available in Switzerland for patients with mild COVID-19 and who do not require hospitalisation, a placebo comparator was chosen for this study. The overall study design and study population is shown in figure 1. The study protocol (V.3.0 of 7 July 2022) was approved by the Geneva Regional Research Ethics Committee (2022-00613) and Swissmedic (701339) and registered at ClinicalTrials.gov and the Swiss National Clinical Portal Registry. Written informed consent will be obtained from all patients by the principal investigator or authorised delegate. During the screening procedures, participants may independently consent to additional procedures called future biomedical research (FBR) and including nasal (or buccal) brush and blood sample collection. We report the study protocol according to the Standard Protocol Items: Recommendations for interventional Trials statement.

Figure 1.

Study design and study population.

Enrolled participants will be randomised to either the intervention arm or the placebo arm in a 1:1 ratio, respectively. Participants in the intervention arm will receive two capsules of CaD 500 mg two times per day (total of 2000 mg/day) and the placebo arm will receive two capsules of placebo (312,5 mg mannitol) two times per day (total of 1250 mg/day). The treatment period is 7 days in both arms, followed by a 77-day observational period without treatment administration.

Patient and public Involvement

None.

Study population

The population will comprise symptomatic COVID-19 individuals aged ≥16 years of age, with documented SARS-CoV-2 positivity as assessed by PCR. Study recruitment will take place at Geneva University Hospitals (Geneva, Switzerland) and other COVID-19 test centres in the canton of Geneva. Study visits and follow-up will be performed only at Geneva University Hospitals. Inclusion and exclusion criteria of the study population are presented in table 1.

Table 1.

Inclusion and exclusion criteria

Inclusion criteria	Exclusion criteria
1. Documented COVID-19 diagnosis (SARS-CoV-2 positivity as assessed by PCR) within≤3 days of symptom appearance, with a cycle threshold (CT) <25.	1. Known hypersensitivity or allergy to any of the study products to be administered.
2. Symptoms related to day 1≤5 days.	2. Participation in any other investigational device or drug study within 30 days preceding the study screening visit.
3. Presents at least one of the following acute COVID-19 symptoms: nasal congestion or runny nose; sore throat; headache; myalgia; dry/productive cough; fever; chills; abdominal symptoms (nausea, vomiting, diarrhoea, abdominal pain); fatigue; chest pain; palpitations; and shortness of breath.	3. Treatment with CaD or related molecules (eg, etamsylate) within 30 days preceding screening visit or current treatment with any other investigational agent(s).
4. Aged≥16 years.	4. Breastfeeding, unless patient agrees to stop breastfeeding.
5. Participant has provided signed informed consent.	5. Treatment with any investigational, emergency approved-use or approved drug for COVID-19 (eg, monoclonal antibody treatment, direct or indirect antiviral treatment, and others according to local guidelines).
	6. Any kind of condition that may compromise the capacity for discernment and consent (eg, dementia), and any other conditions (eg, residing abroad or far from the investigation site) that may be associated with an increased risk to the participant or may affect compliance or may interfere with study assessments or outcomes.
	7. Any hospitalisation criteria according to local guidelines (oxygen saturation<95%, respiratory rate>25) at the time of screening; or signs that the patient’s health may deteriorate in the next 24–48 hours and require hospitalisation according to the investigator’s judgement based on a clinical evaluation (eg, association of critical symptoms: chest pain/difficulty to breath).

CaD, calcium dobesilate.

Randomisation and blinding

Randomisation will be done in variable-sized blocks (size 4) in random sequence. Randomisation will be carried out by the pharmacist preparing the blinded vials using a computer-generated table. Participants will be randomised in a 1:1 ratio to either the CaD or placebo at day 1 (baseline). Participants, investigators and the sponsor or delegates involved in study procedures or outcome assessments will be blinded to the randomisation and group allocation. Blinding will be maintained throughout the entire study period, including the observational period. A request for unblinding of treatment assignment may be made to the pharmacist by the principal investigator only in the setting where the knowledge of the blinded treatment assignment is necessary in decisions about proper subsequent treatments of that participant. In such cases, the sponsor will be immediately informed by the investigator.

Each participant will be assigned a unique identifier on inclusion in the study. This number will serve as the participant’s identifier in the study database. All participant records or data sets transferred to the sponsor will contain the identifier only; participant names or any identifiable information will not be transferred. Only the investigator or designee will be able to link study data to an individual participant via an identification list kept securely at the site.

All study data will be entered in an Electronic Data Capture (EDC) system, REDCap, by the authorised study site team. Case Report Forms (CRFs) will be kept current to reflect subject status during the study. The obtained data will be handled strictly confidential. Personal data will be anonymised for data analysis.

Procedures

At day 1 visit, all required doses of blinded CaD/placebo will be given to the participant for self-administration during the study. However, the first dose is taken as a directly observed therapy at the time of this visit.

During the study, participants will be asked via email notifications to complete online questionnaires using their personal smartphone or other electronic personal device. Data entered by the participant will be directly imported into the study database (REDCap) and evaluated on a daily basis by the study team. Alerts will be automatically sent to the study team in the case of hospitalisation, treatment compliance issues, severe symptoms (chest pain and/or shortness of breath) and if ≥3 notified symptoms. Follow-up with participants by phone will be determined on a case-by-case basis, depending on symptom occurrence, severity and persistence. Safety events will be reported to the Ethics committee and Swissmedic according to the applicable guidelines.

Participants will be withdrawn from the study if there are any clinical reasons judged by the investigator to be life threatening or if the study procedures must be stopped due to safety concerns. Participants will be withdrawn if they revoke their study consent or if they are at significant risk of failing to comply with the provisions of the protocol. Withdrawn participants during the first 4 days will be replaced to achieve the total sample size required for primary analysis.

Outcomes

Primary endpoint

The primary endpoint is the reduction of the SARS-CoV-2 VL as determined by RT-PCR between baseline and day 4.

Secondary endpoints

1. SARS-CoV-2 VL: (1) reduction from baseline of the SARS-CoV-2 VL as determined by RT-PCR at day 8 and (2) the proportion of patients with VL negativity or a very low VL (defined by PCR threshold cycles >32) at days 4, 8 and 21.

2. Symptoms: (1) time to acute symptom resolution (listed in table 2, letter d) after randomisation to treatment and (2) proportion of participants with acute symptom resolution* at days 4, 8, 21.

3. Persistent COVID-19 symptoms: (1) proportion of patients with persistent symptoms (≥1 of the persistent symptoms listed in table 2) at day 84 (week 12) and (2) SF12 score at days 21 and 84 (week 12) compared with day 1.

Table 2.

Study procedures

Visit	Screening *	Baseline *	Day 4	Day 8	Day 21	Day 84	ET †
Details	On-site	On-site	On-site	On-site	On-site	Phone call	On-site
Time (day) window	1 –	1 –	4 –	8 –	21 ± 3 days	84 ± 7 days	–
Procedures
Informed consent	X
Eligibility criteria check	X
Randomisation		X
COVID-19 symptom assessment	X
SARS-CoV-2 PCR ‡ (nasopharyngeal swab)		X	X	X	X		X
Online COVID-19 questionnaire§		X	X	X	X	X
Online persistent COVID-19 symptoms questionnaire¶		X			X	X
Online SF-12 survey**		X			X	X	X
Self-reported temperature		X	X	X	X
Concomitant medications and adverse event/s review††	X		X	X	X	X	X
Blood sample collection (FBR) ‡‡		X	X		X		X
Nasal brush (or buccal brush) ‡‡		X	X		X		X

*Screening and baseline (day 1) visits will be performed during the same day.

†Participants who discontinue the study during the treatment and short-term follow-up periods will be asked to return to site for an early termination visit. Participants who discontinue the study after day 21 will not be required to return to an on-site early termination visit.

‡Depending on the duration of the recruitment phase, the identification of the SARS-CoV-2 variant by sequencing could be carried out.

§The COVID-19 questionnaire will be administered once daily during the first 21 days and once weekly from weeks 4 to 12. The questionnaire includes symptom assessment (see below §§), temperature measurement, and reporting of concomitant medications and adverse events. A study medication compliance check is included during the first 7 days (treatment period).

¶Persistent COVID-19 symptom assessment: fatigue; headache; intermittent fever; palpitations/tachycardia; sleep disturbance; anxiety; blurred vision; depression; brain fog (difficulty concentrating); loss of memory; dizziness; tinnitus (and other hearing issues); altered smell; altered taste; shortness of breath; chest pain; cough; myalgia (and spasms); neuralgias; arthralgia (joint pain); paraesthesia; nausea; vomiting; diarrhoea; constipation; abdominal pain; menstrual and period problems; as well as new onset of allergies.

**The SF-12 Survey is a self-reported measure of physical and mental health status. The SF-12 is often used as a measure of a person or population’s quality of life. It comprises 12 questions that cover various health domains, including limitations of physical and social activities, body pain, mental health, vitality and general health perception.

††There are no prohibited medications; participants will be allowed to continue with their chronic illness medications. However, they will be advised to limit to the maximum extent possible the intake of paracetamol or ibuprofen to reduce any impact on the secondary outcomes.

‡‡Nasal brush (or if medically not indicated, can be substituted by buccal brush) and blood sample collection are optional and are performed under a separate patient consent (FBR).To ensure quality of study conduct and study data retrieval, all study sites will have regular monitoring activities performed by appropriately trained and qualified monitors. Monitoring activities will consist of on-site monitoring visits. Prior to study start (prior to first participant enrolled), a central monitoring plan will be developed detailing all monitoring-related procedures and activities planned during the study. The study documentation, the source data/documents and study-related reports will be accessible to auditors or inspectors and regulatory authorities in case of an audit or an inspection.

§§Nasal congestion or runny nose, sore throat, dry or productive cough, shortness of breath, chest pain, muscle or body aches, fatigue, headache, palpitations, chills/sweats, feeling hot or feverish, nausea, vomiting, abdominal pain, diarrhoea, decrease or loss of smell, decrease or loss of taste.

FBR, future biomedical research; SF-12, Short Form 12-Item.

Safety endpoints

The study will aim to evaluate the long-term safety of CaD compared with placebo in patients with COVID-19 as assessed by the incidence and severity of adverse events.

Exploratory endpoints

During the screening procedures, we will propose to participants to independently consent to additional procedures (FBR). The aims of this additional research are:

1. To identify CaD-induced changes on nasal (or buccal) epithelial cells;

2. To assess the effect of CaD on candidate blood biomarkers of severe COVID-19 and persistent COVID-19 symptoms compared with placebo.

To that end, nasal (or buccal) epithelial cells will be collected for transcriptome analysis. Nasal brushes (or buccal swab) will be stored at −80°C until shipment to an external laboratory for analysis.

Blood sampling (up to 14 mL of peripheral venous blood) will be collected for analysis: D-dimers, von Willebrand factor, fibrinogen, CRP and ferritin dosage, dosage of protein biomarkers including, but not limited to IFN-β, PTX3, IFN-γ IFN-λ2/3, IL6, VEGF-D, VEGF-A, sTIM-3, galectin-3m and for whole blood transcriptome analysis. These samples will be stored at −80°C until shipment to an external laboratory for analysis.

Statistical considerations

Sample size calculation

To detect a difference of 1 log (or three cycle thresholds (CT)) in the mean reduction of SARS-CoV-2 VL determined by RT-PCR at day 4, with a 80% power, a one-sided significance level of 2.5% and assuming an SD of 1.5 log (difference to detect of 0.67 SD), calculated sample size is 37 in each arm, total of 74 patients. Patients for whom the main outcome will not be completed (VL at day 4) will be replaced.

Primary endpoint analysis

For the analysis of the primary endpoint, we will perform an intention to treat analysis, including all patients who were randomised and who have received at least one dose of the study medication.

CT values obtained from the SARS-CoV-2 PCR will be log transformed to obtain VL values (log IU/mL). CT values will be transformed into VL values using the formulation: VL= (CT–43.324)/–3.1486 based on linear regression analysis.

The difference between baseline and day four will be obtained for each participant. The difference in means (of the delta over time) and a 95% CI will be first obtained via a fitted linear model. Then, depending on distribution (symmetrical/non-symmetrical), we will use parametric (t-test for normal distribution) or non-parametric tests (Mann-Whitney for non-symmetrical distribution) to compare the intervention arm with the placebo arm.

No stratification will be applied in the primary analysis. Statistical adjustment will be done using a linear regression model based on immunity (vaccination and prior infection), immunosuppression and age variables:

VL (day 4)=b0+b1*VL (day 0)+b2*treatment+b3*immunity+b4*immunosuppression+b5*age

Compliance with the study intervention (CaD/placebo) will be verified through online questionnaires, and at the end of the treatment period, based on returned CaD/placebo bottles. In the case of non-compliance to the intervention, study participants will not be discontinued or excluded from the study. Non-compliant participants will be analysed through per protocol analysis. Non-compliant participants will be considered as those who have had less than 80% compliance throughout the treatment period or who have missed two consecutive doses.

Secondary endpoint analysis

The same analysis as for the primary endpoint will be repeated for VL at days 8 and 21. The duration of symptoms will be analysed using the same linear model as for the primary analysis and adjusting for VL. Time-to-event variables (duration of symptoms) will be analysed using a Cox model. The SF-12 survey score (mental and physical scores) will be analysed using a linear regression model comparing the intervention arm with the placebo arm.

Interim analyses are not planned. Available data will be analysed at the end of the study. No imputation will be performed.

Ethics and dissemination

This study will be conducted in compliance with the protocol, the current version of the Declaration of Helsinki, the ICH-Good Clinical Practice guidelines and all national legal and regulatory requirements. Substantial amendments to the study protocol are only implemented after the obtention of the competent authorities’ approvals. Under emergency circumstances, deviations from the protocol to protect the rights, safety and well-being of human subjects may proceed without prior approval of the authorities. Such deviations shall be documented and reported to the authorities within 72 hours.

Results from this study will be presented at national and international scientific conferences and disseminated via articles published in scientific journals. No patient data that could permit study participant identification will be presented. Publication of data derived from this study will be supervised by the sponsor. No publication will be made without the prior approval of the sponsor. The sponsor will have access to the final trial dataset.

Ethics statements

Patient consent for publication

Consent obtained directly from patient(s).