Use of convalescent plasma in the treatment of critically ill COVID-19 patients: a study protocol

1. Introduction

In December 2019, an outbreak of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection was discovered in Wuhan, China, and rapidly spread globally [1]. The World Health Organization (WHO) named the disease caused by this infection coronavirus disease 2019 (COVID-19). On 31 January 2020, the WHO classified the COVID-19 outbreak as a pandemic [2]. COVID-19 remains a significant threat to global health and the global economy. The use of convalescent plasma (CP) has shown promise in early studies with COVID-19 patients. CP is the liquid portion of blood from patients who have recovered from confirmed SARS-CoV-2 infection, which can be collected using a cell separator machine (apheresis). CP is thought to act through classical adoptive immunotherapy because the plasma collected from donors who have recovered from COVID-19 likely contains antibodies (proteins) that might provide passive immunity. CP can mediate immune mechanisms such as the neutralisation of viremia, the initiation of antibody-dependent cellular toxicity, and the modification of the inflammatory response. CP can also serve a non-immune system role through the restoration of coagulation factors [3]. Many patients diagnosed with COVID-19 develop antibodies one to two weeks after the presentation of symptoms [4, 5]. According to the United States Food and Drug Administration (FDA) recommendations, the SARS-CoV-2 neutralising antibody titre should be at least 1:160 [6]. Most individuals with previous COVID-19 developed measurable antibody responses and neutralizing antibodies; however, antibody titre measurement can be variable. Therefore, comparative studies between serological assays are needed to reach a consensus. Recent studies [7, 8, 9, 10, 11, 12] have indicated that the administration of CP to critically ill patients has resulted in improved clinical, laboratory, and radiological status of patients, accompanied by decreased mortality. There is a need for more information to guide the treatment of COVID-19 patients. Several drugs have not yet proven to be effective, and current treatment consists of supportive care. This study aims to help gain evidence on the effect of CP in the treatment of COVID-19 patients at their early stage of critical illness. Until medications are available, CP is a potential therapy for COVID-19 patients. However, the safety, efficacy, precise indications of use, optimal time, dose, and titre for CP therapy are not well characterized. Thus, a protocol regarding the use of CP could help to provide treating doctors with scientific evidence on a potential therapeutic option and better clinical management of critically ill COVID-19 patients. Furthermore, COVID-19 is becoming more aggressive with increased numbers of critically ill patients. The objective of this study was to prepare a guiding protocol for the use of CP in the treatment of critically ill COVID-19 patients to assess efficacy and safety.

2. Methods

2.6 Intervention

Donor CP from fully recovered COVID-19 patients will be collected using a cell separator machine (Trima Accel, Automatic Blood Collection System) (Fig. 1). A fully recovered donor is defined as any healthy adult who has recently recovered a confirmed COVID-19 infection, who is clinically asymptomatic for a minimum of 14 days and who has tested negative for SARS-CoV-2 twice using a real-time reverse transcriptase-polymerase chain reaction (RT-PCR) assay on a respiratory tract sample, with a minimum of 24 hours between the two sample collections. The collection of plasma will occur a minimum of four weeks after the initial onset of symptoms and at least two weeks after disease recovery. In addition, we will adhere to the WHO blood donor selection criteria for the assessment of donor suitability for blood donation [14, 15]. The COVID-19 unit will be responsible for the identification and targeted recruitment of convalesced patients who qualify to serve as CP donors. The donor antibody titre will be measured by an IgG ELISA detecting spike receptor-binding domain (S-RBD). A cut-off of CP with anti-SARS-CoV-2 antibody titre $\ge$1:160 will be used according to FDA recommendations. The expert blood bank staff are responsible for donor selection, pre-donation screening, and CP collection using standard operating procedures and according to international guidelines. CP will be labelled (as CP for investigational use only) using a Zebra Designer 2 system, and product safety and distribution to the hospital will follow the same protocols that were established by our study hospital [16, 17]. Donors who have not previously received a transfusion are preferable, to avoid the occurrence of transfusion-related acute lung injury (TRALI). Six hundred ml of CP will be collected from each donor in a single sitting. Viral inactivation through irradiation is mandatory if the CP will be used within six hours of collection, without freezing, to reduce any possibility of infection transmission [18]. The 600 mL CP will be divided into 200 mL doses. Donors can donate twice each month for up to 12 months, based on the American Association of Blood Bank (AABB) guidelines. The CP will be tested for transfusion-transmitted infections, according to the standard procedure for blood transfusion. Finally, CP will be stored as fresh frozen plasma (FFP) at $\le$–18 ${}^{\circ }$C, separate from plasma for routine use [19]. Selection of blood groups, according to ABO compatibility, is illustrated in (Table 1). Based on published studies examining the use of CP to treat previous respiratory viral epidemics, the proposed volume of plasma that will be used for each patient is 6 mL/kg of compatible CP, from a single donor, divided into two doses separated by 24 hours. The decision to administer CP will be approved by the treating team doctors, including a pulmonologist, infectious disease specialist, intensive care specialist, and haematologist. The CP request will require that all study criteria are fulfilled, and the CP request form will include the patient’s demographic and clinical information (Table 2). Other staff that will participate in the trial will include physicians, immunologists, nurses, laboratory scientists, statisticians, and infectious diseases experts.

Fig. 1.Collection and transfusion of two units of convalescent plasma.
The image was reproduced with the permission of David Spach, MD.

Table 1.ABO group selection order for the transfusion of fresh frozen plasma.

Patient ABO group	Fresh frozen plasma
O
	First choice	O
	Second choice	A or B
	Third choice	AB
A
	First choice	A
	Second choice	AB
	Third choice	B*
B
	First choice	B
	Second choice	AB
	Third choice	A*
AB
	First choice	AB
	Second choice	A*
	Third choice	B*
* Tested and negative for high-titre anti-A and anti-B. Adapted from: Joint United Kingdom (UK) Blood Transfusion and Tissue Transplantation. Services Professional Advisory Committee. https://www.transfusionguidelines.org/transfusion-handbook/2-basics-of-blood-groups-and-antibodies/2-4-the-abo-system

Table 2. Convalescent plasma request form.

Convalescent plasma request form
Patient information
	MRNO……………………..……………Name……………………………………………
	Age…………………………………………………………………………………………
	Gender…………..…………………………………………………………………..…..…
	Contact address…………..……………………………………………………………..…
	Blood group……………………………….………………………………………….……
	Date of admission…………………………………………………………………………
	Ward/ICU……………………………………………………………………….…………
	COVID-19 diagnosis:	Confirmed ( )	Suspected ( )
	IgA deficiency:	Yes ( )	No ( )
Clinical history
	Pregnancy ( )	Breastfeeding ( )	Transfusion reaction ( )	Alloantibody ( )
Comorbidities
	Hypertension ( )	Diabetes mellitus ( )
	Cancer/other immunosuppressive states ( )	COPD ( )	Other ( )
Factors associated with the severity
	Thrombocytopenia ( )	Lymphopenia ( )
	D-dimer $>$4 ng/mL ( )	CRP $>$30 mg/L ( )
CP team
	Provided consent	from the patient ( )	from his/her first-degree relatives ( )
	The decision of the CP team	Approved ( )	Not approved ( )
	Number of CP units	One ( )	Two ( )	Three ( )
	Date………………………………………………………………………………………
	Intensive Care Specialist	Pulmonologist	Haematologist	Infectious Disease Specialist
	Name:	Name:	Name:	Name:
	Signature:	Signature	Signature	Signature
Abbreviations: MRNO, medical record number; ICU, intensive care unit; COVID-19, coronavirus disease 2019; IgA, immunoglobulin A; COPD, chronic obstructive pulmonary disease; CRP, C-reactive protein; SOFA, sequential organ failure assessment; CP, convalescent plasma.

2.6.1 Relevant concomitant care that will be permitted or prohibited during the trial

$\bullet$ Regular prescription medications will be permitted.

$\bullet$ Over-the-counter medications will be prohibited.

$\bullet$ Herbal treatments/nutritional supplements will be prohibited.

$\bullet$ Blood products, as prescribed by the trial, will be permitted.

2.6.2 Provisions for post-trial care andmonitoring subject compliance

Recipients will be followed up at four hours, seven days, and 30 days after infusion. Follow-up will be mandatory, and AEs will be closely monitored and reported to the institutional review board (IRB) chairman [20]. A follow-up form will be generated and filled out by the treating doctor to evaluate the safety and efficacy of the trial (Table 3).

Table 3.Convalescent plasma assessment and follow-up form.

Convalescent plasma assessment and follow-up form
Patient information
	MRNO……………………………………..……………………………………………
	Name………………………………………………………………………………………
	Age…………………………………………………………………………………………
	Gender………………………………………………………………………………………..
	Contact address……………………………………………………………………………
	Blood group……………..…………………………………………………………………
	Date of admission……………..……………………………………………………………..
	Ward/ICU……………………………………………………………………………………
	COVID-19 diagnosis:	Confirmed ( )	Suspected ( )
	IgA deficiency:	Yes ( )	No ( )
Clinical Assessment and follow-up
	Parameters	Pre-infusion of CP	Four hours after the first dose	Seven days after the first dose	30 days after the first dose	On discharge day
	Breathlessness
	Temperature
	Respiratory rate
	Pulse rate
	Blood pressure
	O2 saturation
	PaO2/Fio2
	Partial O2 pressure
	SOFA score
	CBC
	CRP
	D-dimer
	Chest X-ray
	Outcome
CP team
	Date:
	Intensive Care Specialist	Pulmonologist	Haematologist	Infectious Disease Specialist
	Name:	Name:	Name:	Name:
	Signature:	Signature	Signature	Signature
Abbreviations: MRNO, medical record number; ICU, intensive care unit; COVID-19, coronavirus disease 2019; IgA, immunoglobulin A; CP, convalescent plasma; SOFA, sequential organ failure assessment; CBC, complete blood count; CRP, C-reactive protein.

2.6.3 Outcomes measurements

The result assessed at the end of this trial will be as follows.

2.6.3.1 Efficacy indicators

The efficacy indicators include the primary endpoint parameters.

Primary endpoints will be:

$\bullet$ Discharge from the ICU

$\bullet$ Discharge from the hospital

$\bullet$ Days of hospitalisation

$\bullet$ Number of days on mechanical ventilation after the administration of CP

$\bullet$ Death

$\bullet$ SARS-CoV-2 viral clearance and viral load

$\bullet$ Inflammatory markers analyses

$\bullet$ Cytokine analyses

$\bullet$ Difference in oxygen saturation

2.6.3.2 Safety evaluation

The safety evaluation will include the secondary endpoints.

Secondary endpoints will be:

$\bullet$ Severe transfusion reactions

$\bullet$ Transfusion-transmitted diseases

2.6.4 Assignment of interventions

2.6.4.1 Allocation

The allocation to the intervention or control groups will occur in a randomised and blinded manner. The randomisation will be performed using computer-generated random numbers.

Patients who sign the informed consent will be randomised by the team into one of the two study arms:

$\bullet$ Experimental arm: will receive the standard treatment as usual, according to the local protocol, and two doses of compatible CP.

$\bullet$ Control arm: will receive only the standard treatment as usual, without the addition of CP (Fig. 2).

Fig. 2.Diagram illustrating the steps for convalescent plasma donation and transfusion.

2.6.4.2 Blinding

Based on the objectives of the study, the identity of the intervention and control treatments will not be known to investigators, research staff, or patients to ensure the double-blinded administration of the study treatments. Access to the randomisation code will be strictly controlled to eliminate any forms of bias. The packaging of the CP concentrates and labelling will only be known to the principal investigator (PI). The clinical measurements obtained to describe the study results will be maintained confidential to avoid breaking the blind. The study blind will be broken upon completion of the clinical study after the study database has been locked. Only the PI will be aware of each subject’s treatment assignments.

Double Blinding has taken care of the unanimity. High ethical standards will be maintained.

2.6.4.3 Procedure for unblinding if needed

During the study, the blind may be broken only in emergencies, when knowledge of the patient’s treatment group becomes necessary for further patient management. In that case, the PI will discuss the emergency with the medical monitor before unblinding.

The study will be discontinued, and a review will be performed if any of the following events occur or if subject safety is at risk of becoming compromised:

(1) The unexpected death of any subject, in association with CP infusion.

(2) The occurrence of a life-threatening allergic/hypersensitivity reaction (anaphylaxis).

(3) Observed bronchospasm, urticaria, or angioedema requiring urgent support.

(4) At least one subject with an unexpected SAE associated with CP.

(5) An overall pattern of symptomatic, clinical, or laboratory events that the medical monitor considers to be associated with CP.

(6) Any other event that is considered to be an SAE in the good clinical judgment of the physician. All AEs will be appropriately documented.

Due to the risk of AEs associated with CP treatment, out of an abundance of caution, we will monitor the number of subjects in each trial arm that progress to an indication for the need for mechanical ventilation.

3. Results

The results assessed at the end of this trial will be the primary and secondary endpoints. The primary endpoints (efficacy indicators) will be the discharge from the ICU, discharge from the hospital, days of hospitalisation, days on mechanical ventilation and death. The main secondary endpoints (safety indicators) will be the severe transfusion reactions and transfusion-transmitted diseases.

4. Discussion

The presented study protocol was developed by the authors using standardised criteria for the development of a clinical trial protocol. This manuscript describes a study protocol to assess the effectiveness and safety of a CP treatment program that can be used during both the current and future pandemics. Because of the current worldwide COVID-19 outbreak a wide range of solutions must be explored to treat the increasing numbers of critically ill patients. The Food and Drug Administration (FDA) recently announced that CP is among the therapeutic approaches that are being actively considered until the successful development of specific treatment drugs. Many authors have explored this concept recently. The current study considers critically ill patients to represent the appropriate target treatment group for the designed CP protocol. Strict criteria for the enrolment of critically ill patients were considered and are in agreement with the available literature [21, 22]. The use of CP is a well-established technique that has been applied with varying success to treat several viral respiratory epidemics. Based on recently published trial studies, the administration of CP is expected to be safe and efficient, and critically ill patients who received CP have been associated with better outcomes than matched control patients. Preparation of a detailed protocol is essential before the implementation of this technique. The protocol that has been developed by the current study is feasible, with satisfactory outcomes, due to collaboration and teamwork, and the availability of potential donors. Our protocol aligns with other published protocols that have been released to address the current pandemic [20, 22, 23]. The present protocol is consistent with the protocol developed by Eckhardt and his team. The only differences are: Eckhardt’s was a single-centre randomised, controlled trial and the study participants will receive one unit (200–250 mL) of CP versus one unit (200–250 mL) of the control plasma [22]. Our protocol is aligned with the protocol developed by Albalawi et al. [23] but differs in the sense that in Albalawi et al. [23] protocol, the volume of the CP will be 300 ml (200-400 ml/ treatment dose) at least once, and if needed daily for up to 5 sessions. Furthermore the anti-SARS-CoV-2 antibody titre was not determined [23]. The current protocol is also consistent with the recently published protocol apart from the source of CP [24]. In the recently published protocol, the CP will be from different donors, while in the present study the CP will be from a single donor [24]. The slight variation between the published study protocols and the present study allows a comparison between different protocols to reach a consensus about several variables such as the precise indications of use, the optimal time of CP infusion, dose, the concentration of anti-SARS-CoV-2 antibody titre.

This study protocol had some limitations. The generalizability of the results may be limited, as the patients participating in the study will include only critically ill patients.

5. Conclusions

The developed protocol will be used in a clinical trial to evaluate the efficacy and safety of CP used in the treatment of critically ill COVID-19 patients. The details that will be obtained from this study protocol can support its application in similar environments. This study protocol addressed all aspects that are necessary for the standardised implementation of CP for the treatment of COVID-19 patients, including CP collection, administration, evaluation, and follow up.

Author contributions

Conceptualization—AIOY, AYA, AMJ, EIM, HKK. Methodology—AIOY, AJ. Validation—AIOY, AJ. Investigation—AIOY. Data curation—AIOY, AJ. Writing original draft—AIOY. Writing-review and editing—AIOY, AYA, AJ, EM, HK. Project administration—AIOY. Supervision—AIOY. All authors approved the final version of the manuscript for publication.

Ethics approval and consent to participate

This study protocol was reviewed and approved by the institutional review board of the King Abdullah Hospital, Saudi Arabia (IRB NO. 002/07/21). Written informed consent will be obtained from the subject or the subject’s legal representative, and the ability for the subject to comply with the requirements of the study will be evaluated (see supplementary files). Participants recruited to a study will be informed of the objectives, methodology and interventions that they may receive if they agree to participate. Only those who give their informed consent in writing will be included in the study. Participants will be allowed to withdraw at any time in the study. However, follow-up will continue for withdrawn recipients to ensure their safety and to document any SAEs.

Acknowledgment

The authors extend their appreciation to the Deanship of Scientific Research at University of Bisha Saudi Arabia for funding this work through COVID-19 Initiative Project under Grant Number (UB – COVID – 01 – 1441).

Funding

This work was supported by the Deanship of Scientific Research at University of Bisha Saudi Arabia [grant number UB – COVID – 01 – 1441].

Conflict of interest

The authors declare no conflict of interest.

Supplementary material

Supplementary material associated with this article can be found, in the online version, at https://oss.signavitae.com/mre-signavitae/article/1443153896320974848/attachment/Supplementary_Files.docx.

Data availability

The data used in this study are available from the corresponding author upon request.