In December 2019, the first patients were diagnosed with coronavirus disease 2019
(COVID-19), a disease caused by severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2).[1] In only a few months, COVID-19 has spread globally, affecting over 3 million individuals
with over 250,000 deaths (data updated on May 2).[2] This has prompted the World Health Organization (WHO) to declare the COVID-19 outbreak
a pandemic.[3] Unfortunately, no uniformly standardized therapy has been endorsed for COVID-19
and a number drugs are currently being investigated in several nonrandomized or randomized
trials for patients with life-threatening COVID-19.[3]
[4] These agents includes steroids, chloroquine, antiviral medications (i.e., remdesivir,
loparinir/ritonavir), and anti-inflammatory agents (i.e., tocilizumab, sarilumab,
baricitinib), the latter being utilized following the discovery of the involvement
of SARS-CoV-2-induced inflammatory response in the pathogenesis of SARS.[5]
[6]
Experiences from previous coronavirus epidemics (SARS and Middle East Respiratory
Syndrome) indicate that convalescent plasma collected from recovered COVID-19 patients,
containing neutralizing antibodies against SARS-CoV-2, could be a potentially effective
therapeutic weapon.[7]
[8]
[9]
[10]
[11]
[12]
[13] In a pilot study of 10 patients with severe COVID-19,[14] classified according to WHO Interim Guidance,[15] the investigators collected convalescent plasma with neutralizing antibody titers
≥ 1:640 dilution. All 10 patients had a rapid improvement in symptoms and pulmonary
lesions (within 1–3 days).[14] A case series by Shen and colleagues on five critically ill patients with COVID-19
further confirmed the potential benefit of this biological antiviral therapy.[16] With this background,[17] considering the dramatic situation and the high lethality rate of COVID-19 in Italy,
we have planned a multicenter interventional single-arm trial (Clinicaltrials.gov:NCT04321421)
to produce hyperimmune plasma for treating critical patients with COVID-19.[18] Briefly, 500 to 600 mL plasmaphereses are collected from adult male donors, recovered
from COVID-19 (defined as two consecutive nasopharyngeal swabs negative for reverse
transcriptase-polymerase chain reaction [RT-PCR] testing), aged 18 years or older,
through a cell separator device and evaluated for transmissible diseases (human immunodeficiency
virus, hepatitis C virus, hepatitis B virus, and syphilis) according to Italian laws.
Additional molecular tests for detection of hepatitis A virus, hepatitis E virus,
and parvovirus B19, and neutralizing titer for SARS-CoV-2 will be performed according
to the recommendations from Italian National Blood Center (CNS) for donation of anti–COVID-19
convalescent plasma.[19] Pathogen reduction of plasma is also performed before freezing or transfusion. Every
plasmapheresis is divided into two 250 to 300 mL units, each of which is used to treat
one patient with severe COVID-19 infection. A second infusion is also contemplated
after 48 hours, in the case of lack of response to the first unit. The primary endpoint
of the study is reduction of mortality rate. Other clinical endpoints, viral load
(quantitative RT-PCR expressed as number of copies/mL), and immune response (expressed
as neutralizing titer, which is the maximum dilution with a reduction of 90% of the
cytopathic effect; a positive titer is equal to or greater than 1:10) are also considered.
The sample size calculation involves the enrolment of 43 patients.[18] Patients are continuously monitored for adverse events; these will be communicated
to the local Transfusion Service staff, who in turn will notify them to the national
electronic system for hemovigilance (SISTRA; https://www.centronazionalesangue.it). In addition, according to directives from CNS,[20] donors are advised to inform their Transfusion Service in the case of reappearance
of symptoms compatible with SARS-CoV-2 infection within 14 days after donation (postdonation
information).
The study started on March 17, 2020. During the conduct of the study, several challenges
have emerged linked to the enrolment of convalescent donors. The first issue was with
regards to the selection of convalescent plasma donors among patients recovered from
COVID-19. Patient-donors aged over 70 years are usually excluded from plasma donation,
according to Italian laws for blood donor selection, due to a hypothetically increased
risk of apheresis-related adverse events. Similarly, pluriparous women are generally
excluded from donation, because of increased risk of transfusion-related acute lung
injury in patients receiving their plasma that contains high levels of antibodies
against recipients' class I and II human leukocyte antigens. In addition, candidates
for hyperimmune plasma donation must be in good health without current or pre-existing
diseases, according to Italian laws on selection of voluntary unpaid blood donors.[18] Information about the course of COVID-19 disease for the patient (e.g., severity,
treatment, dates of onset and resolution of clinical symptoms) is usually requested
prior to plasma collection. In our experience, approximately 30 to 40% of recovered
patients initially screened are finally eligible for convalescent plasma donation,
with comorbidities (i.e., cardiovascular, neoplastic, or autoimmune disorders), age
(> 70 years) and sex (pluriparous females) the main causes for exclusion from plasma
donation. In addition, CNS recommends that donors should be completely recovered by
at least 14 days without a positive anamnesis for contacts with subjects with documented
SARS-CoV-2 infection during this period. This could generate a problem for recovered
health care personnel who thus must donate plasma before their return to operational
medical COVID units.[19]
[20] A consistent proportion of convalescent plasma donors is represented by recovered
periodic voluntary blood donors belonging to blood donor associations and this is
very important as it increases the safety of collected blood. Potential convalescent
plasma donors are retrieved from a local registry of patients who were infected with
COVID-19 and are usually contacted by telephone by a doctor or health care personnel
who performs an initial anamnestic screening to discover possible contraindications
to plasma donations. In our experience, due to the high number of phone calls required
to provide adequate daily plasma apheresis donations (8–10/day), a health care professional
must be dedicated exclusively to this activity.
Another critical point is the titer of anti–SARS-CoV-2 neutralizing antibodies. In
our protocol, a minimum titer of 1:160 is required to collect plasma. Several serologic
tests based on enzyme-linked immunosorbent assay (ELISA) technology for detection
of specific immunoglobulin A (IgA), IgG, and IgM anti–SARS-CoV-2 antibodies are available
and several studies have tried to correlate their titers with patients' disease severity,
clinical outcomes, and with persistent immune protection of recovered patients against
SARS-CoV-2.[21]
[22]
[23]
[24]
[25] However, although these assays might be useful for preliminary screening of seropositive
donors, the neutralization test (see Duan et al[14] for a detailed description of the method) remains the gold standard for identification
and quantification of anti–SARS-CoV-2 neutralizing antibodies. This test, based on
measurement of reduction of viral cytopathic effect in cell culture following addition
of recovered patient's serum, does not necessarily correlate with anti–COVID-19 antibody
levels measured by ELISA assays, but accurately quantifies the anti–COVID-19 neutralizing
properties of donors' plasma, thereby giving a reliable indication of the in vivo
antiviral potency of the convalescent plasma. Although very specific, this test is
very technically demanding and requires 72 hours of incubation before results are
available. The neutralization test is therefore very useful for clinical and experimental
studies, but not for epidemiological investigations of seroprevalence. Our very preliminary
experience suggests that anti–COVID-19 neutralizing antibodies reach their peak early,
approximately 10 to 20 days from the disappearance of symptoms. Since we do not have
clear data on the persistence of neutralizing levels in plasma over time, we recommend
collecting convalescent plasma just after the 14-day period required by CNS for convalescent
plasma donation, providing that two consecutive nasopharyngeal swabs are negative
for COVID-19. Neutralization tests are usually performed on donors before convalescent
plasma collection, to select those donors with titers ≥1:160 and to eliminate as few
donations as possible.
In our opinion, beside the challenges previously mentioned, which will be mitigated
by implementing our expertise in managing convalescent donors, the collection of plasma
represents an attractive therapeutic option, also considering the possibility of collecting
plasma by plasmapheresis procedures quickly and effectively. Furthermore, the possibility
of having local donors offers the added value of providing specific immunity acquired
against the infectious agent of the local strain, considering that in other areas
the strain may be different. The final results of our trial will help to clarify the
therapeutic role of anti–COVID-19 convalescent plasma.
