CP donors
CP was collected by plasmapheresis from 9 patients (7 males and 2 female). Median age was 55 years (IQR, 45 to 63 years). Median time from diagnosis to plasmapheresis was 50 days (IQR, 38 to 56 days). Median levels of IgA and IgG antibodies were 7.52 (range: 1.68–12.35) and 10.96 (range: 2.48–12.68) respectively.
Patients‘ characteristics
From May 7, 2020 to July 24, 2020, 9 patients (4 males and 5 females) with WHO grade 4 COVID-19 disease were enrolled and received CP transfusion. Patient characteristics at diagnosis are shown in Table 1. Median age was 61 years (interquartile range [IQR], 60–62 years). Median time from symptom onset to hospital admission and CP transfusion was 1 day (IQR, 0–3 days) and 6 days (IQR, 4–7 days), respectively. Most common symptoms at disease onset were fever (8 cases), cough (7 cases), shortness of breath (3 cases) and headache (3 cases), while less common symptoms included diarrhea (2 cases), loss of taste (2 cases), loss of smell (2 cases), arthralgias (1 case), myalgias (1 case), and sore throat (1 case). Four patients were overweight, 4 had normal weight and 1 was obese. Four patients had comorbidities: 1 pulmonary fibrosis, 1 essential hypertension and diabetes mellitus, 1 chronic obstractive disease (COD) and lung cancer and 1 essential hypertension only. Antibacterial treatment was used in 5 patients with coexistent bacterial infection. One patient received treatment with tocilizumab (single dose of 8 mg/kg) after clinical deterioration and intubation, 6 days after the first CP infusion.
Table 1
Clinical characteristics of patients
Patient no | Sex | Age, y | BMI category | Time to admission from symptom onset (days) | Time to CP treatment from symptom onset (days) | Principal symptoms | Oxygen support | SOFA score | Comorbidities |
1 | M | 62 | Overweight | 0 | 4 | Fever, cough, shortness of breath, headache | Low-flow nasal cannula | 4 | None |
2 | F | 60 | Overweight | 1 | 3 | Fever, cough, malaise | Low-flow nasal cannula | 4 | None |
3 | M | 60 | Overweight | 0 | 6 | Fever, cough, lack of taste, lack of smell, diarrhea | High-flow nasal cannula | 7 | Pulmonary fibrosis |
4 | F | 62 | Overweight | 5 | 6 | Fever, cough | Low-flow nasal cannula | 7 | None |
5 | M | 52 | Normal weight | 11 | 18 | Cough, headache | High-flow MV | 7 | Hypertension, diabetes mellitus |
6 | F | 29 | Normal weight | 0 | 11 | Fever, myalgias, arthralgias, headache, lack of taste, lack of smell | None | 2 | None |
7 | F | 73 | Obese | 3 | 5 | Fever, cough, sore throat | Low-flow nasal cannula | 3 | Hypertension |
8 | F | 61 | Normal weight | 3 | 7 | Fever, cough, shortness of breath | Low-flow nasal cannula | 5 | None |
9 | M | 70 | Normal weight | 1 | 1 | Fever, shortness of breath | High-flow MV | 5 | COD, lung cancer |
On computer-assisted tomography (CT), all patients showed bilateral ground-glass opacities and/or pulmonary parenchymal consolidation with predominantly subpleural and bronchovascular bundle distribution. Seven patients had multiple lobe involvement, and five patients had interlobular septal thickening. At the time of enrolment no patient had life threatening disease.
Effects of CP transfusion
Improvement of clinical symptoms. All symptoms, including fever and cough, improved significantly after a median of 8 days (IQR, 6.5–15.5 days). In 6 out of 9 patients (66.6%), symptomatic improvement was observed already after the 1st CP dose. Prior to CP transfusion, 8/9 patients needed oxygen treatment: 3 patients were on Venturi Mask and 5 received low-flow nasal cannula oxygenation as shown in Table 2. Following CP treatment, 4 patients experienced deterioration of their respiratory function associated with progression of infection and received mechanical ventilation after 2, 3, 5 and 7 days after the 1st dose of CP transfusion, respectively. All patients were extubated after 2, 5, 7 and 43 days, respectively. Median length of stay in ICU was 9.5 days.
Table 2
Comparison of SOFA score and oxygen support before and after CP transfusion
| Day 1: before CP transfusion | After CP transfusion day 7 | After CP transfusion day 14 | After CP transfusion day 21 | After CP transfusion day 28 | After CP transfusion day 35 |
Patient no | SOFA score | Oxygen support | SOFA score | Oxygen support | SOFA score | Oxygen support | SOFA score | Oxygen support | SOFA score | Oxygen support | SOFA score | Oxygen support |
1 | 4 | nasal cannula | 2 | nasal cannula | 0 | None | 0 | None | 0 | None | 0 | None |
2 | 4 | nasal cannula | 0 | None | 0 | None | 0 | None | 0 | None | 0 | None |
3 | 7 | MV 50% | 8 | Mechanical ventilation | 7 | Mechanical ventilation | 5 | Mechanical ventilation | 6 | Mechanical ventilation | 2 | Mechanical ventilation |
4 | 7 | nasal cannula | 5 | Mechanical ventilation | 2 | MV 40% | 2 | nasal cannula | 2 | nasal cannula | 2 | nasal cannula |
5 | 7 | MV 60% | 5 | nasal cannula | 0 | None | 0 | None | 0 | None | 0 | None |
6 | 2 | None | 2 | None | 0 | None | 0 | None | 0 | None | 0 | None |
7 | 3 | nasal cannula | 4 | Mechanical ventilation | 2 | nasal cannula | 0 | None | 0 | None | 0 | None |
8 | 5 | MV 35% | 2 | None | 2 | nasal cannula | 2 | nasal cannula | 0 | None | 0 | None |
9 | 5 | nasal cannula | 6 | Mechanical ventilation | 4 | MV 35% | 3 | nasal cannula | 0 | None | 0 | None |
MV = Venturi Mask |
Of the 4 remaining patients needing oxygen supply, all became free of need for oxygen administration within a median time of 9.5 days following CP infusion (5–28 days). Oxygen saturation improved consistently from pre-treatment levels through day 14 (median pre-treatment: 96%, day 7: 97%, day 14: 98%). (Fig. 1)
SOFA score after CP therapy decreased gradually to zero in 6/9 patients within a median time of 17.5 days (14–21 days), while in the remaining 3 patients it improved consistently within a median of 35 days. SOFA score significantly decreased on day 7 (2 vs 5, p = 0.047), on day 14 (2 vs 5 p = 0.011), on day 21 (0 vs 5, p = 0.017), and on day 28 compared to day 1 (0 vs 5, p = 0.027) as shown (Table 2, Fig. 1).
Changes in laboratory parameters after CP transfusion. Median lymphocyte count increased significantly after CP transfusion compared to baseline levels, starting on day 14 [median 2.10 × 109/lt vs 1.39 × 109/lt (p = 0.036), and remained significantly higher throughout days 21 and 28 [1.95 × 109/lt, p:0.035; and 2.165 × 109/l, p = 0.028, respectively). As shown in Fig. 1 and Table 3, several inflammatory markers including CRP, fibrinogen, LDH, and IL-6 decreased significantly on days 7, 14, 21, and 28 compared to baseline.
Table 3
Comparison of laboratory parameters before (day 1) and after (day 7 and day 14) CP transfusion
Parameters | Day 1: before CP transfusion | Day 7 after CP transfusion | Day 14 after CP transfusion | Day 21 after CP transfusion | P value |
Lymphocytes, (109/L, NR: 1.1- 4.0) | 1.39 (0.45–1.69) | 1.53 (0.33–2.1) | 2.10 (0.4–3.6)# | 1.95 (0.98–2.14)** | 0.036# 0.035** |
Platelets, (109/L, NR: 130–400) | 204 (133–522) | 395 (343–572)* | 433.5 (314–966)# | 273 (217–298)** | 0.008* 0.025# 0.025** |
CRP, (mg/L, NR: 0.00–6.00) | 23.1 (0.3–146) | 4.135 (0.1–284) | 2.025 (0.1–161) | NA | - |
Fibrinogen, (mg/dL, NR: 200–400) | 538 (334–1026) | 495 (367–811) | 330 (320–420)# | 367 (282- 472.2)** | 0.018# 0.036** |
LDH, (U/L, NR: 135–225) | 294 (162–354) | 212 (173–467) | 235 (138–303) | 146 (140–235)** | 0.012** |
Ferritin, (ng/mL, NR: 13–150) | 411 (264–1406) | 600 (215–1848) | 513 (86- 2413) | 479 (313–847) | - |
Intereukin-6, (pg/mL, NR: < 7) | 34.6(18.8–70.6) | 10.2 (1.5- 259.5) | 5.0 (1.5–14) | NA | - |
SaO2, (%, NR: ≥ 95) | 96 (93–98) | 97 ((94–98) | 98 (97–98)# | NA | 0.011# |
NA: not applicable |
*: p value for the comparison between Day 1 and Day 7. |
# : p value for the comparison between Day 1 and Day 14. |
**: p value for the comparison between Day 1 and Day 21. |
Increase of IgA and IgG antibodies and decrease of SARS-CoV-2 RNA
IgA and IgG antibody levels were determined at baseline and days 7, 14, 21, and 28 in 8/9 patients. All evaluable patients exhibited significant increases in SARS-CoV-2 IgA and IgG antibodies at days 7, 14, and 21 compared to pre-treatment levels (Fig. 2). Median IgA levels were: pre-treatment, 1.44 (0.48–7.75); day 7, 12.9 (5.34-45), p = 0.012); day 14, 28.75 (5.45–72.2), p = 0.008; day 21, 33.64 (5.56–69.2), p = 0.018, (all p values compared to baseline). Median IgG levels were: pre-treatment, 0.39 (0.2-8.0); day 7, 6.02 (0.81–39.32), p = 0.008; day 14, 9.585 (4.78–38.84), p = 0.012; day 21, 9.52(6.46–13.59), p = 0.018. A nonsignificant trend to increased IgA levels was observed on day 28 compared to day 1 [45.74 (4.11–56.4), p = 0.08], whereas IgG levels increased significantly [10.42 (7.79–13.4), p = 0.045]. Compared to day 7, IgG and IgA levels were significantly increased at day 14 (p = 0.038 and p = 0.015, respectively), while IgG was significantly increased also at day 21 (p = 0.018). Median time of detection of antibodies following CP infusion was 11 days from symptom onset (8–19 days). No significant correlation was found between levels of antibodies in the infused CP and recipient antibody levels on days 7, 14 and 21.
Concurrently, 9/9 patients showed reduction of SARS-CoV-2 RNA, as shown by increased Ct values at day 14 and day 21 compared to day 1 [day 14: 33.71 (26.99-45) p = 0.028, day 21: 45 (36.1–45) vs day 1 : 26.66 (19.22–37.53]. However, there was no significant correlation between the increase of SARS-CoV-2 antibodies and the reduction of viral RNA.
As shown in Table 4, 2/9 (22.2%) patients had undetectable SARS-CoV-2 RNA on day 7, while 4/9 (44.4%) had undetectable RNA on day 14 following CP infusion. Median time to PCR SARS-CoV-2 negativity was 14 days after CP infusion.
Table 4
Comparison of serum IgA and IgG antibodies and SARS-Cov-2 RNA status before and after (day 7 and day 14) CP transfusion.
| Day 1: before CP transfusion | Day 7 after CP transfusion | Day 14 after CP transfusion |
Patient no | Serum IgA antibodies | Serum IgG antibodies | Serum SARS-CoV-2 RNA load (Ct value) | Serum IgA antibodies | Serum IgG antibodies | Serum SARS-CoV-2 RNA load (Ct value) | Serum IgA antibodies | Serum IgG antibodies | Serum SARS-CoV-2 RNA load (Ct value) |
1 | 0.49 | 0.2 | 19.22 | 6.74 | 6.02 | 29.26 | > 14 | 12.29 | 26.99 |
2 | 7.75 | 8.0 | 29.23 | > 14 | 12.29 | 22.98 | > 14 | 11.60 | Negative |
3 | 1.66 | 0.42 | 24.32 | > 14 | 3.62 | 28.99 | > 14 | 7.09 | 27.14 |
4 | 0.48 | 0.28 | 24.26 | NA | NA | 29.13 | NA | NA | 31.2 |
5 | NA | NA | NA | > 12 | 39.32 | Negative | > 12 | 38.84 | Negative |
6 | 1.47 | 0.91 | 37.53 | 5.72 | 4.27 | Negative | 5.45 | 5.97 | Negative |
7 | 1.24 | 0.2 | 23.24 | 5.02 | 0.81 | 33.1 | > 12 | 4.78 | 35.42 |
8 | 2.91 | 0.4 | 34.0 | 43.5 | 11.63 | 31.0 | 57.8 | 12.55 | 32.0 |
9 | 0.6 | 0.26 | 29.0 | 5.34 | 4.19 | NA | 5.59 | 6.6 | Negative |
NA: not available |
Outcomes
After a median follow-up of 66 days (IQR 59–70 days), 9/9 patients remain alive. Eight patients recovered completely and were discharged from hospital after a median hospital length of stay of 21 days (IQR, 15.75 days to 27.5 days). One patient with a history of pulmonary fibrosis was extubated and remains hospitalized in regular ward on low oxygen for 72 days, as of today.
No correlation was found between comorbidities and length of hospital or ICU stay or between the pre-treatment levels of antiSARS-CoV2 antibodies and disease severity.
Adverse events
No adverse events related to CP transfusion have been recorded in any patient throughout the follow-up period with the exception of a mild erythema in one patient following the infusion of the first CP unit resolved upon administration of antihistaminic drugs.