Intractable Coronavirus Disease 2019 (COVID-19) and Prolonged Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) Replication in a Chimeric Antigen Receptor-Modified T-Cell Therapy Recipient: A Case Study

Abstract

A chimeric antigen receptor-modified T-cell therapy recipient developed severe coronavirus disease 2019, intractable RNAemia, and viral replication lasting >2 months. Premortem endotracheal aspirate contained >2 × 1010 severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) RNA copies/mL and infectious virus. Deep sequencing revealed multiple sequence variants consistent with intrahost virus evolution. SARS-CoV-2 humoral and cell-mediated immunity were minimal. Prolonged transmission from immunosuppressed patients is possible.

Keywords: COVID-19; SARS-CoV-2 RNAemia; SARS-CoV-2 immune responses; SARS-CoV-2 infectivity; SARS-CoV-2 intrahost variation; severe acute respiratory syndrome coronavirus 2.

Figure 1.

I, Clinical timeline showing chimeric antigen receptor-modified T-cell infusion (days –25 to 0), first hospital admission (days 0–17), home stay (days 18–41), and second hospital admission (days 41–74). Day 0 denotes day of first positive nasopharyngeal (NP) swab polymerase chain reaction (PCR) for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) RNA. The top panel shows the clinical course and timeline, with results of clinical SARS-CoV-2 NP swab reverse-transcription PCR (RT-PCR) testing and clinical immunoglobulin G and immunoglobulin A testing. The bottom graph shows serial plasma SARS-CoV-2 RNA quantification obtained using a research quantitative RT-PCR assay, with viral quantification from endotracheal aspirate fluid using research assays (quantitative RT-PCR and infectious viral titer). Inset shows a coronal view of chest computed tomography from day 41. Green squares: convalescent plasma; red squares: remdesivir; yellow squares: dexamethasone. II, Indirect immunofluorescence of SARS-CoV-2 (green) isolated in Vero E6 cells (red/blue) from day 72 endotracheal aspirate samples. Green: antibody directed against SARS CoV-2 spike protein (Sinobiologicals). Blue: DNA counterstained with DAPI. Red: filamentous actin counterstained with phalloidin. A syncytium is shown in the inset (right) at higher magnification. Scale bars: 100 µm. III, Electron microscopy of endotracheal aspirate sample obtained on day 72. A, Comparison of virus localization in a bronchial epithelial cell (top) and a transient secretory cell (bottom). Each image is shown as montaged 2D overviews of the whole cell in a 200-nm section (left), overlaid with colored dots to indicate positions of virions (center) and 3D tomographic reconstructions detailing virus populations within cytoplasmic compartments. B, 2D overview of SARS-CoV-2–infected ciliated epithelial cell in a 200-nm semi-thick section. The apical side of the cell is characterized by numerous membrane-bound compartments that are filled with SARS-CoV-2 virions. C, Montaged tomogram of the apical side of the cell shown in (B); hundreds of virus particles are contained within smooth-walled cytoplasmic compartments. Inset: Detail of a single virion with spikes indicated by red dots. D, Tomogram detail of a smooth-walled cytoplasmic compartment containing at least 30 SARS-CoV-2 virions within the shown 15-nm thick volume. Abbreviations: CAR, chimeric antigen receptor; COVID-19, coronavirus disease 2019; CP, convalescent plasma; CT, computed tomography; IgA, immunoglobulin A; IgG, immunoglobulin G; NP, nasopharyngeal; PCR, polymerase chain reaction; PFU, plaque-forming units; SARS-CoV-2, severe acute respiratory syndrome coronavirus 2.

Figure 2.

Mutations and deletions in the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) Spike gene identified in the patient’s samples compared to the SARS-CoV-2 GISAID GH Clade circulating in Pittsburgh. The full genome at the top shows the GH clade sequence. The enlarged S gene shows all of the mutations identified in the patient’s samples compared with the GH clade. The sequence alignments in S compared to the GH clade are shown for each of the multiple (6) sequence variants (var) identified by deep next-generation sequencing (Illumina). All of the sequence variants were detected in plasma. Day 72 (*) shows the matching sequence variants identified in the endotracheal aspirate sample. The D614G substitution was found in all samples. #Mutations detected as mixed populations <100% but >20%. Abbreviations: E, envelope; FP, fusion peptide; HR 1, heptad repeat 1; M, membrane; N, nucleocapsid; NTD, N-terminal domain; ORF, open reading frame; RBD, receptor binding domain; RBM, receptor binding motif; SP, signal peptide; TM, transmembrane domain; Var, sequence variant.