Molecular Characterization of BK Polyomavirus Replication in Allogeneic Hematopoietic Cell Transplantation Patients

• Published in: The Journal of infectious diseases Vol. 227; no. 7; pp. 888 - 900
• Main Authors: Leuzinger, Karoline, Kaur, Amandeep, Wilhelm, Maud, Frank, Konstantin, Hillenbrand, Caroline A, Weissbach, Fabian H, Hirsch, Hans H
• Format: Journal Article
• Language: English
• Published: United States Oxford University Press 12.04.2023
• Subjects: Allografts; Antibodies; Antibodies, Neutralizing; BK Virus - genetics; CD8 antigen; CD8-Positive T-Lymphocytes; Cell culture; Cell therapy; Cell-mediated immunity; Cytotoxicity; Deoxyribonuclease; Genetic diversity; Genomes; Genotype & phenotype; Genotypes; Hematopoietic Stem Cell Transplantation; Hematopoietic stem cells; Hemorrhagic cystitis; Humans; Lymphocytes T; Major; Next-generation sequencing; Polyomavirus Infections; Replication; Stem cell transplantation; Urine; VP1 protein; BKPyV; hematopoietic cell transplantation; T cell; CD8; Vp1; HCT; neutralizing antibody; immune escape; hemorrhagic cystitis; epitope; LTag; BK polyomavirus
• ISSN: 0022-1899; 1537-6613
• DOI: 10.1093/infdis/jiac450

Abstract Background High-level BK polyomavirus (BKPyV) replication in allogeneic hematopoietic cell transplantation (HCT) predicts failing immune control and BKPyV-associated hemorrhagic cystitis. Methods To identify molecular markers of BKPyV replication and disease, we scrutinized BKPyV DNA-loads in longitudinal urine and plasma pairs from 20 HCT patients using quantitative nucleic acid testing (QNAT), DNase-I treatment prior to QNAT, next-generation sequencing (NGS), and tested cell-mediated immunity. Results We found that larger QNAT amplicons led to under-quantification and false-negatives results (P < .001). DNase-I reduced urine and plasma BKPyV-loads by >90% (P < .001), indicating non-encapsidated BKPyV genomes. DNase-resistant urine BKPyV-loads remained infectious in cell culture. BKPyV genome fragmentation of ≤250 bp impaired NGS coverage of genetic variation using 1000-bp and 5000-bp amplicons. Conversely, 250-bp amplicons captured viral minority variants. We identified genotype-specific and genotype-independent changes in capsid Vp1 or T-antigen predicted to escape from antibody neutralization or cytotoxic CD8 T-cells, respectively. Genotype-specific changes in immunodominant 9mers were associated with reduced or absent CD8 T-cell responses. Thus, failure to control BKPyV replication in HCT Patients may involve insufficient genotype-specific cytotoxic CD8 T-cell responses, potentially predictable by low neutralizing antibodies as well as genotype-independent immune escape. Conclusions Our results provide new insights for patient evaluation and for designing immune protection through neutralizing antibodies, adoptive T-cell therapy, or vaccines. BKPyV genome loads in allogeneic hematopoietic cell transplantation are DNase-I sensitive, nonencapsidated DNA fragments of ≤250 bp. This can impair detection of BKPyV diversity by next-generation sequencing and specifically genotype-associated changes mediating BKPyV immune escape from cytotoxic CD8 T-cell killing.