HLA tetramers and anti-CMV immune responses: from epitope to immunotherapy.

BACKGROUND: Identification of HLA class I-restricted CMV epitopes, and the subsequent synthesis of HLA class I-peptide tetrameric complexes, have provided investigators with an important tool for visualising and quantifying the precise in vivo CTL response to CMV reactivation following stem cell transplantation. In conjunction with PCR-monitoring of the viral load, the magnitude and dynamic of the host's specific CD8(+) T cell response to viral replication can be studied.
METHODS: CMV peptide epitopes can be identified be searching the CMV-pp65 antigen for HLA class I allele binding motifs, by testing their binding affinity and ability to generate CTLs, and by screening for CTL responses in as many individuals as possible to assess their general applicability for monitoring large number of patients. HLA tetramers are synthesized by refolding recombinant class I heavy chains and beta(2)m with CMV-pp65(495-503) peptide. After biotinylation and tetramerisation to PE-conjugated streptavidin, they are used to stain CD8(+) T cells taken from patients at different time points after SCT.
RESULTS: The T-cell mediated immune response is mainly directed against epitopes derived from the CMV tegument protein pp65. CMV-specific CTL's confer protection against CMV reactivation above a threshold level of 10(7) to 2 x 10(7)/L. CMV reactivation is required to stimulate CTL responses. Transfer of CMV immunity from seropositive donors is associated with better outcome and steroids suppress the Ag-specific immune response. DISCUSSION: Initial studies with CMV-specific HLA class I tetramers have helped to define the nature of anti-CMV T cell response in SCT patients and to determine a threshold CTL level required for controlling CMV reactivation. Monitoring patients with HLA-tetramers should therefore allow clinicians to predict and assess the risk of reactivation and to balance the risks and benefits of early anti-viral treatment, thereby avoiding the hazards of anti-viral prophylaxis. HLA-tetramers can also be used to isolate antigen-specific cells for further in vitro expansion and transfer to patients for antiviral immunotherapy. The threshold level determined from patient monitoring can be used as a guide for estimating an effective target cell dose.