Study Design
This was a prospective observational study conducted at Seoul National University Hospital (SNUH), which is a tertiary care university-affiliated hospital in South Korea. The study period spanned July 2018 to June 2019.
Individuals aged > 50 years, who were diagnosed with a hematologic malignancy and had been cured with either autologous or allogeneic HSCT, were eligible. These patients were stratified according to time since transplantation: 2–5 years and > 5 years (hereafter referred to as HSCT 2–5 year and HSCT > 5 year, respectively). Controls included patients with a hematologic malignancy who had undergone cytotoxic chemotherapy and were cured, without relapse for at least 6 months (referred to hereafter as the chemotherapy group). Moreover, healthy volunteers with normal immune status were recruited (referred to hereafter as the healthy group). Exclusion criteria included GvHD, use of immunosuppressants or antiviral agents, HZ reactivation within 1 year of the study period, or receipt of HZ vaccines. All eligible cases were screened during the study period.
Participants who met eligible criteria were given a single dose (0.65 mL) of ZOSTAVAX®. Blood samples were collected to test both humoral and cellular immune responses against VZV prior to vaccination and at 6 weeks post-vaccination. Baseline characteristics included age, sex, underlying diseases, type of HSCT or cytotoxic chemotherapy, and previous history of HZ.
Glycoprotein ELISA (gpELISA)
VZV-specific antibodies were measured quantitatively using a SERION ELISA classic Varicella Zoster Virus IgG kit (Institut Virion/Serion GmbH, Würzburg, Germany). This gpELISA assay uses a lentil-lectin affinity-purified preparation of glycoprotein from VZV-infected MRC-5 cells as the antigen [16, 17]. Antigen-coated 96-well plates were incubated with test sera. Human immunoglobulin G antibodies (IgGs) bound to antigen were detected by incubation with anti-human IgG antibodies. Color was developed after reaction with a substrate. Optical density was read at 405 nm.
ELISPOT Assay
Peripheral blood mononuclear cells (PBMCs) collected and frozen during the study period were tested using a BD™ IFN-γ ELISPOT kit (BD Bioscience, San Jose, CA, USA). Briefly, PBMCs were activated with VZV and exposed to control antigens on anti-IFN-γ coated plates. After washing, a solution containing a biotinylated anti-IFN-γ detection antibody was added to each well, and streptavidin-HRP solution and substrate were used for color development. Spots were counted with a CTL-ImmunoSpot® reader (CTL ImmunoSpot, Cleveland, OH, USA) and reported as the net number of VZV-specific IFN-γ spot-forming cells (sfc) per 106 PBMCs (the difference between responses to VZV antigen and control antigen) [18]. Samples lacking sufficient PMBCs and results with phytohemagglutinin responses < 300 sfc were not included in the analysis [19].
Safety
All vaccinated individuals were evaluated for adverse events using a self-reported structured questionnaire administered 6 weeks post-vaccination. The type and severity of local and systemic adverse events were assessed. Adverse events were graded on a standard scale [20]. The causality of an adverse event after immunization was classified as follows: unlikely (inconsistent), possible (indeterminate), or likely (consistent) [21]. Subjects also informed the study nurse or physician if they noticed any serious adverse reactions immediately after vaccination.
Statistical Analysis
For continuous variables, mean (standard deviation, SD) and median (interquartile range, IQR) were used for normally and abnormally distributed data. Categorical variables were expressed as numbers and percentages. A t-test was used to compare continuous variables and the Chi-square or Fisher’s exact test was used to compare categorical variables. One-way ANOVA with Dunnett’s adjustment or Kruskal-Wallis test were used to calculate P-values for comparisons of more than two variables. All tests were two-sided and P-values < 0.05 were considered statistically significant. All statistical analyses were performed using SPSS for Windows (version 22; IBM Corp., Armonk, NY, USA).