Humoral anti-KLH responses in cancer patients treated with dendritic cell-based immunotherapy are dictated by different vaccination parameters

Abstract

PurposeKeyhole limpet hemocyanin (KLH) attracts biomedical interest because of its remarkable immunostimulatory properties. Currently, KLH is used as vaccine adjuvant, carrier protein for haptens and as local treatment for bladder cancer. Since a quantitative human anti-KLH assay is lacking, it has not been possible to monitor the dynamics of KLH-specific antibody (Ab) responses after in vivo KLH exposure. We designed a quantitative assay to measure KLH-specific Abs in humans and retrospectively studied the relation between vaccination parameters and the vaccine-induced anti-KLH Ab responses.Experimental designAnti-KLH Abs were purified from pooled serum of melanoma patients who have responded to KLH as a vaccine adjuvant. Standard isotype-specific calibration curves were generated to measure KLH-specific Ab responses in individual serum samples using ELISA.ResultsKLH-specific IgM, IgA, IgG and all IgG-subclasses were accurately measured at concentrations as low as 20 μg/ml. The intra- and inter-assay coefficients of variation of this ELISA were below 6.7 and 9.9 %, respectively. Analyses of 128 patients demonstrated that mature DC induced higher levels of KLH-specific IgG compared to immature DC, prior infusion with anti-CD25 abolished IgG and IgM production and patients with locoregional disease developed more robust IgG responses than advanced metastatic melanoma patients.ConclusionsWe present the first quantitative assay to measure KLH-specific Abs in human serum, which now enables monitoring both the dynamics and absolute concentrations of humoral immune responses in individuals exposed to KLH. This assay may provide a valuable biomarker for the immunogenicity and clinical effectiveness of KLH-containing vaccines and therapies.Electronic supplementary materialThe online version of this article (doi:10.1007/s00262-012-1263-z) contains supplementary material, which is available to authorized users.

Species-Specific and Cross-Reactive IgG1 Antibody Binding to Viral Capsid Protein 1 (VP1) Antigens of Human Rhinovirus Species A, B and C

“…The total IgG1 binding by the HRV-A and -C antigens varied over a 2-log range but at high titre; the highest titres probably reflect antibodies induced by multiple infections with related viruses and recent infection causing transient elevations. The range of binding of specific titres at 250 - 1×10 7 ng/ml is similar to values obtained for other microbial and vaccine antigens [24], [31]–[34]. All the antibody binding could be competitively inhibited with low concentrations of the homologous antigen.…”

Skin-Test Infiltrating Lymphocytes Early Predict Clinical Outcome of Dendritic Cell–Based Vaccination in Metastatic Melanoma

“…Secondly, these immunogenic proteins contain a multitude of predominantly T helper cell epitopes; ergo, it functions as a nontumor specific adjuvant. We have previously investigated the magnitude and dynamics of humoral responses in this cohort of metastatic melanoma patients who underwent DC-based therapy (27). Our findings show that humoral anti-KLH responses are dictated by different vaccination parameters, such as route of administration and anti-CD25 mAb pretreatment.…”

Effective Clinical Responses in Metastatic Melanoma Patients after Vaccination with Primary Myeloid Dendritic Cells

“…Samples were tested for the presence of KLH-specific T cells by proliferation and 3 H-thymidine incorporation, and the presence of KLH-specific antibodies in serum using ELISA (22).…”