Serological and genetic complement alterations in infection-induced and complement-mediated hemolytic uremic syndrome.

BACKGROUND: The role of complement in the atypical form of hemolytic uremic syndrome (aHUS) has been investigated extensively in recent years. As the HUS-associated bacteria Shiga-toxin-producing Escherichia coli (STEC) can evade the complement system, we hypothesized that complement dysregulation is also important in infection-induced HUS. METHODS: Serological profiles (C3, FH, FI, AP activity, C3d, C3bBbP, C3b/c, TCC, αFH) and genetic profiles (CFH, CFI, CD46, CFB, C3) of the alternative complement pathway were prospectively determined in the acute and convalescent phase of disease in children newly diagnosed with STEC-HUS or aHUS. Serological profiles were compared with those of 90 age-matched controls. RESULTS: Thirty-seven patients were studied (26 STEC-HUS, 11 aHUS). In 39 % of them, including 28 % of STEC-HUS patients, we identified a genetic and/or acquired complement abnormality. In all patient groups, the levels of investigated alternative pathway (AP) activation markers were elevated in the acute phase and normalized in remission. The levels were significantly higher in aHUS than in STEC-HUS patients. CONCLUSIONS: In both infection-induced HUS and aHUS patients, complement is activated in the acute phase of the disease but not during remission. The C3d/C3 ratio displayed the best discrepancy between acute and convalescent phase and between STEC-HUS and aHUS and might therefore be used as a biomarker in disease diagnosis and monitoring. The presence of aberrations in the alternative complement pathway in STEC-HUS patients was remarkable, as well.

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

PURPOSE: Keyhole 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 DESIGN: Anti-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. RESULTS: KLH-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. CONCLUSIONS: We 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.

Decreased renal excretion of soluble interleukin-2 receptor alpha after treatment with daclizumab.

BACKGROUND: Daclizumab (+/-150 kD), a humanized monoclonal antibody (mAb) against the alpha-chain of the membrane-bound interleukin-2 (IL-2) receptor (IL-2R) also binds soluble interleukin-2R alpha (sIL-2R alpha; +/-45 kD), and thus may influence the glomerular filtration of sIL-2R alpha. METHODS: We have studied the influence of daclizumab on the renal excretion of sIL-2R alpha in 38 recipients of a renal transplant (32 treated with daclizumab and six controls). sIL-2R alpha was measured every 2 weeks after transplantation in serum and urine with Immulite IL-2R, a solid-phase enzyme-linked immunosorbent assay (ELISA). RESULTS: In the control population, the fractional excretion of sIL-2R alpha was relatively constant with a median value of 1.7%+/- 0.5%. In daclizumab-treated patients, sIL-2R alpha was not detectable in the urine immediately after the administration of daclizumab. sIL-2R alpha became detectable in the urine at a mean of 8 +/- 3 weeks after transplantation. In additional experiments, serum compounds were separated by size-exclusion chromatography and sIL-2R alpha was measured in the collected fractions. In the control patients, sIL-2R alpha was only present in the low-molecular-weight fractions of serum. In contrast, in daclizumab-treated patients evaluated several weeks after transplantation, sIL-2R alpha was merely detected in the high-molecular-weight fractions of serum. During follow-up there was a relative shift of sIL-2R alpha from the high- to the low-molecular-weight fractions and this coincided with normalization of sIL-2R alpha excretion. CONCLUSION: Daclizumab inhibits the renal excretion of sIL-2R alpha by the formation of a complex with sIL-2R alpha in serum, which is too large for glomerular filtration. Measurement of urinary sIL-2R alpha may provide information on the concentration of anti-IL-2R alpha mAb in serum.