The role of dendritic cell precursors in tumour vasculogenesis

In this review, we discuss the recent identification in vivo of a population of CD11c+ cells exhibiting simultaneous expression of both endothelial and dendritic cell markers, termed vascular leukocytes (VLCs). VLCs are highly represented in human ovarian carcinomas and, depending on the milieu, can assemble into functional blood vessels or act as antigen-presenting cells. The identification of dendritic cell precursors as bipotent cells has important implications for the physiopathology and therapy of tumours. VLCs emerge as a novel therapeutic target against tumour vascularisation

Brugia malayi microfilariae adhere to human vascular endothelial cells in a C3-dependent manner

Brugia malayi causes the human tropical disease, lymphatic filariasis. Microfilariae (Mf) of this nematode live in the bloodstream and are ingested by a feeding mosquito vector. Interestingly, in a remarkable co-evolutionary adaptation, Mf appearance in the peripheral blood follows a circadian periodicity and reaches a peak when the mosquito is most likely to feed. For the remaining hours, the majority of Mf sequester in the lung capillaries. This circadian phenomenon has been widely reported and is likely to maximise parasite fitness and optimise transmission potential. However, the mechanism of Mf sequestration in the lungs remains largely unresolved. In this study, we demonstrate that B. malayi Mf can, directly adhere to vascular endothelial cells under static conditions and under flow conditions, they can bind at high (but not low) flow rates. High flow rates are more likely to be experienced diurnally. Furthermore, a non-periodic nematode adheres less efficiently to endothelial cells. Strikingly C3, the central component of complement, plays a crucial role in the adherence interaction. These novel results show that microfilariae have the ability to bind to endothelial cells, which may explain their sequestration in the lungs, and this binding is increased in the presence of inflammatory mediators

Detection of mRNA for the terminal complement components C5, C6, C8 and C9 in human umbilical vein endothelial cells in vitro.

Human umbilical vein endothelial cells (HUVEC) have previously been shown to synthesize the functional terminal pathway of complement based on the detection by radioimmunoassay of the terminal complement complex (TCC) on coincubated agarose beads. In addition, C7 secretion by these cells in amounts comparable to C3, as well as C7 mRNA, has recently been demonstrated. However, it has not been possible to detect C5-6 and C8 in the fluid phase, and only trace amounts of soluble C9. Against this background we examined whether mRNA for the remaining terminal complement factors was present in HUVEC. By the use of reverse transcription (RT)-polymerase chain reaction (PCR) and Northern blot the presence of mRNA for complement factors C5, C6, C8 and C9 was demonstrated

The endothelium is an extrahepatic site of synthesis of the seventh component of the complement system

The level of the terminal complement components secreted by human umbilical vein endothelial cells (HUVEC) was measured by a sensitive ELISA which allows the detection of 30–50 pg/ml of these components. C7 was the only terminal component detected in measurable amounts in the cell supernatant. The mean value was 11 ng/106 cells at 96 h and was slightly higher than that of C3 (9 ng/106 cells). HUVEC and serum C7 analysed by SDS–PAGE and immunoblot exhibited the same electrophoretic mobility. A proportion of C7 secreted by HUVEC was incorporated into the terminal complement complex (TCC) assembled spontaneously in the supernatant of cells cultured in C7-deficient human serum, and was not detected by the standard ELISA for C7 measurement. By adding the amount of C7 present in the TCC to that of free C7, the total amount of the component released by HUVEC was calculated to be approximately 35 ng/106 cells. Further TCC was produced following complement activation of the cell supernatant through the alternative pathway. Synthesis of C7 by HUVEC was confirmed by inhibition experiments in the presence of cycloheximide and by reverse transcriptase-polymerase chain reaction (RT-PCR) analysis of C7 mRNA expression. Addition of IL-1α and tumour necrosis factor-alpha to the cell culture stimulated the secretion of C3, but had no effect on the synthesis of C7. By contrast, interferon-gamma had only a marginal effect on the production of C3, but markedly down-regulated the synthesis of C7 as assessed both by ELISA and RT-PCR

Bispecific antibodies targeting tumor-associated antigens and neutralizing complement regulators increase the efficacy of antibody-based immunotherapy in mice

The efficacy of antibody-based immunotherapy is due to the activation of apoptosis, the engagement of antibody-dependent cellular cytotoxicity and complement-dependent cytotoxicity (CDC). We developed a novel strategy to enhance CDC using bispecific antibodies (bsAbs) that neutralize the C-regulators CD55 and CD59 to enhance C-mediated functions. Two bsAbs (MB20/55 and MB20/59) were designed to recognize CD20 on one side. The other side neutralizes CD55 or CD59. Analysis of CDC revealed that bsAbs could kill 4-25 times more cells than anti-CD20 recombinant antibody in cell lines or cells isolated from patients with chronic lymphocytic leukemia. The pharmacokinetics of the bsAbs was evaluated in a human-SCID model of Burkitt lymphoma. The distribution profile of bsAbs mimics the data obtained by studying the pharmacokinetics of anti-CD20 antibodies, showing a peak in the tumor mass 3-4 days after injection. The treatment with bsAbs completely prevented the development of human/SCID lymphoma. The tumor growth was blocked by the activation of the C cascade and by the recruitment of macrophages, polymorphonuclear and natural killer cells. This strategy can easily be applied to the other anti-tumor C-fixing antibodies currently used in the clinic or tested in preclinical studies using the same vector with the appropriate modifications