Abstract

Background

Proteins within the complement system have complex effects on cellular immune responses. In previous studies, we found that active complement components, especially C5a, can dampen the development of antigen-specific immune responses following vaccination with a model antigen, in part by promoting generation of APC-induced T regulatory (Treg) cells. These studies also demonstrated that B lymphoma cell lines exposed to complement can induce Treg generation in vitro. The current study was designed to address whether depletion of C5a could enhance development of a cellular anti-lymphoma immune response in vivo.

Methods

Immunocompetent Balb/C mice were inoculated subcutaneously with syngeneic A20 B lymphoma cells mixed with either 10 μg of rat anti-mouse C5a monoclonal antibody (mAb) or 10 μg of isotype-matched Rat IgG2a control mAb. Tumor growth was followed. In select experiments, mice were sacrificed and analyzed for the percentage and activity of tumor-infiltrating T cells and A20-specific splenic T cell responses.

Results

1. Tumor progression. Lymphoma grew more slowly in mice treated with anti-C5a mAb compared to mice treated with control mAb (p<0.05) {Fig. 1).

2. Intratumoral T cells. Tumors from mice treated with anti-C5a mAb had higher CD8+ T cell infiltration compared to mice treated with control mAb (p=0.002) (Fig. 2). Tumor-infiltrating CD8+ T cells showed a trend towards higher intracellular IFNg production in mice treated with anti-C5a mAb compared to control mAb (p=0.051).

3. Splenic T cells. Splenic T cells from mice treated with anti-C5a mAb produced IFNg to a greater degree than did splenic T cells from control mice when splenocytes were cultured with irradiated A20 cells in vitro (p=0.041) (Fig. 3). There was a trend towards decreased numbers of splenic CD4+CD25highFoxp3+ Tregs in C5a-depleted mice compared to control mice.

Conclusions

Depletion of C5a at the site of tumor inoculation slows tumor growth and increases the number of tumor infiltrating CD8 T cells in a syngenic immunocompetent model of lymphoma. A trend towards enhanced production of IFNg in the tumor infiltrating T cells, increased numbers of tumor-specific splenic T cells, and reduced numbers of splenic Tregs, suggests intratumoral C5a depletion can enhance tumor-specific immune responses both within the tumor and systemically. Ongoing studies are exploring the molecular mechanisms involved in C5a-promoted tumor progression and the use of C5a depletion as a novel strategy to improve anti-tumor immunity.

Disclosures:

No relevant conflicts of interest to declare.

Author notes

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Asterisk with author names denotes non-ASH members.