Evidence for pathogenic effect of anti-recoverin antibodies in Cancer Associated Retinopathy (CAR). In vitro study

Purpose. To study the role of anti-recoverin antibodies in cancer associated retinopathy (CAR). Recoverin was identified as an autoantigen in CAR and anti-recoverin antibodies were found in the sera of some CAR patients. Methods. Human, rat, or rabbit antibodies against recoverin were purified using a recoverin-affinity column. Titer and specificity of antibodies were measured by ELISA. Normal human, rat, and rabbit IgG were used in control experiments. Purified antibodies were cultured with a recoverin positive rat retinal cell line EIA-NR3 (10⁵ cells/ml). Antibody uptake in vitro was analyzed by immunocytochemistry. Cytotoxic effect of antibodies on retinal cells was measured by MTT colorimetric method. Results. Our data demonstrated that anti-recoverin antibodies obtained from a CAR patient's serum or from sera of immunized animals were taken up by photoreceptor cells. Interestingly, all antibodies showed similar fine specificities to recoverin in a pin ELISA test. Moreover, these antibodies produced destruction of the cells in a dose and time-dependent manner. Normal IgG did not have such affects on retinal cells. Conclusions. Our studies showed that antibodies specific to recoverin are able to enter and cause death of cells expressing recoverin. In humans, the autoantibodies originally elicited against recoverin expressed in tumor cells may damage the retinal photoreceptors and play a role in pathogenesis of CAR.