342 (PB122) - Neutralizing antibody for S100A8/A9 soil sensing signal to prevent metastatic disease

Abstract

Organotropism of cancer metastasis is driven by the ‘seed and soil’ theory where the soil (metastatic microenvironment) provides signals to attract the seeds (cancer cells) to grow on a certain location. S100A8/A9 is a ‘soil sensing signal’ that binds to receptors expressed by many cancer types. We aimed to understand the role of S100A8/A9 in melanoma, breast and lung cancer, to develop a neutralizing antibody, and to provide a proof-of-concept for the antibody for inhibiting metastasis. Cell lines were obtained from ATCC. Overexpression of S100A8/A9 receptors MCAM and NPTNβ was induced by cDNA vector and MCAM expression was silenced by a siRNA. Recombinant S100A8/A9 protein was produced in-house. Migration/invasion assays were performing with Boyden Chamber method, colony formation assay with agarose gel cultures, and the results were quantitated by counting crystal violet stained cells/colonies. mRNA expression was analyzed by qPCR. In mouse lung metastasis models, cancer cells were injected into tail vein, and lung metastases were monitored by X-ray Computed Tomography and quantitated by counting metastatic foci at endpoint. A chimeric S100A8/A9 antibody was produced in CHO-S cells by incorporating genomic sequences of heavy and light chains of a mouse monoclonal S100A8/A9 antibody to the Fab domain of human cDNA-encoded IgG2-Fc part. S100A8/A9 was expressed in monocytes and attracted cancer cells expressing its receptors to immunosuppressed premetastatic niche. Of these receptors, MCAM was highly expressed in metastatic, but not in non-metastatic melanoma and breast cancer cells. Overexpression of MCAM in non-metastatic melanoma cells increased migration, and downregulation in metastatic cells eliminated migration, but also resulted in lost sensitivity to S100A8/A9 stimulation. In breast cancer cells, overexpression of MCAM in non-metastastic cells increased migration, which was abrogated by addition of S100A8/A9 decoy receptor to the cultures. Overexpression of MCAM induced EMT-like expression profile in the breast cancer cells. In lung cancer, overexpression of S100A8/A9 receptor NPTNβ induced anchorage independent growth, migration and invasion, which were further increased by addition of S100A8/A9 ligand to the cultures. From the produced antibodies, the clones that efficiently bound S100A8 and A9 heterodimer most effectively inhibited migration and cytokine production. The S100A8/A9 antibody effectively prevented formation of lung metastases in melanoma and breast cancer lung metastasis models. These results demonstrate a role for S100A8/A9 and its receptors in metastasis of melanoma, breast and lung cancer. A neutralizing antibody for S100A8/A9 prevented formation of lung metastases, suggesting pharmacological inhibition of S100A8/A9 as a potential option for preventing metastatic disease. No conflict of interest.