OK-432, penicillin-killed Streptococcus pyogenes, is used in treating lymphangiomas and carcinomas. We have studied in vitro the role of mononuclear phagocytes (MNPs), including purified monocytes (MOs), in the immune response to OK-432. MIP-1alpha/beta and MCP-1 secretions were assessed in whole blood (WB), peripheral blood mononuclear cells (PBMCs) and purified MOs, after in vitro stimulation with OK-432 with or without adherence for 24 hours.OK-432 stimulated MNPs to secrete MCP-1 and MIP-1alpha/beta in healthy individuals and in head and neck squamous cell carcinoma (HNSCC) patients, except for OK-432 stimulation of WB giving a minimal MIP-1alpha/beta response. Upon culture on low-attachment wells, a spontaneous chemokine secretion was observed, with an unchanged secretion following OK-432 stimulation. Inhibition of Syk kinase and/or PI-3 kinase did not significantly change the chemokine response to OK-432, except for MIP-1alpha production being increased upon Syk inhibitor addition and an increased MCP-1 response upon addition of both inhibitors. Adhesion may possibly involve beta1 and/or beta3 integrins, not beta2, whereas beta(1-3) integrins may act as co-stimulatory receptors for OK-432. Based on direct blockage of CD36 or CD18 by antibodies, MCP-1 production may be mediated by CD18 while MIP-1beta and MCP-1 production may occur upon binding to CD36.Adherent human MOs produce MCP-1 and MIP-1alpha/beta upon stimulation with OK-432. CD36 modulates MIP-1beta and MCP-1 response. Thus, to some extent OK-432 acts as a substance whereby only MOs adhered to surfaces secrete MCP-1 and MIP-1alpha/beta, in part explaining why OK-432 is suited as a biological response modifying drug.