Erythropoietin and Cyclophosphamide Combined Treatment Additively Enhances Immunoglobulin Production in Mice

Erythropoietin (EPO) is an important component in the treatment of cancer patients for improvement of cancer related anemia. EPO treatment for cancer related anemia is usually combined with chemotherapy. Cyclophosphamide (CP) is a known cytotoxic alkylating agent widely used in cancer chemotherapy. While at high doses it functions as an immunosuppressive agent, the anti-neoplastic activities of CP at low doses are attributed to enhancement of cellular and humoral immunity e.g. (Berd et al., Cancer Res; 1984). We have previously shown that EPO displays anti-neoplastic activities (Mittelman, 2001, 2004) and that EPO treatment is associated with enhancement of both the humoral and cellular immune responses (Prutchi-Sagiv 2006, Katz 2007). Here we focused on a murine model of DNP-KLH-injection, used to assess the humoral response in mice. Recently we demonstrated that administration of high doses of EPO (180U×3) to DNP-KLH-injected C57BL mice resulted in an increase in anti-DNP immunoglobulin G1 (IgG1) production. The present study was designed to examine the effect of combining low dose CP (12.5mg/kg ×2) used to achieve an anti-neoplastic activity, with a lower dose of recombinant human EPO (rHuEPO; 90U×3) on the humoral immune response of the DNP-KLH-injected mice, thus simulating the conditions of patient care. Hence, we compared anti-DNP Ig serum levels in DNP-KLH-injected C57BL mice that were treated with either EPO or CP alone, or the combination of CP and EPO (CP-EPO). CP treatment alone resulted in increased levels of serum anti-DNP IgG1 (O.D._(CP) = 0.38±0.06 vs O.D._{(Non treated)} = 0.18±0.064). In contrast, EPO treatment alone enhanced serum levels of IgG2 (O.D._(EPO) = 0.47±0.09 vs O.D._{(Non treated)} = 0.18±0.069). CP or EPO alone did not affect the total levels of anti-DNP total Ig (O.D._(EPO) = 0.37±0.07 vs O.D._{(Non treated)} = 0.28±0.04). Yet, the combined CP-EPO treatment resulted in increased levels of anti-DNP total Ig (O.D._(EPO+CP) = 0.48±0.05 vs O.D._{(EPO or CP)} = 0.37±0.04), maintaining the higher levels of IgG1 (O.D._(EPO+CP) = 0.38±0.06) and IgG2 (O.D._(EPO+CP) = 0.49±0.1). In summary, the combined CP-EPO treatment additively improved immunoglobulin production, compared to treatment with CP or EPO alone. We thus demonstrate that in context of chemotherapy treatment as usually administered in the clinic, EPO can enhance humoral immunity alongside its erythropoietic activities. Our findings emphasize the role of EPO as an immunomodulator, particularly when given as treatment in a combined therapeutic panel