RESUMEN
Immunogenic cell death is a cell death modality that stimulates the immune system to combat cancer cells. IMMUNEPOTENT CRP (ICRP) is a mixture of substances of low molecular weight obtained from bovine spleens that exhibits in vitro cytotoxic activity on different tumor cell lines and modulates the immune response in vivo. The aim of the present study was to determine whether the cytotoxic effect of ICRP and its combination with oxaliplatin (OXP) on murine melanoma B16F10 cells was due to immunogenic cell death. The cytotoxic assay was performed using flow cytometry to detect Annexin V and propidium iodide staining, and calreticulin (CRT) exposure. Adenosine triphosphate, heat shock protein (HSP) 70, HSP90 and high mobility group box 1 (HMGB1) release were identified using bioluminescence, western blot and ELISA assays, respectively. The present in vitro study demonstrated that treatments with ICRP or OXP induced cell death in a time-dependent manner, but treatment with the combination of ICRP + OXP increased the cytotoxic effect following 24 h of treatment. CRT exposure and release of adenosine triphosphate (ATP), HSP70, HSP90 and HMGB1 were induced by treatment with ICRP, and the combination of ICRP + OXP increased the exposure and release of damage-associated molecular patterns (DAMPs), while OXP treatment only induced CRT exposure, ATP and HMGB1 release. The in vivo experiments demonstrated that administration of tumor-derived DAMP-rich cell lysates derived from B16F10 cells treated with ICRP and the combination of ICRP + OXP prevented melanoma growth; however, OXP treatment did not. These results suggested that IMMUNEPOTENT CRP may be used as an agent to increase the ability of antitumor drugs to induce immunogenic cell death and prevent the growth of melanoma.
RESUMEN
Chemotherapy treatments induce a number of side effects, such as leukopenia neutropenia, peripheral erythropenia, and thrombocytopenia, affecting the quality of life for cancer patients. 5-Fluorouracil (5-FU) is wieldy used as myeloablative model in mice. The bovine dialyzable leukocyte extract (bDLE) or IMMUNEPOTENT CRP® (ICRP) is an immunomodulatory compound that has antioxidants and anti-inflammatory effects. In order to investigate the chemoprotection effect of ICRP on bone marrow cells in 5-FU treated mice, total bone marrow (BM) cell count, bone marrow colony forming units-granulocyte/macrophage (CFU-GM), cell cycle, immunophenotypification, ROS/superoxide and Nrf2 by flow cytometry, and histological and hematological analyses were performed. Our results demonstrated that ICRP increased BM cell count and CFU-GM number, arrested BM cells in G0/G1 phase, increased the percentage of leukocyte, granulocytic, and erythroid populations, reduced ROS/superoxide formation and Nrf2 activation, and also improved hematological levels and weight gain in 5-FU treated mice. These results suggest that ICRP has a chemoprotective effect against 5-FU in BM cells that can be used in cancer patients.