Hematopoietic stem cells, kind of adult stem cells, have a potential for differentiating into almost every blood forming cells (including erythrocytes, leucocytes, platelets and lymphocytes) and are constantly auto-regenerated as immune cells from the hematopoietic stem cell in bone marrow. The cells forming immune system, in particular natural killer cells (referred as “NK cells” hereinafter), have an ability to kill tumor cells non-specifically.
This tumor cell killing activity of NK cells has been used for the treatment of a solid tumor using lymphokine activated killer cells (LAK) and tumor infiltration lymphocytes (TIL) or for the immune therapy via donor lymphocyte infusion (Itoh K, et al., J. Immunol., 136: 3910-3915, 1986; Bordignon C, et al., Hematologia, 84: 1110-1149, 1999), which has drawn our attention as an advanced cell therapy for preventing rejection from bone marrow transplantation or organ transplantation. It was also reported that the defect in NK cell differentiation or activity is related to various diseases including breast cancer (Konjevic G, et al., Breast Cancer Res. Treat., 66: 255-263, 2001), melanoma (Ryuke Y, et al., Melanoma Res., 13: 349-356, 2003) and lung cancer (Villegas F R, et al., Lung Cancer, 35: 23-28, 2002). So, correction of NK cell functions seems to lead the way to treat these diseases.
NK cells are derived from hematopoietic stem cells in bone marrow. The differentiation from hematopoietic stem cells into NK cells is composed of many steps, which have not been completely explained, yet.
IL-21 (Interleukin-21) is a cytokine secreted by activated CD4+ T cells (Warren J., et al., Nature, 5: 688-697, 2005). IL-21 receptor (IL-21R) is expressed in lymphocytes such as dendrite cells, NK, T, and B cells (Takaki R., et al., J. Immunol., 175: 2167-2173, 2005). The structure of IL-21 is very similar to those of IL-2 and IL-15, and IL-21R shares γ-chain with IL-2R, IL-15, IL-7R or IL-4R (Ensminger S M, et al., J. Immunol. 167 (1):532-541, 2001).
According to the previous reports, IL-21 induces maturation of NK cell precursor in bone marrow (Parrish-Novak J., et al., Nature, 408: 57-63, 2000) and characteristically increases effector functions of NK such as cytokine generation ability and killing activity (Strengell M, et al., J. Immunol., 170: 5464-5469, 2003; Brady J, et al., J. Immunol., 172: 2048-2058, 2004). IL-21 also increases effector functions of CD8+ T cells, leading to the promotion of anticancer response of innate or adaptive immune system (Takaki R., et al., J. Immunol., 175: 2167-2173, 2005; Moroz A., et al., J Immunol, 173: 900-909, 2004). In addition, IL-21 activates NK cells separated from human peripheral blood (Parrish-Novak J., et al., Nature, 408: 57-63, 2000) and plays an important role in differentiation of NK cells from hematopoietic stem cells separated from umbilical cord blood (Sonia A. P, et al., Int. immunol., 18: 49-58, 2006).
YC-1 [3-(5′-hydroxymethyl-2′-furyl)-1-benzylindazole] was first developed as an agent for treating circulatory diseases after it was proved to inhibit coagulation of platelets and activate soluble GC (guanylyl cyclase) to inhibit contraction of blood vessel (Ko F N, et al., Blood 84. No. 12:4226-4233 1994; Teng C M, et al., Eur J Pharmacol, 320:161-6, 1997; Galle J., et al., Br J Pharmacol, 127: 195-203, 1999). It was additionally found in recent studies that it inhibits HIF-1α accumulation induced in hypoxia, reduces expressions of HIF-1 target genes (VEGF, erythropoietin, etc.) (Chun Y S, et al., Biochem Pharmacol, 61:947-954, 2001), and inhibits tumor growth and angiogenesis in animals (Yeo E J, et al., J Natl Cancer Inst, 95: 516-525, 2003; Pan S L, et al., J pharmacol Exp Ther, 314:35-42, 2005). So, YC-1 becomes a leading compound for the development of an anticancer agent targeting HIF-1α (Yeo E J, et al., Cancer Res, 66: 6345-6352, 2006). Even though YC-1 is very useful for the study of HIF-1α (Moeller B J, et al., Cancer cell 5: 429-441, 2004; Funasaka T, et al., FASEB J, 19:1422-1430, 2005), the mechanism of anticancer activity of YC-1 has not been disclosed, yet (Sun. H. L., et. al. Oncogene 1-11, 2006).
The present inventors completed this invention by confirming that IL-21 (Interleukin-21) and YC-1 [3-(5′-hydroxymethyl-2′-furyl)-1-benzylindazole] promote the differentiation of hematopoietic stem cells into natural killer cells and increase the killing activity of natural killer cells.