We purified a hybrid cytokine that contains interleukin-7 (IL-7) and the beta-chain of hepatocyte growth factor (HGFβ) from a unique long-term murine bone marrow culture system. We have cloned and expressed the human form of IL-7/HGFβ in which the IL-7 and HGFβ genes are connected by a flexible linker to produce a single-chain recombinant human IL-7/HGFβ protein (hrIL-7/HGFβ). To determine whether hrIL-7/HGFβ has antitumor activity, we injected this hybrid cytokine into melanoma and colon cancer animal models, and then assessed the local tumor growth and tumor metastasis. We show here that administration of hrIL-7/HGFβ significantly inhibited the growth and metastasis of malignant melanoma and colon cancer in mice. The antitumor activity was involved in a marked increase in the number of tumor-infiltrating CD4 and CD8 T cells and activated dendritic cells. The immunological mechanism by which hrIL-7/HGFβ inhibits tumor growth was confirmed by its inability to inhibit tumor growth and in immunodeficient mice. Furthermore, immune cells from hrIL-7/HGFβ-treated cancer-bearing mice can be adoptively transferred into naïve mice to resist same tumor cell challenge. Therefore, hrIL-7/HGFβ has potential applications in the treatment of cancer patients.