Immune checkpoint inhibitors have drawn a consider attention as an effective cancer immunotherapy, and several monoclonal antibodies targeting the immune checkpoint receptors, such as human programmed cell death-1 (hPD-1) and cytotoxic T-lymphocyte-associated protein 4 (CTLA-4), are clinically used for treatment of various cancers. Here we present the development of a small-sized protein binder which specifically binds to hPD-1. The protein binder, which is composed of leucine-rich repeat (LRR) modules, was selected against hPD-1 through phage display, and its binding affinity was maturated up to 17 nM by modular evolution approach. The protein binder was shown to be highly specific for hPD-1, effectively inhibiting the interaction between hPD-1 and its ligand, hPD-L1. The protein binder restored T-cell function in vitro, and exhibited a strong anti-tumour activity in tumour mouse model, indicating that it acts as an effective checkpoint blockade. Based on the results, the developed protein binder specific for hPD-1 is likely to find a potential use in cancer immunotherapy.