Objective To investigate the effect of a neoadjuvant treatment strategy of an anti-PD-1 monoclonal antibody combined with cisplatin on postoperative recurrence and metastasis of non-small cell lung cancer (NSCLC) in a humanized mouse model. Methods Human peripheral blood mononuclear cells and human NSCLC cells were sequentially transplanted into the severe combined immunodeficiency mouse (NOD/ ShiLtJGpt-Prkdcem26Cd52 Il2rgem26Cd22 / Gpt, NCG) to establish an immune system-tumor double humanized NSCLC model. We elevated the treatment effect of various neoadjuvant strategies in the double humanized NSCLC model, including cisplatin, anti-PD-1 ( pembrolizumab), and combined treatments. Tumor recurrence and metastasis were observed by optical imaging. Ki67 expression in tumor tissue was analyzed by immunohistochemistry to assess tumor cell proliferation. Changes in the tumor immune microenvironment were detected by immunofluorescence. Results The double humanized NSCLC model was successfully established. Immune cells had infiltrated into peripheral blood, organs, and tumors. Optical imaging showed that neoadjuvant anti-PD-1 treatment inhibited tumor recurrence and metastasis, and the effect of combined treatment was more obvious. Moreover, combination treatment significantly decreased Ki67 expression in tumor tissues. Large numbers of CD8⁺ and CD45⁺ cells in tumors were observed after anti-PD-1 treatment, and many more CD8⁺ and CD45⁺ T cells were found in the combined therapy group. Granzyme B plays an important role in tumor killing by CD8⁺ T cells. Immunohistochemical staining showed that anti-PD-1 treatment increased granzyme B in tumor tissues, which was more obvious after combined treatment. Conclusions Neoadjuvant pembrolizumab combined with cisplatin effectively improves the anti-tumor effect in humanized mice and significantly inhibits tumor recurrence and metastasis.