Ovarian cancer is the second leading cause of cancer-related mortality among gynaecological cancers in the Western world. CAR-T cell therapy is adoptive immunotherapy currently being used to treat some blood cancers. This study has investigated whether leucine-rich repeat-containing G protein-coupled receptor 5 (LGR5) is a suitable target for CAR-T cell therapy for ovarian cancer. The aim of the current study is to investigate the relationship between LGR5 protein and mRNA expression and clinical outcomes and chemotherapy resistance in ovarian cancer patients by immunohistochemistry, western and qRT-PCR. We assessed the cytotoxic effects of LGR5-CAR-T cells on a range of ovarian cancer cell lines and primary serous ovarian cancer cells derived from patient ascites in monolayer culture, 3D spheroid assays and patient-derived explant assays. OVCAR3 and patient-derived xenograft (PDX) models have been used for assessing whether LGR5 CAR-T cells can inhibit growth and metastasis in vivo. We found that LGR5 is highly expressed in high-grade serous ovarian carcinoma (HGSOC) cancer cell lines and chemo-resistant relapse tissues. We demonstrated that LGR5-CAR-T cells were cytotoxic and significantly inhibited the survival of ovarian cancer cells lines and chemotherapy-resistant primary ovarian cancer cells with high LGR5 express levels in both monolayer culture and 3D spheroids compared to un-transduced CD3+ T cells. In patient-derived explant assays, we observed increased apoptosis in HGSOC tissues expressing high LGR5. In vivo, LGR5-CAR-T cell treatment decreased the tumour burden in mice with OVCAR3 xenografts and a chemotherapy-resistant patient-derived xenograft with high LGR5 expression, compared to CD3+ T cells. In summary, LGR5-CAR-T cells have great potential to be developed as a novel immunotherapy for ovarian cancer.