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Background Nilogen Oncosystems’ 3D-EXact ex vivo tumoroid platform preserves key features of the tumor microenvironment, enabling assessment of patient-specific responses to clinically relevant therapies. In this study, we evaluated the feasibility of using fresh patient-derived ovarian serous carcinoma tumoroids to predict the efficacy of first-line chemotherapeutic and immuno-oncologic treatment regimens.Methods Tumoroids approximately 150 µm in size were generated using a proprietary mechanical dissociation process, without enzymatic digestion or in vitro propagation, from freshly resected ovarian serous carcinoma tissue collected with appropriate patient consent. Ex vivo treatment was conducted with carboplatin, docetaxel, pembrolizumab, and the selective estrogen receptor modulator tamoxifen – both singly and in combinations. High-content confocal imaging was used to quantify tumor cell death. Multi-parameter flow cytometry characterized treatment-induced changes in tumor-resident T cell populations, and multiplex cytokine assays were performed on culture supernatants to assess treatment-associated cytokine responses. Additional studies using cryo-preserved tumoroids from the same patient were employed to repeat the study and to understand the correlation of results with first-round findings.Results Confocal imaging revealed substantial tumor cell death in response to both single-agent and combination treatment with carboplatin and docetaxel. Multiparameter flow cytometry demonstrated a heterogeneous immune microenvironment, including diverse myelomonocytic and leukocytic lineages, with minimal alterations in immune cell composition following treatment. Notably, PD-1 was highly expressed on both CD8 + and CD4+ tissue-resident T cells, suggesting chronic antigen exposure and potential T cell exhaustion. Cytokine profiles mirrored the tumoricidal effects observed via imaging, supporting the functional relevance of treatment-induced responses.Conclusions These results support the utility of the 3D-EXact platform in identifying effective therapeutic regimens for individual patients. In this case, carboplatin and docetaxel demonstrated potent tumoricidal activity in ovarian cancer tumoroids, a response that was recapitulated in cryopreserved tumoroids from the same patient. This approach offers a valuable tool for optimizing personalized cancer treatment while minimizing exposure to ineffective therapies.Ethics Approval Ohio State University Ethics Board: 2014H0130. Informed consent was collected prior to tissue collection.