Plasticity and immune marker profiling of tumour- and ascites-derived microtumours in serous ovarian cancer: relevance for short-term culture models.

Introduction: Serous ovarian cancer (SOC) is characterized by tumour heterogeneity, cellular plasticity, and immune suppression, contributing to limited therapeutic responses. Understanding tumour -immune dynamics at the cellular level is essential for developing personalized treatment strategies. Methods: We established a patient-derived microtumour (MT) platform from solid tumour tissue and malignant ascites of 18 treatment-naïve SOC patients. Using multiparametric flow cytometry, we analysed plasticity-associated markers (CD10, CD24, CD44, CD90, CD133), immunoregulatory molecules (PD-L1, PVR, PVRL2, CD47, Calreticulin, CD39, CD73), and antigen presentation molecules (HLA-A/B/C, HLA-DR/DP/DQ) during short-term culture. Results: Plasticity-associated marker expression was largely conserved across compartments, with substantial interpatient heterogeneity and only modest differences in an exploratory composite plasticity score. Compared with tumour-derived MTs, ascites-derived MTs exhibited significantly higher calreticulin expression and lower expression of the ectonucleotidase CD39. Short-term culture induced CD44 upregulation and CD90 downregulation as well as dynamic modulation of immune markers, such as increased PVR and PVRL2 expression. HLA expression showed substantial interindividual heterogeneity rather than clear compartment-specific differences. Co-expression analyses revealed associations between plasticity-associated and immunoregulatory features, such as CD10/CD24 with PD-L1 and CD47. Finally, exploratory autologous co-cultures demonstrated the feasibility of combining patient-derived MTs with matched immune cells. Two of five samples showed moderate responses to anti-PD-L1 and anti-TIGIT, reflected by increased granzyme B expression and accompanied by compensatory changes in TIGIT and PD-L1 expression. Discussion: Taken together, these findings indicate an association between tumour cell plasticity and an immune-evasive phenotype in ovarian cancer and support patient-derived MTs as an exploratory short-term platform for capturing patient-specific tumour -immune interactions, while highlighting dynamic immune checkpoint molecule expression during cultivation.