Background: Ovarian cancer (OC) presents a complex landscape shaped by diverse molecular mechanisms across various histotypes. Exploring intra-tumor heterogeneity (ITH), characterized by distinct populations of molecularly or phenotypically diverse tumor cells and their microenvironments, remains a significant challenge.
Method: Digital Spatial Profiling (DSP) platform is used to explore ITH in a retrospective cohort of 10 OC patients, including 6 early-stage ovarian clear cell carcinoma (OCCC) cases, 3 advanced-stage OCCC cases, and 1 advanced-stage high-grade serous carcinoma (HGSC). Formalin-fixed paraffin-embedded (FFPE) samples from primary ovarian and metastatic sites were analyzed. A total of 1,145 regions of interest (ROIs), including 669 PanCK+ and 476 CD45+ areas of illumination (AOIs), were profiled using a 28-plex immune-related protein panel. Protein expression patterns were examined based on the presence or absence of tumor-infiltrating immune cells (TII), disease stage, and histotype.
Results: In the PanCK+ compartment, three clusters emerged: C1 (immune-cold), C2a, and C2b (immune-hot). C1, representing 50% of ROIs, showed increased OX40L, while C2a had elevated SMA, PanCK, PD-1, HLA-DR, CD8, and CD3. C2b was characterized by higher levels of Ki-67, CD56, CTLA4, and CD68. PanCK+ AOIs without CD45+ cells were predominantly in C1, reflecting its immune-cold feature. PanCK+ AOIs with either TII or non-TII were more associated with C2a. Stage 2 tumors and HGSC were mainly linked to immune-hot clusters. In the CD45+ compartment, four clusters were identified: C1 (CD8, CD3, CD20), C2 (LAG3), C3a (SMA, Fibronectin, CTLA4), and C3b (HLA-DR, CD68, CD56, CD11c, B7-H3). C1 was more prevalent in non-TII samples, while C2 was dominant in Stage 1. OCCC was primarily linked to C1 and C2, while HGSC was associated with C3a and C3b.
Conclusion: OC exhibits extensive ITH in both tumor and immune cell compartments, with distinct associations based on disease stage and histotype.