Polyaneuploid Cancer Cells (PACCs) as metastasiscompetent cells

Abstract Metastasis is responsible for 90% of cancer deaths, yet on a cellular level, successful metastasis is rare. Barriers to metastasis exist at every step of the metastatic cascade (invasion, intravasation, survival in the circulation, extravasation, and colonization.) Modeling predicts that only one of every 1.5 billion circulating tumor cells results in clinically-detectable metastasis. Identification of the rare subset of metastasis-competent cells is the most crucial goal of cancer research. Polyanuploid Cancer Cells (PACCs) are physically enlarged, treatment-resistant cells with increased genomic content that form in response to stress. After stress-induction, PACCs exist in a non-proliferative state for many weeks before undergoing eventual depolyploidization to produce progeny of “typical” cancer cell size, morphology, and nuclear content. PACCs can be identified in both primary and metastatic patient tumor tissues. As such, we propose Polyaneuploid Cancer Cells (PACCs) are metastasis-competent cells. Single-cell tracking reveals that PACCs are more motile than nonPACCs. Additionally, we observe that PACCs exhibit increased directional migration in 2D chemotactic environments. Optical-tracking of spontaneous bead motion reveals that PACCs demonstrate increased cytoskeletal rearrangement, an observation that aligns with increased environment-sensing and directional motility. In total, this predicts successful invasion. Analyses of cellular deformability using Magnetic Twisting Cytometry and Atomic Force Microscopy jointly reveal that cells in the PACC state display hyper-elastic properties. Among these include increased peripheral deformability and maintained peri-nuclear cortical integrity, both of which predict successful intravasation and extravasation. Functional deformability of PACCs navigating through narrow channels in a chemotactic environment was confirmed using a custom microfluids device. RTqPCR, NanoString mRNA quantification, Western blot, and immunofluorescent imaging reveal that PACCs highly overexpress Vimentin, a cytoskeletal component known to confer hyper-elasticity. Notably, there is no correlation between Vimentin content and motility dynamics in PACCs, indicating that the role of Vimentin in PACCs may primarily drive increased hyper-elasticity rather than increased motility. Anoikis-resistance assays and detection of PACCs in the blood of a patient with metastatic prostate cancer using a selection-free circulating tumor cell detection platform reveal that PACCs are capable of surviving in the circulation. Our work to date reports that PACCs demonstrate increased motility, environment-sensing, hyper-elasticity, and anoikis-resistance. Taken together with the knowledge that PACCs exist in a treatment-resistant state and are capable of eventual depolyploidization (as a potential route to successful colony formation), this data suggests PACCs are a candidate rare metastasis-competent cell type worthy of further analysis. Citation Format: Mikaela M. Mallin, Nicholas Kim, Mohammad Ikbal Choudhury, SeJong Lee, Steven S. An, Sean X. Sun, Konstantinos Konstantopoulos, Sarah R. Amend, Kenneth J. Pienta. Polyaneuploid Cancer Cells (PACCs) as metastasis-competent cells [abstract]. In: Proceedings of the AACR Special Conference: Cancer Metastasis; 2022 Nov 14-17; Portland, OR. Philadelphia (PA): AACR; Cancer Res 2022;83(2 Suppl_2):Abstract nr A017.