Abstract 586: Expression of CD68 on CKpos/CD45pos circulating cells in breast cancer patients: An additional hint supporting the fusion hybrid theory
Authors: Reduzzi Carolina , D'Amico Paolo , Zhang Youbin , Cappelletti Vera , Cristofanilli Massimo 
Published: July 2021

Abstract
Background: Cells expressing both cytokeratyins (CK) and CD45 (dual-positive, DPcells), circulating in the blood of patients with solid tumors, have been reported for different cancers. Their origin is not clear yet, but one hypothesis is that they are hybrids, deriving from the fusion of tumor cells and macrophages. Recently, we have demonstrated that DPcells can have aberrant genomes, and that, in metastatic breast cancer, they are associated with shorter progression free survival. However, a direct proof of their origin from tumor/macrophage fusion is lacking. Here, we investigated the expression of a macrophage marker (CD68) on DPcells in breast cancer patients.
Methods: Blood samples collected from BC patients were analyzed using the CellSearch platform, which allowed for the evaluation of the expression of CK, CD45 and CD68 on circulating cells enriched for EpCAM expression. Based on the marker expression, cells were classified as circulating tumor cells (CTCs, CKpos/CD45neg), and DPcells (CKpos/CD45pos).
Results: Twenty-six blood samples were collected from 26 patients with luminal, HER2+, and triple negative BC (n = 13, 5, and 8, respectively). Of these, 6 had non-metastatic (NM) BC (i.e. early breast cancer or locally advanced breast cancer), and 20 had metastatic BC (MBC). Overall, 43 DPcells were identified in 14 (54%) samples (3/6 from NMBC and 11/20 from MBC patients). CD68 expression was detected on 26 DPcells (60%) from 13 samples. In the majority of cases (7/13 samples), CD68pos and CD68neg DPcells were detected together, whereas, in 5 samples, CD68pos DPcells only were present. CD68neg DPcells alone were detected in a single sample. Among the 20 MBC cases, 11 were classified as stage IVaggressive and 9 as stage IVindolent (i.e. with ≥ 5 CTCs and &lt; 5 CTCs, respectively). CD68pos DPcells were present in 5/11 stage IVaggressive and in 5/9 stage IVindolent patients, suggesting that they are independent from CTC-based classification. As expected, CD68 was not detected on the majority of CTCs (796 out of 841), but, interestingly, 5% of CTCs were CD68pos.
Conclusions and future perspectives: These preliminary results suggest that DPcells can derive from heterotypic cell fusion between tumor cells and macrophages. This would provide a new perspective on the unique interaction between tumor cells and the microenvironment and the potential role of these new cell entity in promoting disease progression. However, many questions about this subpopulation of circulating cells remain unanswered and future studies are needed to better characterize DPcells at both phenotypic and genomic level, to understand their biological and clinical relevance.
Citation Format: Carolina Reduzzi, Paolo D'Amico, Youbin Zhang, Vera Cappelletti, Massimo Cristofanilli. Expression of CD68 on CKpos/CD45pos circulating cells in breast cancer patients: An additional hint supporting the fusion hybrid theory [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 586.