Genomic and expression profiling reveal molecular heterogeneity of disseminated tumor cells in bone marrow of early breast cancer

Abstract

Detection of disseminated tumor cells (DTCs) in bone marrow is an established negative prognostic factor. We isolated small pools of (~20) EPCAM-positive DTCs from early breast cancer patients for genomic profiling. Genome-wide copy number profiles of DTC pools (n = 45) appeared less aberrant than the corresponding primary tumors (PT, n = 16). PIK3CA mutations were detected in 26% of DTC pools (n = 53), none of them were shared with matched PTs. Expression profiling of DTC pools (n = 30) confirmed the upregulation of EPCAM expression and certain oncogenes (e.g., MYC and CCNE1), as well as the absence of hematopoietic features. Two expression subtypes were observed: (1) luminal with dual epithelial–mesenchymal properties (high ESR1 and VIM/CAV1 expression), and (2) basal-like with proliferative/stem cell-like phenotype (low ESR1 and high MKI67/ALDH1A1 expression). We observed high discordance between ESR1 (40%) and ERRB2 (43%) expression in DTC pools vs. the clinical ER and HER2 status of the corresponding primary tumors, suggesting plasticity of biomarker status during dissemination to the bone marrow. Comparison of expression profiles of DTC pools with available data from circulating tumor cells (CTCs) of metastatic breast cancer patients revealed gene expression signatures in DTCs that were unique from those of CTCs. For example, ALDH1A1, CAV1, and VIM were upregulated in DTC pools relative to CTCs. Taken together, analysis of pooled DTCs revealed molecular heterogeneity, possible genetic divergence from corresponding primary tumor, and two distinct subpopulations. Validation in larger cohorts is needed to confirm the presence of these molecular subtypes and to evaluate their biological and clinical significance.

Synchronous Detection of Circulating Tumor Cells in Blood and Disseminated Tumor Cells in Bone Marrow Predicts Adverse Outcome in Early Breast Cancer

“…Abbreviation: HRS, HR-positive status. CellSearch (13-17, 25, 26) and IE/FC (18)(19)(20)(21)(22). Our previous studies in triple-negative metastatic breast cancer have demonstrated high concordance between these methods (20).…”

“…Blood and bone marrow samples were subjected to the IE/FC assay to enumerate CTCs and DTCs, respectively (18)(19)(20)22). Briefly, two distinct mAbs against EPCAM, one conjugated to immunomagnetic beads (MJ37) and the other conjugated to phycoerythrin (EBA-1) were added to whole blood or bone marrow.…”

“…We have described an EPCAM-based immunomagnetic enrichment/flow cytometry (IE/FC) for enumeration and isolation of CTCs (18)(19)(20) in blood, and have applied this method as well to DTCs (18,21,22) in bone marrow.…”

Molecular Interplay between Dormant Bone Marrow-Resident Cells (BMRCs) and CTCs in Breast Cancer

“…Expression and functionality of the mTOR pathway were found to be associated with the proliferative switch of BMRC cells. Multiple oncogenic mutations are known to characterize benign breast cancers that do not metastasize [30,50]. Conversely, the activation of some oncogenic pathways repress BC metastasis, but increase tumor cell proliferation [3,50].…”

Predicting therapy response by analysis of metastasis founder cells: emerging perspectives for personalized tumor therapy

“…In melanoma, the presence of BRAFV600E mutation was inconsistent between PT and LN-derived DCCs [28]. Discordance between DCCs and matched PT has also been reported for the expression of estrogen receptor as well as the presence of PI3K mutations in breast cancer [30][31][32]. With regard to HER2 expression, discordance between PT has been observed for both DCCs and CTCs in non-metastatic breast cancer patients [33][34][35] indicating that adjuvant therapies could be improved by implementation of routine CTC/DCC-based stratification.…”