Brief Summary of the Paper:
This paper examines the influence of circadian rhythms on the metastasis of breast cancer, specifically focusing on the role of circulating tumor cells (CTCs). It uncovers that the majority of spontaneous CTC intravasation events occur during the rest phase (sleep) rather than the active phase, with these rest-phase CTCs being highly prone to metastasis. This unexpected finding suggests a concentration of aggressive CTC generation within the rest phase, driven by the upregulation of mitotic genes influenced by key circadian rhythm hormones like melatonin, testosterone, and glucocorticoids, as well as insulin’s direct impact on tumor cell proliferation.
Detailed Summary of Key Findings and Methods:
- Temporal Dynamics of CTCs: The study found that the majority (78.3%) of CTCs in breast cancer patients were found during the rest phase of the day. This was consistent across various human and mouse breast cancer models. CTCs generated during the rest phase were more likely to intravasate and had an increased metastatic potential compared to those from the active phase.
- Metastatic Propensity: Rest-phase CTCs exhibited a significantly higher ability to metastasize compared to active-phase CTCs. This was demonstrated through bioluminescence imaging showing a higher metastatic burden from rest-phase CTCs in various mouse models.
- Gene Expression in CTCs: There was a significant upregulation of mitotic genes in rest-phase CTCs, indicating heightened cell division and metastasis readiness. Conversely, CTCs from the active phase showed increased expression related to protein synthesis.
- Influence of Circadian Rhythms and Hormones: The study indicates that circadian rhythms significantly impact the release and aggressive nature of CTCs, mediated by hormones such as melatonin, testosterone, and glucocorticoids. Furthermore, it demonstrates that insulin directly promotes tumor cell proliferation in a time-dependent manner.
- Sample Collection and Analysis: Blood samples from hospitalized women with progressive breast cancer were collected during active and rest phases. CTCs were captured using antigen-agnostic microfluidic capture and confirmed via immunofluorescence staining. Similar methodologies were applied in various mouse models representing human breast cancer.
- CTC Generation and Intravasation Analysis: The study involved time-kinetic analysis of CTC release over a 24-hour period, observing dramatic oscillations corresponding to the rest phase across different mouse models. The abundance of CTCs and their intravasation rates during different phases were quantified and compared.
- Metastatic Potential Assessment: Using xenograft mouse models, the study isolated CTCs spontaneously shed during different phases of the circadian rhythm and analyzed their metastatic potential via bioluminescence imaging. This was done for single CTCs as well as CTC clusters.
- Gene Expression Profiling: Single-cell resolution RNA sequencing was employed to investigate the gene expression profiles of CTCs isolated during rest and active phases. Differential gene expression analysis and gene set enrichment analysis were conducted to identify the critical pathways and genes involved.
The findings provide a new perspective on the temporal nature of metastatic spread in breast cancer and suggest that time-controlled treatment strategies may be beneficial due to the significant influence of circadian rhythms on cancer progression.
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