Periodic Reporting for period 4 - INJURMET (Impact of tissue injury induced by diagnostic biopsies and surgery on cancer metastasis)
Período documentado: 2024-02-01 hasta 2025-07-31
Background: Prostate and breast cancer belong to the most frequent tumor types occurring in the European Union with an incidence rate of 139.5 and 139.0 cases per 100,000, respectively. Needle biopsy is required for diagnosis of these cancers and primary surgery to remove the malignant lesion. Biopsies and primary surgery cause local trauma of the targeted organ with potential risk of releasing tumor cells into the surrounding tissue or blood vessels. Since the systemic spread of tumor cells from the primary lesion to distant organs is a key event in cancer progression, it is of high importance to investigate the role of tissue biopsy and surgery on the spread of cancer cells and the formation of metastasis. The lack of reliable detection and characterization assays hampered the investigation of circulating tumor cell (CTC) release caused by biopsy or surgery for many years.
Objectives: This project aimed to analyze whether needle biopsies of suspicious lesions in the breast or prostate and surgical removal of the malignant lesion are accompanied by an increased release of tumor cells into the blood circulation. Moreover, we compared different CTC detection methods, characterized the biology of CTCs released into the blood and compared the results with the clinical outcome of the patients. Taken together, the INJURMET project provides a comprehensive set of data on the clinically relevant question whether biopsies and surgery may have a negative side effect in some patients through the release of metastasis-competent CTCs.
Conclusions: Our results indicate that tissue injury through either needle biopsy or surgical manipulation dislodges tumor cells that end up in the blood circulation. However, currently it is unclear whether the release of these cells increases the likelihood of future relapses or metastasis. Although our study does not argue against the safety or necessity of biopsies, it highlights a possible mechanism by which physical disruption can mobilize cancer cells and points to liquid biopsy as a tool for identifying patients who may benefit from closer follow-up or treatments aimed at blocking released tumor cells from seeding new metastases.
Objectives: This project aimed to analyze whether needle biopsies of suspicious lesions in the breast or prostate and surgical removal of the malignant lesion are accompanied by an increased release of tumor cells into the blood circulation. Moreover, we compared different CTC detection methods, characterized the biology of CTCs released into the blood and compared the results with the clinical outcome of the patients. Taken together, the INJURMET project provides a comprehensive set of data on the clinically relevant question whether biopsies and surgery may have a negative side effect in some patients through the release of metastasis-competent CTCs.
Conclusions: Our results indicate that tissue injury through either needle biopsy or surgical manipulation dislodges tumor cells that end up in the blood circulation. However, currently it is unclear whether the release of these cells increases the likelihood of future relapses or metastasis. Although our study does not argue against the safety or necessity of biopsies, it highlights a possible mechanism by which physical disruption can mobilize cancer cells and points to liquid biopsy as a tool for identifying patients who may benefit from closer follow-up or treatments aimed at blocking released tumor cells from seeding new metastases.
Release of tumor cells into the circulation during prostatic surgery: We recruited 103 men undergoing prostate removal surgery for early-stage prostate cancer. We drew blood from both the vessels draining the prostate gland and from an arm vein, collecting samples during and immediately after surgery. Interestingly, most patients had hundreds of prostate‐derived cells in the tumor-draining blood. Patients with high PSA levels before surgery were more likely to have CTCs in peripheral blood, suggesting a connection between tumor burden and the likelihood of finding cancer cells in circulation. Longer patient follow-up observations and larger trials will be essential to determine whether perioperative strategies to reduce tumor cell release can translate into better long-term outcomes for men with prostate cancer.
Release of tumor cells during breast tissue biopsy and surgery: In our study of 206 women undergoing needle or core biopsies for suspected breast lesions, we drew blood samples immediately before and after their biopsy. Of the 118 women whose biopsies confirmed cancer, 17% showed a rise in CTC number after biopsy. This increase in CTCs occurred more often in patients whose tumors lacked E-cadherin, a molecule that helps cells stick together, suggesting that tumors already prone to loosen their cell-to-cell bonds are especially likely to shed cells when disturbed (Figure 1).
In addition to the CTCs, we collected and isolated cell-free DNA from the blood plasma from women that underwent tissue biopsy. We next assessed the methylation status of a large panel of oncology related genes to identify the tumor specific DNA among the normal DNA. With this, we aim to reduce the number of invasive tissue biopsies to women with the highest likelihood of having breast cancer. Namely, from the 206 women we recruited, 43% had no malignant disease and were thus over diagnosed. This highlights the urgent need for more precise and less intrusive diagnostic tools.
Peripheral blood samples were analyzed before and shortly after breast cancer surgery from 52 breast cancer patients. Preoperatively, CTCs were detected less frequently than post-surgery (5.8 vs 13.5%). The clinical follow-up data of the patients examined in this study are currently being evaluated and will then be compared with the results of the CTC analyses.
Molecular CTC characterization: To confirm the malignant identity of CTCs detected in the above studies, we obtained 26 cells and performed whole-genome sequencing. Of the 19 cells extracted from the prostatic plexus, 10 were identified as tumor cells and 9 as normal epithelial cells; of the 7 cells from the peripheral blood, 4 were identified as tumor cells and 3 as normal epithelial cells.
Functional studies: From the CTCs of a breast cancer patient, we established a permanent cell line that we used to study tumor cell extravasation. With it, we showed that the drug bortezomib is able to impair spontaneous lung metastasis in mice, suggesting that this drug can inhibit the formation of distant metastasis. As an alternative to mouse experiments, we started to use zebrafish embryos for our investigations. We could demonstrate that as few as 50–200 injected CTCs faithfully extravasate and proliferate in zebrafish embryos highlighting its value for functional and drug-target discovery.
Release of tumor cells during breast tissue biopsy and surgery: In our study of 206 women undergoing needle or core biopsies for suspected breast lesions, we drew blood samples immediately before and after their biopsy. Of the 118 women whose biopsies confirmed cancer, 17% showed a rise in CTC number after biopsy. This increase in CTCs occurred more often in patients whose tumors lacked E-cadherin, a molecule that helps cells stick together, suggesting that tumors already prone to loosen their cell-to-cell bonds are especially likely to shed cells when disturbed (Figure 1).
In addition to the CTCs, we collected and isolated cell-free DNA from the blood plasma from women that underwent tissue biopsy. We next assessed the methylation status of a large panel of oncology related genes to identify the tumor specific DNA among the normal DNA. With this, we aim to reduce the number of invasive tissue biopsies to women with the highest likelihood of having breast cancer. Namely, from the 206 women we recruited, 43% had no malignant disease and were thus over diagnosed. This highlights the urgent need for more precise and less intrusive diagnostic tools.
Peripheral blood samples were analyzed before and shortly after breast cancer surgery from 52 breast cancer patients. Preoperatively, CTCs were detected less frequently than post-surgery (5.8 vs 13.5%). The clinical follow-up data of the patients examined in this study are currently being evaluated and will then be compared with the results of the CTC analyses.
Molecular CTC characterization: To confirm the malignant identity of CTCs detected in the above studies, we obtained 26 cells and performed whole-genome sequencing. Of the 19 cells extracted from the prostatic plexus, 10 were identified as tumor cells and 9 as normal epithelial cells; of the 7 cells from the peripheral blood, 4 were identified as tumor cells and 3 as normal epithelial cells.
Functional studies: From the CTCs of a breast cancer patient, we established a permanent cell line that we used to study tumor cell extravasation. With it, we showed that the drug bortezomib is able to impair spontaneous lung metastasis in mice, suggesting that this drug can inhibit the formation of distant metastasis. As an alternative to mouse experiments, we started to use zebrafish embryos for our investigations. We could demonstrate that as few as 50–200 injected CTCs faithfully extravasate and proliferate in zebrafish embryos highlighting its value for functional and drug-target discovery.
CTC research has long been constrained by reliance on single‐marker enrichment and peripheral blood sampling, leaving important subpopulations undetected and the impact of diagnostic or surgical intervention on tumor‐cell dissemination largely unexplored. Our integrated body of work surmounts these limitations on three fronts: (1) we pioneered direct sampling of the tumor‐draining prostatic venous plexus during radical prostatectomy, revealing for the first time that surgery mobilizes hundreds of epithelial cells, including genetically confirmed malignant clones, into local circulation without a simple spillover into the peripheral blood, as verified by single‐cell whole‐genome sequencing. (2) we extended these mechanistic insights to breast cancer by showing that routine core‐needle or fine‐needle biopsies provoke a statistically significant surge in peripheral CTC counts, exceeding sampling variability, and identified loss of E-cadherin expression as a critical risk factor for biopsy‐induced tumor‐cell release. Our work transcends the current state of the art by integrating dual‐mode CTC enrichment, real‐time surgical and diagnostic sampling, and high‐resolution genomic profiling to illuminate how clinical procedures themselves influence tumor‐cell dissemination, thereby laying the foundation for perioperative monitoring. Although our study does not argue against the safety or necessity of biopsies, it highlights a possible mechanism by which physical disruption can mobilize cancer cells and points to liquid biopsy as a tool for identifying patients who may benefit from closer follow-up or treatments aimed at blocking released tumor cells from seeding new metastases. Our future work will include the development of anti‐extravasation therapies to prevent metastasis from happening because of surgery induced tissue damage.