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Tumor cell death supports recurrence of cancer

Periodic Reporting for period 4 - Cancer-Recurrence (Tumor cell death supports recurrence of cancer)

Reporting period: 2019-04-01 to 2020-08-31

The most common treatment strategy for cancer is surgical removal combined with therapy to kill metastatic cells. For example, chemotherapy is often given prior to surgery to reduce tumor size in order to improve the success rate of tumor resection and after surgery to reduce potential metastasis. Most therapies aim at inducing cell death, mostly (but not exclusively) by stimulating the intrinsic apoptotic pathway. However, some cells survive the therapy by e.g. being resistant to chemotherapeutic. These cells survive the treatment and manifest later, leading to tumor recurrence. Therefore, it is of utmost importance to exactly understand the cellular and molecular mechanisms that drive the outgrowth of the few surviving cells.

State-of-the–art objectives and hypothesis:
Although all the universal aim of all the different therapies is to kill the tumor cells, the therapie itself can also influence the environment of the surviving tumor cells. This can potentially lead to cell states (e.g. EMT and stemness) that enable the surviving cells to spread and regrow (distant) tumors leading to recurrence. Therefore, we will study the unintended side-effects of therapies on the surviving tumor cells and stroma, identify the key cell types and mechanisms that mediate this effect, and test whether interference with these key cell types and mechanisms leads to reduced recurrence of tumors upon treatment. Since induction of cell death is the universal aim of therapy, interfering with unintended side-effects of tumor cell death may be a therapeutic avenue leading towards improved outcome of a wide range of therapies.

Hypothesis: Although therapies can kill the bulk of cancer cells, it potentially also has unintended and harmful side-effects on the remaining fraction of tumor cells and stroma, with subsequent profound consequences for the long-term outcome of cancer.

Main aim: Gain a better understanding on the unintended side-effects of therapies on the growth and dissemination of surviving tumor cells, with the ultimate goal to improve clinical strategies by reducing tumor recurrence.
Results:
Although biopsies have a prognostic benefit, potential negative effects of inducing cell death in the remaining tumor cells have also been suggested. Using a retrospect study of patients and intravital imaging study of mice, we have identified some of these negative aspects, including stimulation of proliferation and migration of non-resected cells, and provide a strategy to prevent these adverse effects. By repeated high-resolution intravital microscopy, we showed that biopsy-like injury induces migration and proliferation of tumor cells through chemokine (C-C motif) ligand 2 (CCL-2)-dependent recruitment of macrophages. Blocking macrophage recruitment or administrating dexamethasone, a commonly used glucocorticoid to treat inflammation, led to suppression of the observed inflammatory response and subsequent tumor growth upon biopsy both in mice and patients. Taken together, our study suggested that induction of cell death by needle biopsy induces an inflammation response with a subsequent enhancement of proliferation and migration in the non-resected cancer cells, and that inhibiting CCL-2-dependent recruitment of macrophages may further increase the clinical benefits from surgical and biopsy procedures.

Exploration:
For our research, we had to develop different new mouse models, intravital imaging tools, and single cell sequencing approaches to study whether surviving cancer cells undergo a phenotypic change (e.g. undergo epithelial-to-mesenchymal transition (EMT) or acquire stemness). We have used this tools to show the key roles of stem cell properties and EMT states to spread to distant sites and to induce (re)growth of tumors.

Dissemination:
The dissemination was multiple. We have disseminated our results to the scientific community by publishing >20 papers in high-impact journals such as Nature, Cell, and Cell Reports. Moreover, we disseminated our new tools and approaches by publishing protocol papers, and have shared reagents through e.g. Addgene (https://www.addgene.org/Jacco_van_Rheenen/). Moreover, our data was communicated by presentations at international meetings such as the AACR meetings, and to society by giving layman talks.
Cancer therapeutics target the “average” differentiated tumor cell, while tumors are actually very heterogeneous. Therefore many therapies fail to target “non-average” populations of cells with a genetic or epigenetic profile that makes them resistant to therapies. Although important knowledge on how these resistant cells regrow tumors has been obtained using traditional techniques including histochemistry, (q)PCR and Western blotting, these approaches provide snapshots of large populations of cells and therefore fail to provide crucial information about the history of individual cells (e.g. the few surviving cells upon therapy). Over the years, we have pioneered high resolution intravital imaging techniques and combined these with high-end genetic fluorescent mouse models in order to visualize the behavior of individual tumor cells in living mice. Using our state-of-the-art intravital imaging and fluorescent genetic tumor mouse models, we were able to study the plasticity of the behavior of individual cells during tissue development and homeostasis, tumor progression, and therapy responses. In this ERC project, we have used our unique imaging tools and fluorescent mouse models to study the potential non-intended side-effects of therapy on surrounding and distant surviving tumor cells and stromal cells. We have identified key cell types and mechanisms that mediate non-intended side-effects of therapies on surviving tumor cells. We have found that taking a biopsy leads to an inflammatory response with a subsequent recruitment of macrophages by CCL2. This recruitment induces migration and proliferation in the remaining tumor cells both in mice and in patients. When interfering with this recruitment of macrophages, by CCL2 inhibition, macrophages depletion, or anti-inflammation drug, we could block the biopsy-mediated induction of migration and proliferation of the remaining tumor cells. So in addition to identifying that therapies can have adverse effects on the remaining tumor cells, we also identified approaches to block this detrimental effects.