Biology subjects

Condeelis, J. S.

Publications and source records attributed to Condeelis, J. S..

10 recordsLinked to original sources

bioRxiv2026

Tie2 signaling in the tumor microenvironment orchestrates breast cancer cell dissemination through TMEM doorways

During breast cancer metastasis, tumor cells migrate toward intratumoral blood vessels and intravasate through stable structures known as TMEM (Tumor Microenvironment of Metastasis) doorways. TMEM doorways, composed of a Mena-expressing tumor cell, a Tie2hi/VEGFhi macrophage, and an endothelial cell, are clinically validated prognostic markers of distant metastasis in breast cancer and represent the exclusive sites of tumor cell intravasation. We previously demonstrated that Tie2 signaling is essential for TMEM doorway function and tumor cell intravasation. In this study, we investigated how Tie2 signaling promotes tumor cell intravasation and metastasis. Because all three TMEM doorway-associated cell types can express Tie2, we sought to determine which of these cells contribute to the Tie2 signaling-dependent vascular opening at TMEM doorways and tumor cell dissemination. We found that endothelial cells associated with TMEM doorways secrete Ang2, which stimulates VEGF-A expression in Tie2hi macrophages. Elevated VEGF-A levels at TMEM doorways increase vascular permeability, facilitating tumor cell entry into the bloodstream. Using tissue staining and line-scan analysis of Tie2 and lineage markers in human and mouse breast cancer models, we observed Tie2 expression in macrophages, tumor cells, and endothelial cells. To assess functional contributions, we selectively disrupted Tie2 in macrophages, endothelial cells, and cancer cells using CRISPR-Cas9 and RNAi approaches and tested in which of these cell-knockouts of Tie2 expression affected transendothelial migration in vitro. Macrophage-specific Tie2 deletion had the greatest impact on tumor cell intravasation. To confirm this finding in vivo, we generated a mouse model with inducible, macrophage-specific Tie2 knockout. Acute, targeted loss of Tie2 specifically in macrophages significantly reduced TMEM doorway associated vascular opening and tumor cell intravasation. Together, these findings establish macrophage Tie2 signaling as a critical driver of TMEM doorway-mediated vascular permeability and metastatic dissemination in breast cancer.

cancer biology↗

bioRxiv2026

Metastatic dissemination of breast cancer stem cells requires MenaINV for lung extravasation but not survival

Cancer stemness is a pivotal driver of tumor initiation, treatment resistance, and tumor cell survival. Cancer stem cells (CSCs), though constituting only a small fraction of primary tumor cells, are progressively enriched during metastatic progression: from circulating tumor cells traveling in the bloodstream, to disseminated tumor cells lodged in the lung vasculature, to extravasated tumor cells that have entered tissue parenchyma. However, whether CSCs have an intrinsic advantage for extravasation over cancer non-stem cells (CnSCs), or simply their increased representation in circulation renders them more likely to extravasate, remains unresolved. MenaINV, an invasive isoform of the actin regulatory protein Mena, promotes tumor cell transendothelial migration in primary and secondary sites, yet the direct mechanistic link between stemness and MenaINV in lung metastasis remains unresolved. Here, using a validated fluorescent stemness reporter (SORE6) to identify CSCs, we found that CSCs display elevated MenaINV expression relative to CnSCs. High-resolution intravital imaging showed that CSCs extravasate efficiently into lung parenchyma and survive at higher levels, robustly forming metastatic lesions, while CnSCs show limited extravasation, low survival, and poor colonization. Mechanistically, MenaINV disruption in CSCs specifically impaired extravasation without affecting survival, demonstrating that MenaINV is the key extravasation effector downstream of stemness, whereas stemness-associated factors independently confer survival advantages. Moreover, reintroduction of MenaINV in CnSCs restores their extravasation ability upon which extravasated CnSCs reactivate stem program and form metastases. Overall, we discovered a hierarchical framework where stemness regulates both survival and extravasation capacity, with MenaINV as the key CSC extravasation effector. Significance: This study reveals how breast cancer stem cells achieve metastatic dominance through separable pathways: MenaINV-dependent extravasation and MenaINV-independent survival, providing rationale for targeting stem program to improve patient outcome.

cancer biology↗

bioRxiv2025

Paclitaxel causes de novo induction of invasive breast cancer cells by repolarizing tumor-associated macrophages

Metastasis is the leading cause of breast cancer death, and tumor cells must migrate and invade to metastasize. Breast cancer cells that express the pro-metastatic actin regulatory protein MenaINV have an enhanced ability to migrate and intravasate within the primary tumor and extravasate at secondary sites. Though chemotherapy improves patient survival, treatment with paclitaxel leads to upregulation of MenaINV and an increase in metastasis in mice. MenaINV expression can be induced in breast cancer cells through cooperative NF-{kappa}B/ Notch1 signaling with macrophages, which are often increased in tumors in response to chemotherapy. MenaINV-expressing cells are also resistant to paclitaxel, raising the question of whether paclitaxel increases MenaINV by de novo induction or by selectively killing non-MenaINV-expressing cells. We hypothesized that paclitaxel causes de novo MenaINV induction by increasing macrophage-tumor cell NF-{kappa}B/ Notch1 signaling. Understanding this pro-metastatic effect of chemotherapy is crucial to refining treatment strategies. In this study, we demonstrate that paclitaxel increases MenaINV expression by de novo induction. Mechanistically, paclitaxel induces MenaINV by repolarizing tumor-associated macrophages towards a pro-inflammatory phenotype. These pro-inflammatory macrophages then participate in enhanced NF-{kappa}B/ Notch1 signaling with tumor cells, which leads to MenaINV induction in the tumor cells. These results lay the groundwork for novel microenvironment-based therapies to alleviate the pro-metastatic effects of chemotherapy in breast cancer.

cancer biology↗

bioRxiv2024

Intratumoral CXCL12 Gradients Contextualize Tumor Cell Invasion, Migration and Immune Suppression in Breast Cancer

Although the CXCL12/CXCR4 pathway has been prior investigated for its prometastatic and immuno- suppressive roles in the tumor microenvironment, evidence on the spatiotemporal regulation of these hallmarks has been lacking. Here, we demonstrate that CXCL12 forms a gradient specifically around cancer cell intravasation doorways, also known as Tumor Microenvironment of Metastasis (TMEM) doorways, thus facilitating the chemotactic translocation of prometastatic tumor cells expressing CXCR4 toward the perivascular TMEM doorways for subsequent entry into peripheral circulation. Fur- thermore, we demonstrate that the CXCL12-rich micro-environment around TMEM doorways may cre- ate immunosuppressive niches, whereby CD8+ T cells, despite being attracted to these regions, often exhibit reduced effector functions, limiting their efficacy. While the CXCL12/CXCR4 pathway can mini- mally influence the overall composition of immune cell populations, it biases the distribution of CD8+ T cells away from TMEM doorways, justifying its prior-established role as immunosuppressive factor for CD8+ T cells. Our research suggests that the complex interactions between CXCL12 and the various tumor and immune cell types contributes not only to the completion of the initial steps of the metastatic cascade, but also offers an immunological "sanctuary" to prometastatic tumor cells homed around TMEM doorways. Overall, our study enhances our current understanding on the mechanisms, via which CXCL12 orchestrates tumor cell behavior and immune dynamics, potentially guiding future thera- peutic strategies to combat breast cancer metastasis and improve anti-tumor immunity.

cancer biology↗

bioRxiv2024

Signaling events at TMEM doorways provide potential targets for inhibiting breast cancer dissemination

Tumor cell intravasation is essential for metastatic dissemination, but its exact mechanism is incompletely understood. We have previously shown that in breast cancer, the direct and stable association of a tumor cell expressing Mena, a Tie2hi/VEGFhi macrophage, and a vascular endothelial cell, creates an intravasation portal, called a "tumor microenvironment of metastasis" (TMEM) doorway, for tumor cell intravasation, leading to dissemination to distant sites. The density of TMEM doorways, also called TMEM doorway score, is a clinically validated prognostic marker of distant metastasis in breast cancer patients. Although we know that tumor cells utilize TMEM doorway-associated transient vascular openings to intravasate, the precise signaling mechanisms involved in TMEM doorway function are only partially understood. Using two mouse models of breast cancer and an in vitro assay of intravasation, we report that CSF-1 secreted by the TMEM doorway tumor cell stimulates local secretion of VEGF-A from the Tie2hi TMEM doorway macrophage, leading to the dissociation of endothelial junctions between TMEM doorway associated endothelial cells, supporting tumor cell intravasation. Acute blockade of CSF-1R signaling decreases macrophage VEGF-A secretion as well as TMEM doorway-associated vascular opening, tumor cell trans-endothelial migration, and dissemination. These new insights into signaling events regulating TMEM doorway function should be explored further as treatment strategies for metastatic disease.

cancer biology↗

bioRxiv2023

Macrophages Promote Tumor Cell Extravasation across an Endothelial Barrier through Thin Membranous Connections

Macrophages are important players involved in the progression of breast cancer, including in seeding the metastatic niche. However, the mechanism by which macrophages in the lung parenchyma interact with tumor cells in the vasculature to promote tumor cell extravasation at metastatic sites is not clear. To mimic macrophage-driven tumor cell extravasation, we used an in vitro assay (eTEM) in which an endothelial monolayer and a matrigel-coated filter separated tumor cells and macrophages from each other. The presence of macrophages promoted tumor cell extravasation while macrophage conditioned media was insufficient to stimulate tumor cell extravasation in vitro. This finding is consistent with a requirement for direct contact between macrophages and tumor cells. We observed the presence of Thin Membranous Connections (TMCs) resembling similar structures formed between macrophages and tumor cells called tunneling nanotubes which we previously demonstrated to be important in tumor cell invasion in vitro and in vivo (Hanna 2019). To determine if TMCs are important for tumor cell extravasation, we used macrophages with reduced levels of endogenous M-Sec (TNFAIP2), which causes a defect in tunneling nanotube formation. As predicted, these macrophages showed reduced macrophage-tumor cell TMCs. In both, human and murine breast cancer cell lines, there was also a concomitant reduction in tumor cell extravasation in vitro when co-cultured with M-Sec deficient macrophages compared to control macrophages. We also detected TMCs formed between macrophages and tumor cells through the endothelial layer in the eTEM assay. Furthermore, tumor cells were more frequently found in pores under the endothelium that contain macrophage protrusions. To determine the role of macrophage-tumor cell TMCs in vivo, we generated an M-Sec deficient mouse. Using an in vivo model of experimental metastasis, we detected a significant reduction in the number of metastatic lesions in M-Sec deficient mice compared to wild type mice. There was no difference in the size of the metastases, consistent with a defect specific to tumor cell extravasation and not metastatic outgrowth. Additionally, examination of time-lapse intravital-imaging (IVI) data sets of breast cancer cell extravasation in the lung, we could detect the presence of TMCs between extravascular macrophages and vascular tumor cells. Overall, our data indicate that macrophage TMCs play an important role in promoting the extravasation of circulating tumor cells in the lung.

cancer biology↗

bioRxiv2023

Cooperative NF-κB and Notch1 signaling promotes macrophage-mediated MenaINV expression in breast cancer

Metastasis is a multistep process that leads to the formation of clinically detectable tumor foci at distant organs and frequently patient demise. Only a subpopulation of breast cancer cells within the primary tumor can disseminate systemically and cause metastasis. To disseminate, cancer cells must express MenaINV, an isoform of the actin-regulatory protein Mena encoded by the ENAH gene that endows tumor cells with transendothelial migration activity allowing them to enter and exit the blood circulation. We have previously demonstrated that MenaINV mRNA and protein expression is induced in cancer cells by macrophage contact. In this study, we discovered the precise mechanism by which macrophages induce MenaINV expression in tumor cells. We examined the promoter of the human and mouse ENAH gene and discovered a conserved NF-{kappa}B transcription factor binding site. Using live imaging of an NF-{kappa}B activity reporter and staining of fixed tissues from mouse and human breast cancer we further determined that for maximal induction of MenaINV in cancer cell NF-{kappa}B needs to cooperate with the Notch1 signaling pathway. Mechanistically, Notch1 signaling does not directly increase MenaINV expression, but it enhances and sustains NF-{kappa}B signaling through retention of p65, an NF-{kappa}B transcription factor, in the nucleus of tumor cells, leading to increased MenaINV expression. In mice, these signals are augmented following chemotherapy treatment and abrogated upon macrophage depletion. Targeting Notch1 signaling in vivo decreased NF-{kappa}B signaling and MenaINV expression in the primary tumor and decreased metastasis. Altogether, these data uncover mechanistic targets for blocking MenaINV induction that should be explored clinically to decrease cancer cell dissemination and improve survival of patients with metastatic disease.

cancer biology↗

bioRxiv2021

Interleukin 4 controls the role of macrophages in pulmonary metastatic tumor cell seeding and growth

Metastasis is the systemic manifestation of cancer and the main cause of death from breast cancer. In mouse models of lung metastases, recruitment of classical monocytes from blood to the lung and their differentiation to metastasis-associated macrophages (MAMs) facilitate cancer cell extravasation, survival, and growth. Ablation of MAMs or their monocytic progenitors inhibits metastasis. We hypothesized that factors controlling macrophage polarization modulate tumor cell extravasation in the lung. We evaluated whether signaling by Th1 or Th2 cytokines in macrophages affected trans-endothelial migration of tumor cells in vitro. Interferon {gamma} and LPS inhibited macrophage-dependent tumor cell extravasation while the Th2 cytokine interleukin-4 (IL4) enhanced this process. We demonstrated that IL4 receptor (IL4r) null mice develop fewer and smaller lung metastasis. Adoptive transfer of wild type monocytes to IL4r deficient mice rescued this phenotype. IL4 signaling in macrophages controls the expression of the chemokine receptor CXCR2, necessary for IL4-mediated tumor cell extravasation in vitro. Furthermore, IL4 signaling in macrophages transcriptionally regulates several other genes already causally associated with lung metastasis including CCL2, CSF1, CCR1, HGF and FLT1. The central role for IL4 signaling in MAMs was confirmed by high-resolution intravital imaging of the lung in mice at the time of metastatic seeding, which showed reduced physical interaction between tumor cells and IL4r-deficient macrophages. This interaction enhances tumor cell survival. These data indicate that IL4 signaling in monocytes and macrophages is key during seeding and growth of breast metastasis in the lung as it regulates pro-tumoral paracrine signaling between cancer cells and macrophages.

cancer biology↗

bioRxiv2020

SUN-MKL1 crosstalk regulates nuclear deformation and fast motility of breast carcinoma cells in fibrillar ECM micro-environment

Aligned collagen fibers provide topography for the rapid migration of single tumor cells (streaming migration) to invade the surrounding stroma, move within tumor nests towards blood vessels to intravasate and form distant metastases. Mechanisms of tumor cell motility have been studied extensively in the 2D context, but the mechanistic understanding of rapid single tumor cell motility in the in vivo context is still lacking. Here, we show that streaming tumor cells in vivo use collagen fibers with diameters below 3 m. Employing 1D migration assays with matching in vivo fiber dimensions, we found a dependence of tumor cell motility on 1D substrate width, with cells moving the fastest and the most persistently on the narrowest 1D fibers (700 nm - 2.5 m). Interestingly, we also observed nuclear deformation in the absence of restricting extracellular matrix pores during high speed carcinoma cell migration in 1D, similar to the nuclear deformation observed in tumor cells in vivo. Further, we found that actomyosin machinery is aligned along the 1D axis and actomyosin contractility synchronously regulates cell motility and nuclear deformation. To further investigate the link between cell speed and nuclear deformation, we focused on the Linker of Nucleoskeleton and Cytoskeleton (LINC) complex proteins and SRF-MKL1 signaling, key regulators of mechanotransduction, actomyosin contractility and actin-based cell motility. Analysis of The Cancer Genome Atlas dataset showed a dramatic decrease in the LINC complex proteins SUN1 and SUN2 in primary tumor compared to the normal tissue. Disruption of LINC complex by SUN1+2 KD led to multi-lobular elongated nuclei, increased tumor cell motility and concomitant increase in F-actin, without affecting Lamin proteins. Mechanistically, we found that MKL1, an effector of changes in cellular G-actin to F-actin ratio, is required for increased 1D motility seen in SUN1+2 KD cells. Thus, we demonstrate a previously unrecognized crosstalk between SUN proteins and MKL1 transcription factor in modulating nuclear shape and carcinoma cell motility in an in vivo relevant 1D microenvironment.

cancer biology↗

bioRxiv2020

Live imaging of breast tumors shows macrophage-dependent induction and TMEM-mediated enrichment of cancer stem cells during metastatic dissemination

Cancer stem cells (CSCs) play an important role during metastasis, but the dynamic behavior and induction mechanisms of CSCs are not well understood. We employed high-resolution intravital microscopy using a CSC biosensor to directly observe CSCs in live mice with mammary tumors. CSCs display the slow-migratory, invadopod-rich phenotype that is the hallmark of disseminating tumor cells. CSCs are enriched near macrophages, particularly near macrophage-containing intravasation sites called Tumor Microenvironment of Metastasis (TMEM) doorways. A dramatic enrichment of CSCs occurs on association with TMEM doorways, contributing to the finding that CSCs represent [~]>60% of circulating tumor cells. Mechanistically, stemness is induced in non-stem cancer cells upon their direct contact with macrophages via Notch signaling. In breast cancers from patients, the density of TMEM doorways correlates strongly with the proportion of cancer cells expressing stem cell markers, indicating that in human breast cancer TMEM doorways are not only cancer cell intravasation portals but also CSC programming sites. One Sentence SummaryIntravital imaging reveals macrophage-mediated induction of cancer stem cells in vivo and their dramatic enrichment on dissemination through TMEM doorways.

cancer biology↗