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Simultaneous inhibition of interleukin-6 (IL-6) and interleukin-8 (IL-8) signaling diminishes cancer cell migration, and combination therapy has recently been shown to synergistically reduce metastatic burden in a preclinical model of triple-negative breast cancer. Here, we have engineered two novel bispecific antibodies that target the IL-6 and IL-8 receptors to concurrently block the signaling activity of both ligands. We demonstrate that a first-in-class bispecific antibody design has promising therapeutic potential, with enhanced selectivity and potency compared with monoclonal antibody and small-molecule drug combinations in both cellular and animal models of metastatic triple-negative breast cancer. Mechanistic characterization revealed that our engineered bispecific antibodies have no impact on cell viability, but profoundly reduce the migratory potential of cancer cells; hence they constitute a true anti-metastatic treatment. Moreover, we demonstrate that our antibodies can be readily combined with standard-of-care anti-proliferative drugs to develop effective anti-cancer regimens. Collectively, our work establishes an innovative metastasis-focused direction for cancer drug development.

The loss of E-cadherin (E-cad), an epithelial cell adhesion molecule, has been implicated in the epithelial-mesenchymal transition (EMT), promoting invasion and migration of cancer cells and, consequently, metastasis. However, recent studies have demonstrated that E-cad supports the survival and proliferation of metastatic cancer cells, suggesting that our understanding of E-cad in metastasis is far from comprehensive. Here, we report that E-cad upregulates thede novoserine synthesis pathway (SSP) in breast cancer cells. The SSP provides metabolic precursors for biosynthesis and resistance to oxidative stress, critically beneficial for E-cad-positive breast cancer cells to achieve faster tumor growth and more metastases. Inhibition of PHGDH, a rate- limiting enzyme in the SSP, significantly and specifically hampered the proliferation of E-cad- positive breast cancer cells and rendered them vulnerable to oxidative stress, inhibiting their metastatic potential. Our findings reveal that E-cad adhesion molecule significantly reprograms cellular metabolism, promoting tumor growth and metastasis of breast cancers.

Cell proliferation and invasion are two key drivers of tumor progression and are traditionally considered two independent cellular processes regulated by distinct pathways. Throughin vitroandin silicomethods, we provide evidence that these two processes are intrinsically coupled through matrix-adhesion friction. Using novel tumor spheroids, we show that both tumor cell proliferation and invasion are limited by a volumetric carrying capacity of the system, i.e. maximum spatial cell concentration supported by the system’s total cell count, nutrient consumption rate, and collagen gel mechanical properties. To manipulate these phenotypes in breast cancer cells, we modulate the expression of E-cadherin and its associated role in adhesion, invasion, and proliferation. We integrate these results into a mixed-constitutive formulation to computationally delineate the contributions of cellular and extracellular adhesion, stiffness, and mechanical properties of the extracellular matrix (ECM) to the proliferative and invasive fates of breast cancer tumor spheroids. Both approaches conclude that the dominant drivers of tumor fate are system properties modulating cell-ECM friction, such as E-cadherin dependent cell-ECM adhesion and matrix pore size.

The loss of the intercellular adhesion molecule E-cadherin is a hallmark of the epithelial-mesenchymal transition (EMT), during which tumor cells transition into an invasive phenotype. Thus, E-cadherin has long been considered a tumor suppressor gene. However, recent studies have provided evidence that E-cadherin may promote metastasis rather than suppress it, suggesting oncogenic behavior. Here we provide data that E-cadherin plays an oncogenic role in breast cancer by promoting a hyper-proliferative phenotype in breast cancer cells via interaction with EGFR. This interaction results in the activation of the MEK/ERK signaling pathway, leading directly to changes in proliferation via transcription factors such as c-Fos. Pharmacological inhibition of MEK in E-cadherin positive breast cancer cells significantly decreases both tumor growth and macro-metastasis in vivo. This work provides evidence for a novel role of E-cadherin in breast tumor progression and identifies a potential new target to treat hyper-proliferative E-cadherin-positive breast tumors. Citation Format: Gabriella C. Russo, Ashleigh Crawford, David J. Clark, Julie Cui, Ryan Carney, Michelle N. Karl, Boyang Su, Batrholomew Starich, Tung-Shing Lee, Qiming Zhang, Pei-hsun Wu, Meng-Horng Lee, Hon S. Leong, Vito Rebecca, Hui Zhang, Denis Wirtz. E-cadherin and EGFR interactions result in hyper-proliferation via ERK signaling in breast cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 1000.

The loss of the intercellular adhesion molecule E-cadherin is a hallmark of the epithelial-mesenchymal transition (EMT), which promotes a transition of cancer cells to a migratory and invasive phenotype. E-cadherin is associated with a decrease in cell proliferation in normal cells. Here, using physiologically relevant 3D in vitro models, we find that E-cadherin induces hyper-proliferation in breast cancer cells through activation of the Raf/MEK/ERK signaling pathway. These results were validated and consistent across multiple in vivo models of primary tumor growth and metastatic outgrowth. E-cadherin expression dramatically increases tumor growth and, without affecting the ability of cells to extravasate and colonize the lung, significantly increases macrometastasis formation via cell proliferation at the distant site. Pharmacological inhibition of MEK1/2, blocking phosphorylation of ERK in E-cadherin-expressing cells, significantly depresses both tumor growth and macrometastasis. This work suggests a novel role of E-cadherin in tumor progression and identifies a potential new target to treat hyper-proliferative breast tumors. SUMMARY E-cadherin, an extensively studied transmembrane molecule ubiquitously expressed in normal epithelial tissues, promotes and maintains intercellular adhesion. In cancer, the loss of adhesion molecule E-cadherin is associated with onset of invasion via epithelial-to-mesenchymal transition (EMT) process.1 EMT consists of a highly orchestrated cascade of molecular events where epithelial cells switch from a non-motile phenotype to an invasive, migratory phenotype accompanied by a change in cell morphology.1,2 These processes are believed to then trigger metastasis in carcinomas (cancers of epithelial origin). Moreover, the expression of intercellular adhesion molecule E-cadherin (E-cad) is associated with a decrease in cell proliferation in normal cells. Classical experiments in fibroblasts and epithelial cells show that the expression of E-cad not only promotes cell-cell adhesion, but also reduces cell proliferation and onset of apoptosis.3,4 Altogether these results have long supported that E-cad acts as a tumor suppressor gene.1,2 However, despite its role in cell-adhesion the requirement for loss of E-cad in metastasis has recently been re-assessed.5,6,7,8 These investigations focus on E-cad’s role in EMT, even though the relationship between E-cad and proliferation is just as intriguing. While E-cad has been shown to have anit-proliferative effects in normal cells, E-cad also helps maintain a pluripotent and proliferative phenotype in stem cells, and notably is lost during differentiation, a non-proliferative step of stem cell progression.9,10 Yet, despite potentially important implications in our understanding of tumor progression, whether E-cad expression affects growth in cancer cells remains mostly unexplored. Here, utilizing a physiologically relevant 3D in vitro model and multiple in vivo models, we studied the impact of E-cad on cell proliferation at the primary tumor site and proliferation at a secondary site. Remarkably, E-cad upregulates multiple proliferation pathways, including hyper-activation of the ERK cascade within the greater MAPKinase pathway, resulting in a dramatic increase in cell proliferation in vitro and tumor growth in vivo. When the phosphorylation of ERK is blocked utilizing a MEK1/2 inhibitor, PD032590111, this effect is reversed in vitro and in vivo. Thus, E-cad plays an oncogenic role in tumorigenesis and merits evaluation as a potential new drug target.