Genome-wide association studies of complex physiological traits and diseases consistently found that associated genetic factors, such as allelic polymorphisms or DNA mutations, only explained a minority of the expected heritable fraction. This discrepancy is known as “missing heritability”, and its underlying factors and molecular mechanisms are not established. Epigenetic programs may account for a significant fraction of the “missing heritability.” Epigenetic modifications, such as DNA methylation and chromatin assembly states, reflect the high plasticity of the genome and contribute to stably alter gene expression without modifying genomic DNA sequences. Consistent components of complex traits, such as those linked to human stature/height, fertility, and food metabolism or to hereditary defects, have been shown to respond to environmental or nutritional condition and to be epigenetically inherited. The knowledge acquired from epigenetic genome reprogramming during development, stem cell differentiation/de-differentiation, and model organisms is today shedding light on the mechanisms of (a) mitotic inheritance of epigenetic traits from cell to cell, (b) meiotic epigenetic inheritance from generation to generation, and (c) true transgenerational inheritance. Such mechanisms have been shown to include incomplete erasure of DNA methylation, parental effects, transmission of distinct RNA types (mRNA, non-coding RNA, miRNA, siRNA, piRNA), and persistence of subsets of histone marks.

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Trop-2 is a calcium signal transducer that drives tumor growth. Anti-Trop-2 antibodies with selective reactivity versus Trop-2 maturation stages allowed to identify two different pools of Trop-2, one localized in the cell membrane and one in the cytoplasm. Of note, membrane-localized/functional Trop-2 was found to be differentially associated with determinants of tumor aggressiveness and distinct breast cancer subgroups. These findings candidated Trop-2 states to having an impact on cancer progression. We tested this model in breast cancer. A large, consecutive human breast cancer case series (702 cases; 8 years median follow-up) was analyzed by immunohistochemistry with anti-Trop-2 antibodies with selective reactivity for cytoplasmic-retained versus functional, membrane-associated Trop-2. We show that membrane localization of Trop-2 is an unfavorable prognostic factor for overall survival (1+ versus 0 for all deaths: hazard ratio, 1.63; P = 0.04), whereas intracellular Trop-2 has a favorable impact on prognosis, with an adjusted hazard ratio for all deaths of 0.48 (high versus low; P = 0.003). A corresponding impact of intracellular Trop-2 was found on disease relapse (high versus low: hazard ratio, 0.51; P = 0.004). Altogether, we demonstrate that the Trop-2 activation states are critical determinants of tumor progression and are powerful indicators of breast cancer patients survival.

We recently reported that activation of Trop-2 through its cleavage at R87-T88 by ADAM10 underlies Trop-2–driven progression of colon cancer. However, the mechanism of action and pathological impact of Trop-2 in metastatic diffusion remain unexplored. Through searches for molecular determinants of cancer metastasis, we identified TROP2 as unique in its up-regulation across independent colon cancer metastasis models. Overexpression of wild-type Trop-2 in KM12SM human colon cancer cells increased liver metastasis rates in vivo in immunosuppressed mice. Metastatic growth was further enhanced by a tail-less, activated ΔcytoTrop-2 mutant, indicating the Trop-2 tail as a pivotal inhibitory signaling element. In primary tumors and metastases, transcriptome analysis showed no down-regulation of CDH1 by transcription factors for epithelial-to-mesenchymal transition, thus suggesting that the pro-metastatic activity of Trop-2 is through alternative mechanisms. Trop-2 can tightly interact with ADAM10. Here, Trop-2 bound E-cadherin and stimulated ADAM10-mediated proteolytic cleavage of E-cadherin intracellular domain. This induced detachment of E-cadherin from β-actin, and loss of cell-cell adhesion, acquisition of invasive capability, and membrane-driven activation of β-catenin signaling, which were further enhanced by the ΔcytoTrop-2 mutant. This Trop-2/E-cadherin/β-catenin program led to anti-apoptotic signaling, increased cell migration, and enhanced cancer-cell survival. In patients with colon cancer, activation of this Trop-2–centered program led to significantly reduced relapse-free and overall survival, indicating a major impact on progression to metastatic disease. Recently, the anti-Trop-2 mAb Sacituzumab govitecan-hziy was shown to be active against metastatic breast cancer. Our findings define the key relevance of Trop-2 as a target in metastatic colon cancer.

Trop-2 is a transmembrane signal transducer that is overexpressed in most human cancers, and drives malignant progression. To gain knowledge on the higher-order molecular mechanisms that drive Trop-2 signaling, we applied next-generation sequencing, proteomics, and high-resolution microscopy to models and primary cases of human colorectal cancer (CRC). We had previously shown that Trop-2 induces a Ca 2+ signal. We reveal here that Trop-2 binds the cell membrane Na + /K + -ATPase, and that clustering of Trop-2 induces an intracellular Ca 2+ rise followed by membrane translocation of PKCα, which in turn phosphorylates the Trop-2 cytoplasmic tail. This feed-forward signaling is promoted by the binding of Trop-2 to the PKCα membrane-anchor CD9. CRISPRbased inactivation of CD9 in CRC cells shows that CD9 is required by Trop-2 for recruiting PKCα and cofilin-1 to the cell membrane. This induces malignant progression through proteolytic cleavage of E-cadherin, remodeling of the β-actin cytoskeleton, and activation of Akt and ERK. The interaction between Trop-2 and CD9 was validated in vivo in murine models of CRC growth and invasion. Overexpression of the components of this Trop-2-driven super-complex significantly worsened disease-free and overall survival of CRC patients, supporting a pivotal relevance in CRC malignant progression. Our findings demonstrate a previously unsuspected layer of cancer growth regulation, which is dormant in normal tissues, and is activated by Trop-2 in cancer cells.