Breast Cancer: Circular RNAs Mediating Efficacy in Preclinical In Vivo Models.

In order to identify new targets and treatment modalities for breast cancer, we searched the literature for circular RNAs (circRNAs) with efficacy in preclinical breast cancer-related in vivo models. From our search, we identified 26 up-regulated and six down-regulated circRNAs which mediate efficacy in breast cancer-related preclinical in vivo models. We discuss reconstitution and inhibition of the identified circRNAs, as well as druggability and validation of the targets identified in the context of chemoresistance, inhibition of proliferation and metastasis. Pathways driven by suppressors of cytokines and high-mobility group proteins, nuclear factor B and Hippo signaling emerged as important drivers of tumor growth and metastasis. The role of trefoil factor-1 with respect to metastasis of estrogen receptor-positive breast cancer also merits further investigation. In addition, mucin 19 has emerged as an unexplored target for treatment of breast cancer.

Endoglycan (PODXL2) is proteolytically processed by ADAM10 (a disintegrin and metalloprotease 10) and controls neurite branching in primary neurons.

Cell adhesion is tightly controlled in multicellular organisms, for example, through proteolytic ectodomain shedding of the adhesion-mediating cell surface transmembrane proteins. In the brain, shedding of cell adhesion proteins is required for nervous system development and function, but the shedding of only a few adhesion proteins has been studied in detail in the mammalian brain. One such adhesion protein is the transmembrane protein endoglycan (PODXL2), which belongs to the CD34-family of highly glycosylated sialomucins. Here, we demonstrate that endoglycan is broadly expressed in the developing mouse brains and is proteolytically shed in vitro in mouse neurons and in vivo in mouse brains. Endoglycan shedding in primary neurons was mediated by the transmembrane protease a disintegrin and metalloprotease 10 (ADAM10), but not by its homolog ADAM17. Functionally, endoglycan deficiency reduced the branching of neurites extending from primary neurons in vitro, whereas deletion of ADAM10 had the opposite effect and increased neurite branching. Taken together, our study discovers a function for endoglycan in neurite branching, establishes endoglycan as an ADAM10 substrate and suggests that ADAM10 cleavage of endoglycan may contribute to neurite branching.

The tetraspanin Tspan15 is an essential subunit of an ADAM10 scissor complex.

A disintegrin and metalloprotease 10 (ADAM10) is a transmembrane protein essential for embryonic development, and its dysregulation underlies disorders such as cancer, Alzheimer's disease, and inflammation. ADAM10 is a "molecular scissor" that proteolytically cleaves the extracellular region from >100 substrates, including Notch, amyloid precursor protein, cadherins, growth factors, and chemokines. ADAM10 has been recently proposed to function as six distinct scissors with different substrates, depending on its association with one of six regulatory tetraspanins, termed TspanC8s. However, it remains unclear to what degree ADAM10 function critically depends on a TspanC8 partner, and a lack of monoclonal antibodies specific for most TspanC8s has hindered investigation of this question. To address this knowledge gap, here we designed an immunogen to generate the first monoclonal antibodies targeting Tspan15, a model TspanC8. The immunogen was created in an ADAM10-knockout mouse cell line stably overexpressing human Tspan15, because we hypothesized that expression in this cell line would expose epitopes that are normally blocked by ADAM10. Following immunization of mice, this immunogen strategy generated four Tspan15 antibodies. Using these antibodies, we show that endogenous Tspan15 and ADAM10 co-localize on the cell surface, that ADAM10 is the principal Tspan15-interacting protein, that endogenous Tspan15 expression requires ADAM10 in cell lines and primary cells, and that a synthetic ADAM10/Tspan15 fusion protein is a functional scissor. Furthermore, two of the four antibodies impaired ADAM10/Tspan15 activity. These findings suggest that Tspan15 directly interacts with ADAM10 in a functional scissor complex.

Non-cell-autonomous function of DR6 in Schwann cell proliferation.

Death receptor 6 (DR6) is an orphan member of the TNF receptor superfamily and controls cell death and differentiation in a cell-autonomous manner in different cell types. Here, we report an additional non-cell-autonomous function for DR6 in the peripheral nervous system (PNS). DR6-knockout (DR6 KO) mice showed precocious myelination in the PNS Using an in vitro myelination assay, we demonstrate that neuronal DR6 acts in trans on Schwann cells (SCs) and reduces SC proliferation and myelination independently of its cytoplasmic death domain. Mechanistically, DR6 was found to be cleaved in neurons by "a disintegrin and metalloprotease 10" (ADAM10), releasing the soluble DR6 ectodomain (sDR6). Notably, in the in vitro myelination assay, sDR6 was sufficient to rescue the DR6 KO phenotype. Thus, in addition to the cell-autonomous receptor function of full-length DR6, the proteolytically released sDR6 can unexpectedly also act as a paracrine signaling factor in the PNS in a non-cell-autonomous manner during SC proliferation and myelination. This new mode of DR6 signaling will be relevant in future attempts to target DR6 in disease settings.

Ubiquitin E3 ligase Nedd4-1 acts as a downstream target of PI3K/PTEN-mTORC1 signaling to promote neurite growth.

Protein ubiquitination is a core regulatory determinant of neural development. Previous studies have indicated that the Nedd4-family E3 ubiquitin ligases Nedd4-1 and Nedd4-2 may ubiquitinate phosphatase and tensin homolog (PTEN) and thereby regulate axonal growth in neurons. Using conditional knockout mice, we show here that Nedd4-1 and Nedd4-2 are indeed required for axonal growth in murine central nervous system neurons. However, in contrast to previously published data, we demonstrate that PTEN is not a substrate of Nedd4-1 and Nedd4-2, and that aberrant PTEN ubiquitination is not involved in the impaired axon growth upon deletion of Nedd4-1 and Nedd4-2. Rather, PTEN limits Nedd4-1 protein levels by modulating the activity of mTORC1, a protein complex that controls protein synthesis and cell growth. Our data demonstrate that Nedd4-family E3 ligases promote axonal growth and branching in the developing mammalian brain, where PTEN is not a relevant substrate. Instead, PTEN controls neurite growth by regulating Nedd4-1 expression.