An epidermal growth factor motif of developmental endothelial locus 1 protein inhibits efficient angiogenesis in explanted squamous cell carcinoma in vivo

ABSTRACT Background: Cancer gene therapy using a nonviral vector is expected to be repeatable, safe, and inexpensive, and to have long-term effectiveness. Gene therapy using the E3 and C1 (E3C1) domain of developmental endothelial locus-1 (Del1) has been shown to improve prognosis in a mouse transplanted tumor model. Objective: In this study, we examined how this treatment affects angiogenesis in mouse transplanted tumors. Materials and methods: Mouse transplanted tumors (SCCKN human squamous carcinoma cell line) were injected locally with a nonviral plasmid vector encoding E3C1 weekly. Histochemical analysis of the transplanted tumors was then performed to assess the effects of E3C1 on prognosis. Results: All mice in the control group had died or reached an endpoint within 39 days. In contrast, one of ten mice in the E3C1 group had died by day 39, and eight of ten had died or reached an endpoint by day 120 (p < 0.01). Enhanced apoptosis in tumor stroma was seen on histochemical analyses, as was inhibited tumor angiogenesis in E3C1-treated mice. In addition, western blot analysis showed decreases in active Notch and HEY1 proteins. Conclusion: These findings indicate that cancer gene therapy using a nonviral vector encoding E3C1 significantly improved life-span by inhibiting tumor angiogenesis. (REV INVEST CLIN. 2021;73(1):39-51)

An Epidermal Growth Factor Motif of Developmental Endothelial Locus 1 Protein Inhibits Efficient Angiogenesis in Explanted Squamous Cell Carcinoma In Vivo.

BACKGROUND: Cancer gene therapy using a nonviral vector is expected to be repeatable, safe, and inexpensive, and to have longterm effectiveness. Gene therapy using the E3 and C1 (E3C1) domain of developmental endothelial locus-1 (Del1) has been shown to improve prognosis in a mouse transplanted tumor model. OBJECTIVE: In this study, we examined how this treatment affects angiogenesis in mouse transplanted tumors. MATERIALS AND METHODS: Mouse transplanted tumors (SCCKN human squamous carcinoma cell line) were injected locally with a nonviral plasmid vector encoding E3C1 weekly. Histochemical analysis of the transplanted tumors was then performed to assess the effects of E3C1 on prognosis. RESULTS: All mice in the control group had died or reached an endpoint within 39 days. In contrast, one of ten mice in the E3C1 group had died by day 39, and eight of ten had died or reached an endpoint by day 120 (p < 0.01). Enhanced apoptosis in tumor stroma was seen on histochemical analyses, as was inhibited tumor angiogenesis in E3C1-treated mice. In addition, western blot analysis showed decreases in active Notch and HEY1 proteins. CONCLUSION: These findings indicate that cancer gene therapy using a nonviral vector encoding E3C1 significantly improved life-span by inhibiting tumor angiogenesis.

Structural mapping of hot spots within human CASPR2 discoidin domain for autoantibody recognition.

Accumulating evidence has showed that anti-CASPR2 autoantibodies occur in a long list of neurological immune disorders including limbic encephalitis (LE). Belonging to the well-known neurexin superfamily, CASPR2 has been suggested to be a central node in the molecular networks controlling neurodevelopment. Distinct from other subfamilies in the neurexin superfamily, the CASPR subfamily features a unique discoidin (Disc) domain. As revealed by our and others' recent studies, CASPR2 Disc domain bears a major epitope for autoantibodies. However, structural information on CASPR2 recognition by autoantibodies has been lacking. Here, we report the crystal structure of human CASPR2 Disc domain at a high resolution of 1.31 Å, which is the first atomic-resolution structure of the CASPR subfamily members. The Disc domain adopts a total ß structure and folds into a distorted jellyroll-like barrel with a conserved disulfide-bond interlocking its N- and C-termini. Defined by four loops and located in one end of the barrel, the "loop-tip surface" is totally polar and easily available for protein docking. Based on structure-guided epitope prediction, we generated nine mutants and evaluated their binding to autoantibodies of cerebrospinal fluid from twelve patients with limbic encephalitis. The quadruple mutant G69N/A71S/S77N/D78R impaired CASPR2 binding to autoantibodies from eleven LE patients, which indicates that the loop L1 in the Disc domain bears hot spots for autoantibody interaction. Structural mapping of autoepitopes within human CASPR2 Disc domain sheds light on how autoantibodies could sequester CASPR2 ectodomain and antagonize its functionalities in the pathogenic processes.

The C. elegans Discoidin Domain Receptor DDR-2 Modulates the Met-like RTK-JNK Signaling Pathway in Axon Regeneration.

The ability of specific neurons to regenerate their axons after injury is governed by cell-intrinsic regeneration pathways. However, the signaling pathways that orchestrate axon regeneration are not well understood. In Caenorhabditis elegans, initiation of axon regeneration is positively regulated by SVH-2 Met-like growth factor receptor tyrosine kinase (RTK) signaling through the JNK MAPK pathway. Here we show that SVH-4/DDR-2, an RTK containing a discoidin domain that is activated by collagen, and EMB-9 collagen type IV regulate the regeneration of neurons following axon injury. The scaffold protein SHC-1 interacts with both DDR-2 and SVH-2. Furthermore, we demonstrate that overexpression of svh-2 and shc-1 suppresses the delay in axon regeneration observed in ddr-2 mutants, suggesting that DDR-2 functions upstream of SVH-2 and SHC-1. These results suggest that DDR-2 modulates the SVH-2-JNK pathway via SHC-1. We thus identify two different RTK signaling networks that play coordinated roles in the regulation of axonal regeneration.