Improved understanding and management of COVID-19, a potentially life-threatening disease, could greatly reduce the threat posed by its etiologic agent, SARS-CoV-2. Toward this end, we have identified a core peripheral blood immune signature across 63 hospital-treated patients with COVID-19 who were otherwise highly heterogeneous. The signature includes discrete changes in B and myelomonocytic cell composition, profoundly altered T cell phenotypes, selective cytokine/chemokine upregulation and SARS-CoV-2-specific antibodies. Some signature traits identify links with other settings of immunoprotection and immunopathology; others, including basophil and plasmacytoid dendritic cell depletion, correlate strongly with disease severity; while a third set of traits, including a triad of IP-10, interleukin-10 and interleukin-6, anticipate subsequent clinical progression. Hence, contingent upon independent validation in other COVID-19 cohorts, individual traits within this signature may collectively and individually guide treatment options; offer insights into COVID-19 pathogenesis; and aid early, risk-based patient stratification that is particularly beneficial in phasic diseases such as COVID-19.

Metastasis, the cardinal feature of malignant tumors, is an important clinical variable in patient prognosis. To understand the basis for metastasis, we systematically selected for highly invasive cells from breast cancer cell lines, MCF7 and MDA-MB-453, with moderate to low invasive ability using Boyden chamber invasion assay. The four-cycle selected invasive lines, named MCF7-I4 and MDA-MB-453-I4, respectively, displayed epithelial-mesenchymal transition (EMT) and dramatically enhanced invasive ability. EMT changes were corroborated with decreased level of E-cadherin and increased vimentin, fibronectin, and B 1 integrin. Twist, a basic helix-loop-helix transcription factor, and AKT2, a known proto-oncogene, were found to be elevated in the invasive cells compared with the parental. Ectopic expression and knockdown of Twist by short interference RNA resulted in significant increase and reduction, respectively, of AKT2 protein and mRNA expression. Twist bound to E-box elements on AKT2 promoter and enhanced its transcriptional activity. Moreover, silencing AKT2 decreased Twist-promoted migration, invasion, and paclitaxel resistance. Reintroducing AKT2 largely rescued the phenotype resulted from knockdown of Twist in I4 cells, suggesting that AKT2 is a downstream target and functional mediator of Twist. Finally, we observed a 68.8% correlation of elevated Twist and AKT2 expression in latestage breast cancers as oppose to 13% in early-stage breast cancers. Our study identifies Twist as a positive transcriptional regulator of AKT2 expression, and Twist-AKT2 signaling is involved in promoting invasive ability and survival of breast cancer cells.

No abstract

Summary Ancient organisms have a combined coagulation and immune system, and although links between inflammation and hemostasis exist in mammals, they are indirect and slower to act. Here we investigated direct links between mammalian immune and coagulation systems by examining cytokine proproteins for potential thrombin protease consensus sites. We found that interleukin (IL)-1α is directly activated by thrombin. Thrombin cleaved pro-IL-1α at a site perfectly conserved across disparate species, indicating functional importance. Surface pro-IL-1α on macrophages and activated platelets was cleaved and activated by thrombin, while tissue factor, a potent thrombin activator, colocalized with pro-IL-1α in the epidermis. Mice bearing a mutation in the IL-1α thrombin cleavage site (R114Q) exhibited defects in efficient wound healing and rapid thrombopoiesis after acute platelet loss. Thrombin-cleaved IL-1α was detected in humans during sepsis, pointing to the relevance of this pathway for normal physiology and the pathogenesis of inflammatory and thrombotic diseases.

No abstract

Purpose: Ectopic expression of GRM1 in murine melanocytes results in transformation into a form of melanoma, and more than 60% of human melanoma samples tested ectopically express GRM1. Stimulation of this receptor in vitro results in up-regulation of activated extracellular signal-regulated kinase (ERK). Furthermore, a xenograft model of melanoma treated with riluzole, an oral GRM1 blocking agent, showed decreased tumor growth compared with the untreated controls. We have now completed a phase 0 trial of riluzole in patients with melanoma. Experimental Design: Patients enrolled on this trial underwent a pretreatment biopsy, took 200 mg of oral riluzole per day for 14 days, and then underwent resection of their remaining tumor. We compared the levels of pERK and pAKT in the pretreatment and post-treatment samples and assessed the metabolic activity of pretreatment and posttreatment tumors using fluorodeoxyglucose positron emission tomography (FDG-PET) scanning. Results: We accrued 12 patients and all expressed GRM1. We found a significant decrease in pAKT and/or pERK in post-treatment tumor samples as compared with pretreatment samples in 4 (34%) patients. These four patients had a significant decrease in FDG-PET intensity post-treatment as well. Two other patients had a clinical response with no corresponding metabolic response; five patients had similar pretreatment and post-treatment FDG-PET scan findings; and one patient had progressive disease. Conclusions: Our data show that glutamate blockade with riluzole can inhibit signaling through the mitogen-activated protein kinase and phosphatidylinositol 3-kinase/AKT pathways and suppress the metabolic activity of melanoma. The ectopic expression of metabotropic glutamate receptors may be important in the pathogenesis of human melanoma, and targeting this pathway may be an effective therapy. Recently, our group described a heretofore unknown component of melanoma pathogenesis. A transgenic murine model of melanoma was developed by the ectopic expression of metabotropic glutamate receptor 1 (GRM1) in melanocytes (1-3). These mice spontaneously develop melanocytic lesions indistinguishable from human melanoma. We have expanded these original studies and have now shown that more than 60% of human melanomas express GRM1 and that activation of this receptor results in activation of the mitogen-activated protein kinase (MAPK) pathway in a B-Raf-and N-Rasindependent fashion (1). In preclinical studies, we have shown that the ectopic expression of GRM1 in melanocytes is transforming and that inhibition of GRM1 signaling in vitro and in vivo results in cell cycle arrest and subsequent apoptosis in human melanoma (2).We have now translated our findings into the clinic and have completed a phase 0 trial of riluzole in patients with stage III and IV melanoma. Riluzole (2-amino-6-trifluoromethoxybenzothiazole) is a noncompetitive GRM1 receptor antagonist that has been shown to be safe and effective in patients with amylotropic lateral sclerosis (ALS; refs. 4-7). Rilu...

Transforming growth factor (TGF)-b1 has a biphasic effect on rat intestinal epithelial (RIE) cells. By itself, TGF-b1 functions as a tumor suppressor by inhibiting the growth, migration and invasion of RIE cells. We show in this study that in conjunction with epidermal growth factor (EGF), TGF-b1 helped to augment migration, invasion and anchorage-independent growth (AIG) compared to that by EGF alone. EGF plus TGF-b1 induced a dramatic morphological change characteristic of epithelial-mesenchymal transition (EMT). The mechanism for this enhanced effect of TGF-b1 and EGF on oncogenic properties was explored by analysis of EGF-and TGF-b1-mediated signaling pathways and complementary DNA arrays. TGF-b1 augmented EGF-mediated signaling of mitogen-activated protein kinase (MAPK) and AKT by enhancing and prolonging the activation of the former and prolonging the activation of the latter. Inhibition of MAPK, but not phosphoinositide-3 kinase (PI3K), abolished TGF-b1 plus EGF-induced EMT and downregulation of E-cadherin at mRNA and protein levels. By contrast, cell migration and invasion were sensitive to inhibition of either MAPK or PI3 kinase. TGF-b1 plus EGF-induced AIG was significantly more resistant to inhibition of PI3K and MAPK compared to that induced by EGF alone. EGF and TGF-b1 synergistically induced the expression of a series of proteases including matrix metalloproteinase (MMP) 1 (collagenase), MMP3, MMP9, MMP10, MMP14 and cathepsin. Among them, the expression of MMP1, MMP3, MMP9 and MMP10 was MAPK dependent. Inhibition of the MMPs or cathepsin significantly blocked EGF plus TGF-b1-induced invasion, but had no effect on colony formation.Phospholipase C (PLC) and Cox2 induced by EGF plus TGF-b1 also played a significant role in invasion, whereas PLC was also important for colony formation. Our study reveals specific signaling functions and induction of genes differentially required for enhanced effect of EGF-and TGF-b1-induced oncogenic properties, and helps to explain the tumor-promoting effect of TGF-b1 in human cancer with elevated expression or activation of TGF-b1 and receptor protein tyrosine kinases.

No abstract