Deceiving SARS-CoV-2 molecular-tropism clues - A combinational contemporary strategy.

Several attempts to control the dreadfulness of SARS-CoV-2 are still underway. Based on the literature evidences we have speculated a prospective contemporary remedy, which was categorized into Specificity, Remedy, and a Conveyor. In which, pros and cons were discussed and inferred the possible alternatives. (a) Specificity: Implicit to express the ACE2 receptors in conveyor cells to deceive SARS-CoV-2 frompreponetargets. (b) Remedy: As depletion of pulmonary surfactants causes strong acute respiratory distress syndrome, we propose an entity of a cost-effective artificialsurfactantsystem as a remedy to pulmonary complications. (c) Conveyor: We propose red blood cells (RBCs) as a conveyor with embedded artificial surfactant and protruding ACE2 receptors for the target-specific delivery. Overall we postulate focused insights by employing a combinational contemporary strategy to steer towards a prospective direction on combating SARS-CoV-2.

Early Treatment of Coxsackievirus B3-Infected Animals With Soluble Coxsackievirus-Adenovirus Receptor Inhibits Development of Chronic Coxsackievirus B3 Cardiomyopathy.

BACKGROUND: Coxsackie-B-viruses (CVB) are frequent causes of acute myocarditis and dilated cardiomyopathy, but an effective antiviral therapy is still not available. Previously, we and others have demonstrated that treatment with an engineered sCAR-Fc (soluble coxsackievirus-adenovirus receptor fused to the carboxyl-terminus of human IgG) efficiently neutralizes CVB3 and inhibits the development of cardiac dysfunction in mice with acute CVB3-induced myocarditis. In this study, we analyzed the potential of sCAR-Fc for treatment of chronic CVB3-induced myocarditis in an outbred NMRI mouse model. METHODS: NMRI mice were infected with the CVB3 strain 31-1-93 and treated with a sCAR-Fc expressing adeno-associated virus 9 vector 1, 3, and 7 days after CVB3 infection. Chronic myocarditis was analyzed on day 28 after infection. RESULTS: Initial investigations showed that NMRI mice develop pronounced chronic myocarditis between day 18 and day 28 after infection with the CVB3 strain 31-1-93. Chronic cardiac infection was characterized by inflammation and fibrosis as well as persistence of viral genomes in the heart tissue and by cardiac dysfunction. Treatment of NMRI mice resulted in a distinct reduction of cardiac inflammation and fibrosis and almost complete elimination of virus RNA from the heart by day 28 after infection. Moreover, hemodynamic measurement revealed improved cardiac contractility and diastolic relaxation in treated mice compared with mice treated with a control vector (mean±SD; maximal pressure, 81.9±9.2 versus 69.4±8.6 mm Hg, P=0.02; left ventricular ejection fraction, 68.9±8.5 versus 54.2±11.5%, P=0.02; dP/dtmax, 7275.2±1674 versus 4432.6±1107 mm Hg/s, P=0.004; dP/dtmin, -4046.9±776 versus -3146.3±642 mm Hg/s, P=0.046). The therapeutic potential of sCAR-Fc is limited, however, since postponed start of sCAR-Fc treatment either 3 or 7 days after infection could not attenuate myocardial injury. CONCLUSIONS: Early therapeutic employment of sCAR-Fc, initiated at the beginning of the primary viremia, inhibits the development of chronic CVB3-induced myocarditis and improves the cardiac function to a level equivalent to that of uninfected animals.

TIGIT signalling pathway negatively regulates CD4+ T-cell responses in systemic lupus erythematosus.

B-lymphocyte hyperactivity in systemic lupus erythematosus (SLE) is T-cell-dependent, and CD4+ T-cell activation is essential to SLE pathogenesis. However, the mechanism of the deregulation of CD4+ T cells in SLE is largely unknown. T-cell immunoglobulin and ITIM domain (TIGIT) is a new inhibitory receptor preferentially expressed on activated CD4+ T cells. Here, we address the role of TIGIT in the pathogenesis of SLE. Our results showed that TIGIT expression on CD4+ T cells was significantly elevated in patients with SLE and highly correlated with the activity of the disease. TIGIT+ CD4+ T cells from both healthy individuals and patients with SLE had a more activated phenotype than TIGIT- CD4+ T cells. In contrast, the activation, proliferation and cytokine production potential of TIGIT+ CD4+ T cells were significantly lower than those of TIGIT- CD4+ T cells. Furthermore, activation of the TIGIT pathway by using CD155 could substantially down-regulate the activities of CD4+ T cells from SLE patients in vitro, and in vivo administration of CD155 resulted in a delayed development of SLE in MRL/lpr mice. TIGIT is a powerful negative regulator of CD4+ T cells in SLE, which suggests that the TIGIT signalling pathway may be used as a potential therapeutic target for treating this disease.

Enhancement of SIV-specific cell mediated immune responses by co-administration of soluble PD-1 and Tim-3 as molecular adjuvants in mice.

The development of an effective T cell based HIV vaccine would need to elicit cell mediated immune responses with superior magnitude, breadth, and quality. Since blocking the interactions between inhibitory receptors with their associated ligands using soluble PD-1 (sPD-1) and soluble Tim-3 (sTim-3) have been shown to reverse T cell exhaustion and enhance cell mediated immune responses, we tested if co-administration of sPD-1 and sTim-3 with an adenovirus vectored SIV vaccine (rAd5-SIV) can enhance cell mediated immune responses. The frequency of SIV antigen specific IFN-γ spot-forming cells and the secretion of IFN-γ and TNF-α by splenocytes from rAd5-SIV immunized mice was significantly increased when stimulated ex vivo with SIV peptides in the presence of sPD-1 or sTim-3 or both sPD-1 and sTim-3. The magnitude of cell mediated immune responses elicited by rAd5-SIV was enhanced by co-administration of sPD-1 and sTim-3. Co-administration of both sPD-1 and sTim-3 induced higher frequency of SIV antigen specific IFN-γ(+) spot-forming cells to poorly immunogenic Vif and Tat. The percentage of cell mediated responses for each SIV antigen became more balanced, with reduction to Gag but induction to non-structural proteins. Furthermore, co-injection of rAd5-sPD1 and rAd5-sTim3 with rAd5-SIV in mice enhanced T cell proliferation capability and generated more antigen specific IFN-γ(+) CD4(+) and CD8(+) T cells. Our study provided a new approach to enhance vaccine induced cell mediated immune responses, which may be applicable to improve the efficacy of vaccines against SIV/HIV.

Polioviral receptor binding ligand: a novel and safe peptide drug carrier from polioviral capsid.

Cellular uptake enhancement of green fluorescent protein (GFP) into human colon adenocarcinoma (HT-29) and human mouth epidermal carcinoma (KB) cells by a segment of VP1-BC loop polioviral capsid (V), a polioviral receptor binding peptide and HIV-I transactivator of transcription (Tat) was evaluated. HT-29 and KB cells were incubated with various molar concentrations of GFP, V, and Tat mixtures. Both V and Tat showed potent enhancement of GFP uptake into HT-29 and KB cells. In HT-29 cells, the V-GFP, Tat-GFP, and V-Tat-GFP mixtures enhanced the GFP cellular uptake efficiency with the maximum of 3.98-, 4.59-, and 4.08-folds of GFP at 1:3, 1:1/6, and 1/6:1:1/6 molar ratios, respectively. For KB cells, the V-GFP, Tat-GFP, and V-Tat-GFP mixtures enhanced the GFP cellular uptake efficiency with the maximum of 4.05-, 5.09-, and 4.91-folds of GFP at 1:1/6, 1:1, and 1:1:1 molar ratios, respectively. Both V and V-GFP mixtures showed a lower cytotoxicity effect than Tat and Tat-GFP mixture. These studies demonstrated the potential of polioviral capsid, a polioviral receptor binding peptide as a novel, low cytotoxicity carrier for the development of peptide drugs delivery system.

Prevention of cardiac dysfunction in acute coxsackievirus B3 cardiomyopathy by inducible expression of a soluble coxsackievirus-adenovirus receptor.

BACKGROUND: Group B coxsackieviruses (CVBs) are the prototypical agents of acute myocarditis and chronic dilated cardiomyopathy, but an effective targeted therapy is still not available. Here, we analyze the therapeutic potential of a soluble (s) virus receptor molecule against CVB3 myocarditis using a gene therapy approach. METHODS AND RESULTS: We generated an inducible adenoviral vector (AdG12) for strict drug-dependent delivery of sCAR-Fc, a fusion protein composed of the coxsackievirus-adenovirus receptor (CAR) extracellular domains and the carboxyl terminus of human IgG1-Fc. Decoy receptor expression was strictly doxycycline dependent, with no expression in the absence of an inducer. CVB3 infection of HeLa cells was efficiently blocked by supernatant from AdG12-transduced cells, but only in the presence of doxycycline. After liver-specific transfer, AdG12 (plus doxycycline) significantly improved cardiac contractility and diastolic relaxation compared with a control vector in CVB3-infected mice if sCAR-Fc was induced before infection (left ventricular pressure 59+/-3.8 versus 45.4+/-2.7 mm Hg, median 59 versus 45.8 mm Hg, P<0.01; dP/dt(max) 3645.1+/-443.6 versus 2057.9+/-490.2 mm Hg/s, median 3526.6 versus 2072 mm Hg/s, P<0.01; and dP/dt(min) -2125.5+/-330.5 versus -1310.2+/-330.3 mm Hg/s, median -2083.7 versus -1295.9 mm Hg/s, P<0.01) and improved contractility if induced concomitantly with infection (left ventricular pressure 76.4+/-19.2 versus 56.8+/-10.3 mm Hg, median 74.8 versus 54.4 mm Hg, P<0.05; dP/dt(max) 5214.2+/-1786.2 versus 3011.6+/-918.3 mm Hg/s, median 5182.1 versus 3106.6 mm Hg/s, P<0.05), respectively. Importantly, hemodynamics of animals treated with AdG12 (plus doxycycline) were similar to uninfected controls. Preinfection induction of sCAR-Fc completely blocked and concomitant induction strongly reduced cardiac CVB3 infection, myocardial injury, and inflammation. CONCLUSIONS: AdG12-mediated sCAR-Fc delivery prevents cardiac dysfunction in CVB3 myocarditis under prophylactic and therapeutic conditions.

Assessment of stability, toxicity and immunogenicity of new polymeric nanoreactors for use in enzyme replacement therapy of MNGIE.

The lack of a crucial metabolic enzyme can lead to accumulating substrate concentrations in the bloodstream and severe human enzyme deficiency diseases. Mitochondrial Neurogastrointestinal Encephalomyopathy (MNGIE) is such a fatal genetic disorder, caused by a thymidine phosphorylase deficiency. Enzyme replacement therapy is a strategy where the deficient enzyme is administered intravenously in order to decrease the toxic substrate concentrations. Such a therapy is however not very efficient due to the fast elimination of the native enzyme from the circulation. In this study we evaluate the potential of using polymeric enzyme-loaded nanoparticles to improve the delivery of therapeutic enzymes. We constructed new 200-nanometer PMOXA-PDMS-PMOXA polymeric nanoparticles that encapsulate the enzyme thymidine phosphorylase. These particles are permeabilised for substrates and products by the reconstitution of the nucleoside-specific porin Tsx in their polymeric wall. We show that the obtained 'nanoreactors' are enzymatically active and stable in blood serum at 37 degrees C. Moreover, they do not provoke cytotoxicity when incubated with hepatocytes for 4 days, nor do they induce a macrophage-mediated inflammatory response ex vivo and in vivo. All data highlight the potential of such nanoreactors for their application in enzyme replacement therapy of MNGIE.

Expression of coxsackie and adenovirus receptor reduces the lung metastatic potential of murine tumor cells.

The coxsackie and adenovirus receptor (CAR) is involved in the epithelial cell tight junction, the downregulated expression of which is observed in different cancer types. In the present study, we examined CAR's role in tumor metastasis using a B16 melanoma and CT26 colon adenocarcinoma model of experimental metastasis. In lung metastasis, the colony number of B16 cells stably expressing CAR (B16CAR) was significantly lower than that of the control CAR-negative B16 cells. B16 and CT26 cells transiently expressing CAR, which were transduced with adenovirus (Ad) vector expressing CAR, also reduced lung metastasis, suggesting that CAR plays a role in the early stage of metastasis. CAR expression significantly decreased the accumulation of B16 cells in the lung after i.v. injection and the migration in vitro. CAR expression reduced expression of alpha(v), alpha(4), beta(3) and beta(1) integrin, which play important roles in attachment to cells or basement membrane. Thus, CAR expression likely acts as a metastatic suppressor.

The Tim-3 ligand galectin-9 negatively regulates CD8+ alloreactive T cell and prolongs survival of skin graft.

CD8+ alloreactive T cells are the key mediators of accelerated rejection. Vigorous CD8+ alloreactive T cells responses against alloantigens, which is the main effector mechanism in acute allograft rejection, has been well described. But the molecular mechanisms to dampen activated CD8+ T cells are largely unknown. On the other hand, Tim-3 is a molecule expressed on terminally differentiated CD4+ Th1 cells. Engaging Tim-3 with its ligand galectin-9 causes an inhibitory signal, resulting in apoptosis of Th1 cells and negatively regulates Th1 type immunity. However, the question whether CD8+ T cells express surface molecular Tim-3 has not been fully elucidated. In this study, we have investigated which CD8+ subset express molecular Tim-3 by flow cytometric assay. In addition, cytotoxic assay was applied to analyze whether CD8+ alloreactive T cells were sensitive to galectin-9 induced apoptosis. Here, our results demonstrated that Tim-3 was expressed on activated CD8+ alloreactive T cells (CD8+CD44highCD62Llow), but not expressed on naïve CD8+ T cells. Furthermore, alloreactive CD8+ cytotoxic T cells were sensitive to galectin-9 induced apoptosis both in vitro and vivo, resulting in attenuation of CD8+ alloreactive T cells mediated cytotoxicity and prolonged survival of skin graft.

Therapeutic nanoreactors: combining chemistry and biology in a novel triblock copolymer drug delivery system.

Triblock copolymeric nanoreactors are introduced as an alternative for liposomes as encapsulating carrier for prodrug activating enzymes. Inosine-adenosine-guanosine preferring nucleoside hydrolase of Trypanosoma vivax, a potential prodrug activating enzyme, was encapsulated in nanometer-sized vesicles constructed of poly(2-methyloxazoline)-block-poly(dimethylsiloxane)-block-(2-methyloxazoline) triblock copolymers. The nanoreactor is functionalized by incorporation of bacterial porins, OmpF or Tsx, in the reactor wall. Efficient cleavage of three natural substrates and one prodrug, 2-fluoroadenosine, by the nanoreactors was demonstrated.

Designing CD4 immunoadhesins for AIDS therapy.

A newly-constructed antibody-like molecule containing the gp120-binding domain of the receptor for human immunodeficiency virus blocks HIV-1 infection of T cells and monocytes. Its long plasma half-life, other antibody-like properties, and potential to block all HIV isolates, make it a good candidate for therapeutic use.