Manipulating TLR signaling increases the anti-tumor T Cell response induced by viral cancer therapies

The immune response plays a key role in enhancing the therapeutic activity of oncolytic virotherapies. However, to date, investigators have relied on inherent interactions between the virus and the immune system, often coupled to the expression of a single cytokine transgene. Recently, the importance of TLR activation in mediating adaptive immunity has been demonstrated. We therefore sought to influence the type and level of immune response raised after oncolytic vaccinia therapy through manipulation of TLR signaling. Vaccinia naturally activates TLR2, associated with an antibody response, whereas a CTL response is associated with TLR3-TRIF-signaling pathways. We manipulated TLR signaling by vaccinia through deglycosylation of the viral particle to block TLR2 activation and expression of a TRIF transgene. The resulting vector displayed greatly reduced production of anti-viral neutralizing antibody as well as an increased anti-tumor CTL response. Delivery in both naive and pre-treated mice was enhanced and immunotherapeutic activity dramatically improved

Fluoromodule-based reporter/probes designed for in vivo fluorescence imaging

Optical imaging of whole, living animals has proven to be a powerful tool in multiple areas of preclinical research and has allowed noninvasive monitoring of immune responses, tumor and pathogen growth, and treatment responses in longitudinal studies. However, fluorescence-based studies in animals are challenging because tissue absorbs and autofluoresces strongly in the visible light spectrum. These optical properties drive development and use of fluorescent labels that absorb and emit at longer wavelengths. Here, we present a far-red absorbing fluoromodule-based reporter/probe system and show that this system can be used for imaging in living mice. The probe we developed is a fluorogenic dye called SC1 that is dark in solution but highly fluorescent when bound to its cognate reporter, Mars1. The reporter/probe complex, or fluoromodule, produced peak emission near 730 nm. Mars1 was able to bind a variety of structurally similar probes that differ in color and membrane permeability. We demonstrated that a tool kit of multiple probes can be used to label extracellular and intracellular reporter-tagged receptor pools with 2 colors. Imaging studies may benefit from this far-red excited reporter/probe system, which features tight coupling between probe fluorescence and reporter binding and offers the option of using an expandable family of fluorogenic probes with a single reporter gene

A convenient and high-yield synthesis for pyridinium and metal hexafluorocomplexes of vanadium, chromium, iron and cobalt

Pyridinium poly(hydrogen fluoride) reacts with the oxide of vanadium(V) and chlorides of chromium(III), iron (III) and Co(II) at room temperature forming the pyridinium salts of hexafluoro vanadate(V), hexafluorochromate(III), hexafluoroferrate(III) and hexafluorocobaltate(II) in near quantitative yields (80%). These pyridinium salts are the precursors for the preparation of the alkali metal hexafluorometallates by metathetic reactions in acetonitrile medium with the corresponding metal chlorides. The prepared salts have been identified by their infrared spectral data and elemental analysis

Displacement of Lewis Acid Gases - PF5,BF3PF_5, BF_3 and SiF4SiF_4 from Their Ammonium, Alkali Metal and Pyridinium Fluorocomplexes by Sulphur Trioxide at Room Temperature

Lewis acid gases, $PF_5, BF_3$ and $SiF4$ are liberated from their ammonium, alkali metal and pyridinium fluorocomplexes, $NH_4PF_6, NaPF_6, KPF_6, C_5H_5NHPF_6, C_5H_5NHBF_4$ and $(C_5H_5NH)_2SiF_6$, when treated with sulphur trioxide at room temperature. The dissociated ammonium/alkali metal fluorides as well as the pyridinium fluoride react further with the excess sulphur trioxide to form the corresponding fluorosulphates. The products, Lewis acid gases and the fluorosulphates, have been 19 characterised by I. R., F NMR and wet chemical analysis. This is the first report of the synthesis of these gases from pyridinium fluorosulphate at room temperature in good yields

Reaction of thiophosphoryl fluoride with sulphur trioxide

Thiophosphoryl fluoride is observed to undergo a facile reaction with sulphur trioxide forming phosphoryl fluoride, sulphur dioxide and elemental sulphur in quantitative yields. In the presence of excess of sulphur trioxide, however, the elemental sulphur released combines with it to form sulphur sesquioxide which subsequently decomposes and gives off sulphur dioxide. Similar observations are made with oleum

On the mechanism of anti-CD39 immune checkpoint therapy

With the coming of age of cancer immunotherapy, the search for new therapeutic targets has led to the identification of immunosuppressive adenosine as an important regulator of antitumor immunity. This resulted in the development of selective inhibitors targeting various components of the adenosinergic pathway, including small molecules antagonists targeting the high affinity A2A adenosine receptor and low affinity A2B receptor, therapeutic monoclonal antibodies (mAbs) and small molecules targeting CD73 and therapeutic mAbs targeting CD39. As each regulator of the adenosinergic pathway present non-overlapping biologic functions, a better understanding of the mechanisms of action of each targeted approach should accelerate clinical translation and improve rational design of combination treatments. In this review, we discuss the potential mechanisms-of-action of anti-CD39 cancer therapy and potential toxicities that may emerge from sustained CD39 inhibition. Caution should be taken, however, in extrapolating data from gene-targeted mice to patients treated with blocking anti-CD39 agents. As phase I clinical trials are now underway, further insights into the mechanism of action and potential adverse events associated with anti-CD39 therapy are anticipated in coming years

CCL5-armed oncolytic virus augments CCR5-engineered NK cell infiltration and antitumor efficiency

BackgroundNatural killer (NK) cells have potent antitumor activities. Nevertheless, adoptive transfer therapy of NK cells has gained very limited success in patients with solid tumors as most infused NK cells remain circulating in the peripheral blood instead of entering tumor sites. Chemokines and their receptors play important roles in NK cell distribution. Enhancing chemokine receptors on immune cells to match and be driven to tumor-specific chemokines may improve the therapeutic efficacy of NK cells.MethodsThe CCR5-CCL5 axis is critical in NK cell homing to tumor sites. Thus, we analyzed CCR5 expression on NK cells from patients with cancer and healthy donors. We then upregulated CCR5 and CCL5 with lentiviruses and oncolytic viruses in NK and tumor cells, respectively. Animal experiments were also carried out to test the efficacy of the combination of oncolytic virus with NK cells.ResultsIn NK cells from patients with various solid tumors or healthy subjects, CCR5 was expressed at low levels before and after expansion in vitro. CCR5-engineered NK cells showed enhanced tumor infiltration and antitumor effects, but no complete regressions were noted in the in vivo tumor models. To further improve therapeutic efficacy, we constructed CCL5-expressing oncolytic vaccinia virus. In vitro data demonstrated that vaccinia virus can produce CCL5 in tumor cells while infectivity remained unaffected. Supernatants from tumor cells infected by CCL5-modified vaccinia virus enhanced the directional movement of CCR5-overexpressed NK cells but not green fluorescent protein (GFP)-expressing cells. More importantly, NK cells were resistant to the vaccinia virus and their functions were not affected after being in contact. In vivo assays demonstrated that CCL5-expressing vaccinia virus induced a greater accumulation of NK cells within tumor lesions compared with that of the prototype virus.ConclusionEnhancement of matched chemokines and chemokine receptors is a promising method of increasing NK cell homing and therapeutic effects. Oncolytic vaccinia viruses that express specific chemokines can synergistically augment the efficacies of NK cell-based therapy

Oxidation of Phosphorus Trifluoride with Sulphur Trioxide

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Placental TLR/NLR expression signatures are altered with gestational age and inflammation

<p><b>Objective:</b> To quantify changes in placental expression of Toll-like receptors (TLRs) and nuclear oligomerization domain (NOD)-like receptors (NLRs) gene with (1) advancing gestational age (GA) and (2) exposure to chorioamnionitis (CA) and preterm premature rupture of membrane (PPROM).</p> <p><b>Methods:</b> Placental tissue was collected at the time of birth from 83 subjects with live birth pregnancies from 24- to 40-week gestation between 2009 and 2013. Real-time RT-PCR analysis of 13 TLR/NLR genes involved in bacterial sensing was performed using specific probes.</p> <p><b>Results:</b> Of 83 patients enrolled, 61 were preterm (<37 weeks). 23 (27%) had evidence of CA; and 33 (39.8%) had PPROM. 15 (18%) had both CA and PPROM (CP). 42 (50%) had neither CA nor PPROM (C/P). Only RIPK2 (<i>p</i> = 0.0025) and TLR4 (<i>p</i> = 0.0005) were found to increase progressively with GA. We found significant changes in TLR5 (<i>p</i> = 0.01) with CA, NFKBIA (<i>p</i> = 0.016) with PPROM, NKKBIA (<i>p</i> = 0.003), and NFKB1 (<i>p</i> = 0.009) with CA and PPROM.</p> <p><b>Conclusion:</b> RIPK2 (mediator of NOD-dependent NF-kB signaling) and TLR4 progressively increased with GA. We speculate this upregulation may be involved in initiating labor and delivery at term. Increase in NFKBIA seen in PPROM and CA might represent a counter regulatory mechanism to decrease inflammation in these conditions. This study provides new information on relationships between GA, CA/PPROM, and TLR/NLR signaling in the placenta.</p