VTT-006, an anti-mitotic compound, binds to the Ndc80 complex and suppresses cancer cell growth <i>in vitro</i>.

Hec1 (Highly expressed in cancer 1) resides in the outer kinetochore where it works to facilitate proper kinetochore-microtubule interactions during mitosis. Hec1 is overexpressed in various cancers and its expression shows correlation with high tumour grade and poor patient prognosis. Chemical perturbation of Hec1 is anticipated to impair kinetochore-microtubule binding, activate the spindle assembly checkpoint (spindle checkpoint) and thereby suppress cell proliferation. In this study, we performed high-throughput screen to identify novel small molecules that target the Hec1 calponin homology domain (CHD), which is needed for normal microtubule attachments. 4 million compounds were first virtually fitted against the CHD, and the best hit molecules were evaluated in vitro. These approaches led to the identification of VTT-006, a 1,2-disubstituted-tetrahydro-beta-carboline derivative, which showed binding to recombinant Ndc80 complex and modulated Hec1 association with microtubules in vitro. VTT-006 treatment resulted in chromosome congression defects, reduced chromosome oscillations and induced loss of inter-kinetochore tension. Cells remained arrested in mitosis with an active spindle checkpoint for several hours before undergoing cell death. VTT-006 suppressed the growth of several cancer cell lines and enhanced the sensitivity of HeLa cells to Taxol. Our findings propose that VTT-006 is a potential anti-mitotic compound that disrupts M phase, impairs kinetochore-microtubule interactions, and activates the spindle checkpoint

Comparison of Clinically Relevant Oncolytic Virus Platforms for Enhancing T Cell Therapy of Solid Tumors

Despite some promising results, the majority of patients do not benefit from T cell therapies, as tumors prevent T cells from entering the tumor, shut down their activity, or downregulate key antigens. Due to their nature and mechanism of action, oncolytic viruses have features that can help overcome many of the barriers currently facing T cell therapies of solid tumors. This study aims to understand how four different oncolytic viruses (adenovirus, vaccinia virus, herpes simplex virus, and reovirus) perform in that task. For that purpose, an immunocompetent in vivo tumor model featuring adoptive tumor-infiltrating lymphocyte (TIL) therapy was used. Tumor growth control (p < 0.001) and survival analyses suggest that adenovirus was most effective in enabling T cell therapy. The complete response rate was 62% for TILs + adenovirus versus 17.5% for TILs + PBS. Of note, TIL biodistribution did not explain efficacy differences between viruses. Instead, immunostimulatory shifts in the tumor microenvironment mirrored efficacy results. Overall, the use of oncolytic viruses can improve the utility of T cell therapies, and additional virus engineering by arming with transgenes can provide further antitumor effects. This phenomenon was seen when an unarmed oncolytic adenovirus was compared to Ad5/3-E2F-d24-hTNFa-IRES-hIL2 (TILT-123). A clinical trial is ongoing, where patients receiving TIL treatment also receive TILT-123 (ClinicalTrials.gov: NCT04217473).</p

Microbial Expansin Related Proteins for Improved Enzymatic Action for Conversion of Lignocellulose to Platform Sugars

| openaire: EC/H2020/964764/EU//BioUPGRADECost effective enzymatic deconstruction of cellulosic materials for the production of renewable fuels and chemicals is challenged by the limited accessibility of enzymes to cellulose substrates. Recently identified loosenin-like proteins (PcaLOOLs from Phanerochaete carnosa) can promote amorphogenesis and disrupt noncovalent binding between cellulose fibrils, thus promoting cell wall loosening and increasing surface accessibility for enzymatic hydrolysis. Like other microbial expansin-related proteins characterized to date, loosenins do not display hydrolytic activity; instead, they reportedly boost hydrolysis by cellulolytic enzymes although the impact of expansin-related proteins on cellulolytic activity is substrate dependent (Liu et al., 2015). Herein, the impact of four PcaLOOLs on the activity of different cellulolytic enzymes was evaluated using a lignocellulosic substrate from softwood fiber. The PcaLOOLs were heterologously produced in Pichia pastoris in shake-flask and 5-L bioreactor systems, andthen purified by affinity chromatography. Notably, the bioreactor production increased PcaLOOL yield over 10-times. Each PcaLOOL was used to pretreat the lignocellulosic material prior to the addition of cellulolytic enzyme; alternatively, each PcaLOOL was added directly with the cellulolytic enzyme to the lignocellulosic substrate. The release of soluble sugars was quantified at regular time points over 24-hours using the 4-hydroxybenzoic acid hydrazide (PAHBAH) assay and by high-performance anion exchange chromatography/pulsed amperometric detection (HPAEC-PAD). The hydrolysis reactions were performed at 40 °C and 50 °C to evaluate the impact of temperature on the boosting performance of PcaLOOLs. Briefly, all PcaLOOLs increased the enzymatic release of soluble sugars from the lignocellulosic substrate. On-going HPAEC analyses will evaluate the impact of PcaLOOLs on the profile of sugars released from the lignocellulosic substrate. In addition to lignocellulose deconstruction, the potential of PcaLOOLs topromote fiber defibrillation will be investigated.Non peer reviewe