IL-8 correlates with nonresponse to neoadjuvant nivolumab in HPV positive HNSCC via a potential extracellular vesicle miR-146a mediated mechanism.

Therapy using anti-PD-1 immune checkpoint inhibitors (ICI) has revolutionized the treatment of many cancers including head and neck squamous cell carcinomas (HNSCC), but only a fraction of patients respond. To better understand the molecular mechanisms driving resistance, we performed extensive analysis of plasma and tumor tissues before and after a 4-week neoadjuvant trial in which HNSCC patients were treated with the anti-PD-1 inhibitor, nivolumab. Luminex cytokine analysis of patient plasma demonstrated that HPVpos nonresponders displayed high levels of the proinflammatory chemokine, interleukin-8 (IL-8), which decreased after ICI treatment, but remained higher than responders. miRNAseq analysis of tetraspanin-enriched small extracellular vesicles (sEV) purified from plasma of HPVpos nonresponders demonstrated significantly lower levels of seven miRNAs that target IL-8 including miR-146a. Levels of the pro-survival oncoprotein Dsg2, which has been to down-regulate miR-146a, are elevated with HPVpos tumors displaying higher levels than HPVneg tumors. Dsg2 levels decrease significantly following ICI in responders but not in nonresponders. In cultured HPVpos cells, restoration of miR-146a by forced expression or treatment with miR-146a-loaded sEV, reduced IL-8 level, blocked cell cycle progression, and promoted cell death. These findings identify Dsg2, miR-146a, and IL-8 as potential biomarkers for ICI response and suggest that the Dsg2/miR-146a/IL-8 signaling axis negatively impacts ICI treatment outcomes and could be targeted to improve ICI responsiveness in HPVpos HNSCC patients.

Unique Photochemo-Immuno-Nanoplatform against Orthotopic Xenograft Oral Cancer and Metastatic Syngeneic Breast Cancer.

Sophisticated self-assembly may endow materials with a variety of unique functions that are highly desirable for therapeutic nanoplatform. Herein, we report the coassembly of two structurally defined telodendrimers, each comprised of hydrophilic linear PEG and hydrophobic cholic acid cluster as a basic amphiphilic molecular subunit. One telodendrimer has four added indocyanine green derivatives, leading to excellent photothermal properties; the other telodendrimer has four sulfhydryl groups designed for efficient intersubunit cross-linking, contributing to superior stability during circulation. The coassembled nanoparticle (CPCI-NP) possesses superior photothermal conversion efficiency as well as efficient encapsulation and controlled release of cytotoxic molecules and immunomodulatory agents. CPCI-NP loaded with doxorubicin has proven to be a highly efficacious combination photothermal/chemotherapeutic nanoplatform against orthotopic OSC-3 oral cancer xenograft model. When loaded with imiquimod, a potent small molecule immunostimulant, CPCI-NP is found to be highly effective against 4T1 syngeneic murine breast cancer model, particularly when photothermal/immuno-therapy is given in combination with PD-1 checkpoint blockade antibody. Such triple therapy not only eradicates the light-irradiated primary tumors, but also activates systemic antitumor immunoactivity, causing tumor death at light-unexposed distant tumor sites. This coassembled multifunctional, versatile, and easily scalable photothermal immuno-nanoplatform shows great promise for clinical translation.

Validation of a Novel Cognitive Simulator for Orbital Floor Reconstruction.

PURPOSE: The increasing focus on patient safety in current medical practice has promoted the development of surgical simulation technology in the form of virtual reality (VR) training designed largely to improve technical skills and less so for nontechnical aspects of surgery such as decision making and material knowledge. The present study investigated the validity of a novel cognitive VR simulator called Touch Surgery for a core maxillofacial surgical procedure: orbital floor reconstruction (OFR). MATERIALS AND METHODS: A cross-sectional study was carried out on 2 groups of participants with different experience levels. Novice graduate dental students and expert surgeons were recruited from a local dental school and academic residency programs, respectively. All participants completed the OFR module on Touch Surgery. The primary outcome variable was simulator performance score. Post-module questionnaires rating specific aspects of the simulation experience were completed by the 2 groups and served as the secondary outcome variables. The age and gender of participants were considered additional predictor variables. From these data, conclusions were made regarding 3 types of validity (face, content, and construct) for the Touch Surgery simulator. Dependent-samples t tests were used to explore the consistency in simulation performance scores across phases 1 and 2 by experience level. Two multivariate ordinary least-squares regression models were fit to estimate the relation between experience and phase 1 and 2 scores. RESULTS: Thirty-nine novices and 10 experts who were naïve to Touch Surgery were recruited for the study. Experts outperformed novices on phases 1 and 2 of the OFR module (P < .001), which provided the measurement of construct validation. Responses to the questionnaire items used to assess face validity were favorable from the 2 groups. Positive questionnaire responses also were recorded from experts alone on items assessing the content validity for the module. Participant age and gender were not relevant predictors of performance scores. CONCLUSION: Construct, content, and face validities were observed for the OFR module on a novel cognitive simulator, Touch Surgery. Therefore, OFR simulation on the smart device platform could serve as a useful cognitive training and assessment tool in maxillofacial surgery residency programs.

Cytokine Profiling in Low- and High-Density Small Extracellular Vesicles from Epidermoid Carcinoma Cells.

Exosomes or small extracellular vesicles (sEVs) are membrane-bound nanoparticles that carry various macromolecules and act as autocrine and paracrine signaling messengers. In this study, sEVs from epidermoid carcinoma cells influenced by membrane presentation of the glycoprotein desmoglein 2 and its palmitoylation state were investigated. In this study, sEVs were isolated by sequential ultracentrifugation followed by iodixanol density gradient separation. They were then subjected to multiplex profiling of cytokines associated with the surface of intact sEVs. The results revealed a previously undescribed active sorting of cytokines onto the surface of low-density and high-density sEV subpopulations. Specifically, an altered surface presentation of desmoglein 2 decreased FGF-2 and VEGF in low-density sEVs. In addition, in response to desmoglein 2, IL-8 and RANTES were increased in low-density sEVs but only slightly decreased in high-density sEVs. Finally, IL-6 and G-CSF were increased dramatically in high-density sEVs. This comprehensive analysis of the cytokine production profile by squamous cell carcinoma‒derived sEVs highlights their contribution to immune evasion, pro-oncogenic and proangiogenic activity, and the potential to identify diagnostic disease biomarkers.

Neural capacity limits on the responses to memory interference during working memory in young and old adults.

Advancing age affects the recruitment of task related neural resources thereby changing the efficiency, capacity and use of compensatory processes. With advancing age, brain activity may therefore increase within a region or be reorganized to utilize different brain regions. The different brain regions may be exclusive to old adults or accessible to young and old alike, but non-optimal. Interference during verbal working memory information retention recruits parahippocampal brain regions in young adults similar to brain activity recruited by old adults in the absence of external interference. The current work tests the hypothesis that old adults recruit neural resources to combat increases in age-related intrinsic noise that young adults recruit during high levels of interference during information retention. This experiment administered a verbal delayed item recognition task with low and high levels of an interfering addition task during information maintenance. Despite strong age-related behavioral effects, brain imaging results demonstrated no significant interaction effects between age group and the interference or memory tasks. Significant effects were only found for the interaction between interference level and memory load within the inferior frontal cortex, supplementary motor cortex and posterior supramarginal regions. Results demonstrate that neural resources were shared when facing increasing memory load and interference. The combined cognitive demands resulted in brain activity reaching a neural capacity limit which was similar for both age groups and which brain activation did not increase above. Despite significant behavioral differences the neural capacity limited the detection of age group differences in brain activity.

miRNA- and cytokine-associated extracellular vesicles mediate squamous cell carcinomas.

Exosomes, or small extracellular vesicles (sEVs), serve as intercellular messengers with key roles in normal and pathological processes. Our previous work had demonstrated that Dsg2 expression in squamous cell carcinoma (SCC) cells enhanced both sEV secretion and loading of pro-mitogenic cargo. In this study, using wild-type Dsg2 and a mutant form that is unable to be palmitoylated (Dsg2cacs), we investigated the mechanism by which Dsg2 modulates SCC tumour development and progression through sEVs. We demonstrate that palmitoylation was required for Dsg2 to regulate sub-cellular localisation of lipid raft and endosomal proteins necessary for sEV biogenesis. Pharmacological inhibition of the endosomal pathway abrogated Dsg2-mediated sEV release. In murine xenograft models, Dsg2-expressing cells generated larger xenograft tumours as compared to cells expressing GFP or Dsg2cacs. Co-treatment with sEVs derived from Dsg2-over-expressing cells increased xenograft size. Cytokine profiling revealed, Dsg2 enhanced both soluble and sEV-associated IL-8 and miRNA profiling revealed, Dsg2 down-regulated both cellular and sEV-loaded miR-146a. miR-146a targets IRAK1, a serine-threonine kinase involved in IL-8 signalling. Treatment with a miR-146a inhibitor up-regulated both IRAK1 and IL-8 expression. RNAseq analysis of HNSCC tumours revealed a correlation between Dsg2 and IL-8. Finally, elevated IL-8 plasma levels were detected in a subset of HNSCC patients who did not respond to immune checkpoint therapy, suggesting that these patients may benefit from prior anti-IL-8 treatment. In summary, these results suggest that intercellular communication through cell-cell adhesion, cytokine release and secretion of EVs are coordinated, and critical for tumour growth and development, and may serve as potential prognostic markers to inform treatment options. ABBREVIATIONS: Basal cell carcinomas, BCC; Betacellulin, BTC; 2-bromopalmitate, 2-Bromo; Cluster of differentiation, CD; Cytochrome c oxidase IV, COX IV; Desmoglein 2, Dsg2; Early endosome antigen 1, EEA1; Epidermal growth factor receptor substrate 15, EPS15; Extracellular vesicle, EV; Flotillin 1, Flot1; Glyceraldehyde-3-phosphate dehydrogenase, GAPH; Green fluorescent protein, GFP; Head and neck squamous cell carcinoma, HNSCC; Interleukin-1 receptor-associated kinase 1, IRAK1; Interleukin 8, IL-8; Large EV, lEV; MicroRNA, miR; Palmitoylacyltransferase, PAT; Ras-related protein 7 Rab7; Small EV, sEV; Squamous cell carcinoma, SCC; Tissue inhibitor of metalloproteinases, TIMP; Tumour microenvironment, TME.