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Tumour-host immune interactions lead to complex changes in the tumour microenvironment (TME), impacting progression, metastasis and response to therapy. While it is clear that cancer cells can have the capacity to alter immune landscapes, our understanding of this process is incomplete. Herein we show that endocytic trafficking at the plasma membrane, mediated by the small GTPase ARF6, enables melanoma cells to impose an immunosuppressive TME that accelerates tumour development. This ARF6-dependent TME is vulnerable to immune checkpoint blockade therapy (ICB) but in murine melanoma, loss of Arf6 causes resistance to ICB. Likewise, downregulation of ARF6 in patient tumours correlates with inferior overall survival after ICB. Mechanistically, these phenotypes are at least partially explained by ARF6-dependent recycling, which controls plasma membrane density of the interferon-gamma receptor. Collectively, our findings reveal the importance of endomembrane trafficking in outfitting tumour cells with the ability to shape their immune microenvironment and respond to immunotherapy.
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Factor 6 de Ribosilación del ADP , Factores de Ribosilacion-ADP , Membrana Celular , Inhibidores de Puntos de Control Inmunológico , Melanoma , Microambiente Tumoral , Microambiente Tumoral/inmunología , Animales , Humanos , Ratones , Factores de Ribosilacion-ADP/metabolismo , Factores de Ribosilacion-ADP/genética , Inhibidores de Puntos de Control Inmunológico/farmacología , Inhibidores de Puntos de Control Inmunológico/uso terapéutico , Melanoma/genética , Melanoma/tratamiento farmacológico , Melanoma/metabolismo , Melanoma/patología , Melanoma/inmunología , Línea Celular Tumoral , Membrana Celular/metabolismo , Receptor de Interferón gamma , Receptores de Interferón/metabolismo , Receptores de Interferón/genética , Transporte de Proteínas , Melanoma Experimental/inmunología , Melanoma Experimental/metabolismo , Melanoma Experimental/patología , Melanoma Experimental/genética , Ratones Endogámicos C57BL , FemeninoRESUMEN
Nasopharyngeal carcinoma (NPC) is closely associated with Epstein-Barr Virus infection (EBV). Despite ubiquitous EBV infection worldwide, NPC displays a unique geographical distribution in Southern China and Southeast Asia. This observed phenomenon can be attributed to the interplay of different strains of EBV infection with host genetics and environmental factors. Polymorphisms on the EBV BALF2 gene have been shown to influence risk of nasopharyngeal carcinoma (NPC). Notably, two non-synonymous EBV polymorphisms (162476T>C, 163364C>T) account for majority of NPC risk in endemic regions. These polymorphisms confer amino acid changes (I1613V, V317M) within the BALF2 protein. However, their impact on NPC tumor biology is unknown. We evaluated the distribution of BALF2 risk polymorphisms in five independent genomic datasets comprising 351 NPC clinical samples, confirming the high prevalence of high-risk EBV strains in NPC. Importantly, we observed two biologically distinct groups of tumors based on their gene expression profiles when grouped by their EBV risk strains. NPC tumors with the V317M substitution demonstrated increased proliferation processes including cell cycle (NES = 1.71, p = 5.64x10-24) and keratinization (NES = 2.42, p = 6.95x10-17). In contrast, NPC tumors without the V317M substitution demonstrated increased immune-related processes, including cell activation (NES = 1.85, p = 8.29x10-31), myeloid leukocyte activation (NES = 2.16, p = 6.51x10-24) and leukocyte mediated immunity (NES = 1.99, p = 1.05x10-23). These findings provide further insight on the influence of BALF2 variants on NPC tumor biology. EBV risk strains may have the potential to define biologically important groups in NPC.
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Prodrugs have been explored as an alternative to conventional chemotherapy; however, their target specificity remains limited. The tumor microenvironment harbors a range of microorganisms that potentially serve as tumor-targeting vectors for delivering prodrugs. In this study, we harness bacteria-cancer interactions native to the tumor microbiome to achieve high target specificity for prodrug delivery. We identify an oral commensal strain of Lactobacillus plantarum with an intrinsic cancer-binding mechanism and engineer the strain to enable the surface loading of anticancer prodrugs, with nasopharyngeal carcinoma (NPC) as a model cancer. The engineered commensals show specific binding to NPC via OppA-mediated recognition of surface heparan sulfate, and the loaded prodrugs are activated by tumor-associated biosignals to release SN-38, a chemotherapy compound, near NPC. In vitro experiments demonstrate that the prodrug-loaded microbes significantly increase the potency of SN-38 against NPC cell lines, up to 10-fold. In a mouse xenograft model, intravenous injection of the engineered L. plantarum leads to bacterial colonization in NPC tumors and a 67% inhibition in tumor growth, enhancing the efficacy of SN-38 by 54%.
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Lactobacillus plantarum , Profármacos , Ensayos Antitumor por Modelo de Xenoinjerto , Profármacos/farmacología , Profármacos/uso terapéutico , Animales , Humanos , Ratones , Línea Celular Tumoral , Neoplasias Nasofaríngeas/tratamiento farmacológico , Neoplasias Nasofaríngeas/terapia , Neoplasias Nasofaríngeas/patología , Neoplasias Nasofaríngeas/microbiología , Carcinoma Nasofaríngeo/tratamiento farmacológico , Carcinoma Nasofaríngeo/terapia , Carcinoma Nasofaríngeo/patología , Microambiente Tumoral/efectos de los fármacos , Antineoplásicos/farmacología , Antineoplásicos/uso terapéutico , Ratones Desnudos , Femenino , Ratones Endogámicos BALB CRESUMEN
BACKGROUND: Epstein-Barr virus (EBV) IgA serology for viral capsid antigen (VCA) and early antigen (EA) aids early detection of nasopharyngeal cancer (NPC), resulting in improved survival. We evaluated the diagnostic performance of a prefabricated immunofluorescent assay (IFA) for NPC screening in high-risk individuals. METHODS: Sera from 96 biopsy-proven patients with NPC diagnosed at the outpatient clinic and 96 healthy family members were tested for EBV-VCA IgA and EBV-EA IgA using the prefabricated IFA from EUROIMMUN (EI) and the traditional immunofluorescence method. RESULTS: The AUC of EI EBV-VCA IgA and EBV-EA IgA was 0.907 (95% confidence interval [CI]: 0.894-0.965) and 0.898 (95% CI: 0.848-0.947), respectively. Combined testing with the prefabricated assay at a threshold of VCA ≥1:320 or EA ≥1:10 showed 92.7% sensitivity and 81.2% specificity. Overall, the traditional EBV-EA IgA assay demonstrated the best accuracy (sensitivity 91.7% and specificity 96.9%) at a threshold of ≥1:5. CONCLUSION: While the traditional IFA method was more accurate, the prefabricated IFA test kit can be a useful tool for NPC screening in high-risk populations.
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Detección Precoz del Cáncer , Inmunoglobulina A , Neoplasias Nasofaríngeas , Humanos , Neoplasias Nasofaríngeas/diagnóstico , Neoplasias Nasofaríngeas/virología , Masculino , Femenino , Persona de Mediana Edad , Detección Precoz del Cáncer/métodos , Adulto , Inmunoglobulina A/sangre , Antígenos Virales/inmunología , Infecciones por Virus de Epstein-Barr/diagnóstico , Técnica del Anticuerpo Fluorescente , Sensibilidad y Especificidad , Anciano , Herpesvirus Humano 4/inmunología , Herpesvirus Humano 4/aislamiento & purificación , Anticuerpos Antivirales/sangre , Proteínas de la Cápside/inmunologíaRESUMEN
Nasopharyngeal carcinoma (NPC) is an Epstein-Barr virus (EBV) driven malignancy arising from the nasopharyngeal epithelium. Current treatment strategies depend on the clinical stage of the disease, including the extent of the primary tumour, the extent of nodal disease, and the presence of distant metastasis. With the close association of EBV infection with NPC development, EBV biomarkers have shown promise in predicting treatment outcomes. Among the omic technologies, RNA and miRNA signatures have been widely studied, showing promising results in the research setting to predict treatment response. The transformation of radiology images into measurable features has facilitated the use of radiomics to generate predictive models for better prognostication and treatment selection. Nonetheless, much of this work remains in the research realm, and challenges remain in clinical implementation.
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OBJECTIVES: To characterize longitudinal changes in Epstein-Barr virus (EBV) DNA post-radiotherapy in nasopharyngeal carcinoma (NPC) patients, and investigate whether an early (0-2 weeks) or delayed (8-12 weeks) EBV DNA result better predicts for disease-free survival (DFS). MATERIALS AND METHODS: Histologically-confirmed NPC patients with ≥1 EBV DNA test quantified using the harmonized BamHI-W polymerase chain reaction-based assay at 0-2 and 8-12 weeks post-radiotherapy were included. RESULTS: We identified 302 patients with EBV DNA measured at 0-2 weeks post-radiotherapy; of which, 110 (36.4 %) underwent a repeat test at 8-12 weeks post-treatment. Patients harboring a detectable EBV DNA at 0-2 weeks experienced an inferior DFS (adjusted HR1-264 copies 1.72 [95 %CI: 1.05-2.83], P = 0.031; AHR≥265 copies 4.39 [95 %CI: 1.68-11.44], P = 0.002 relative to 0 copies/mL). At 8-12 weeks, we observed substantial shifts in EBV DNA readings from 0 to 2 weeks; 76/110 (69.1 %) and 34/110 (30.9 %) patients at 0-2 weeks versus 90/110 (81.8 %) and 20/110 (18.2 %) at 8-12 weeks recorded undetectable and detectable EBV DNA, respectively. Positive EBV DNA at 8-12 weeks was strongly associated with relapse (73.3 % [11/15] for 1-264; 80.0 % [4/5] for ≥265 subgroups had relapses versus 15.6 % [14/90] for 0 copies/mL). Area under receiver operating curve values for 2-year relapse rates were 0.817 (95 %CI: 0.725-0.909) for stage + EBV DNA8-12w versus 0.654 (95 %CI: 0.542-0.765) for stage + EBV DNA0-2w. CONCLUSION: EBV DNA is dynamic post-radiotherapy, and delayed EBV DNA testing better enriched for higher-risk NPC patients. This implicates trials investigating adjuvant chemotherapy intensification based on early EBV DNA testing.
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Infecciones por Virus de Epstein-Barr , Neoplasias Nasofaríngeas , Humanos , Carcinoma Nasofaríngeo/patología , Herpesvirus Humano 4/genética , Infecciones por Virus de Epstein-Barr/complicaciones , Neoplasias Nasofaríngeas/patología , Pronóstico , ADN Viral , Recurrencia , Medición de RiesgoRESUMEN
The Epstein-Barr virus (EBV) is associated with various tumor types, including nasopharyngeal carcinoma and lymphoproliferative disorders. While much is known about EBV-related epithelial and lymphoid tumors, there is a paucity of knowledge concerning EBV-associated mesenchymal tumors. This review aims to provide a comprehensive overview of EBV-associated mesenchymal tumors, encompassing their clinical features, pathological characteristics, pathophysiology, prognostic factors, and current treatment approaches. Through an extensive literature search using the PubMed database, we were able to identify three distinct EBV-associated mesenchymal tumors: EBV-associated smooth muscle tumors, inflammatory pseudotumor-like follicular dendritic cell sarcomas, and EBV-associated osteosarcomas. Although this review extensively explored the different aspects of these mesenchymal tumors, our comprehension of the underlying pathophysiology in this context is still incomplete. Therefore, we hope that this review paper will not only serve as a valuable repository of information but also serve as a catalyst for prospective in vitro and in vivo research studies to bridge the existing knowledge gap surrounding pathophysiology, ultimately making an important contribution to shaping future therapeutic approaches.
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Adaptive immune resistance (AIR) is a protective process used by cancer to escape elimination by CD8+ T cells. Inhibition of immune checkpoints PD-1 and CTLA-4 specifically target Interferon-gamma (IFNγ)-driven AIR. AIR begins at the plasma membrane where tumor cell-intrinsic cytokine signaling is initiated. Thus, plasma membrane remodeling by endomembrane trafficking could regulate AIR. Herein we report that the trafficking protein ADP-Ribosylation Factor 6 (ARF6) is critical for IFNγ-driven AIR. ARF6 prevents transport of the receptor to the lysosome, augmenting IFNγR expression, tumor intrinsic IFNγ signaling and downstream expression of immunosuppressive genes. In murine melanoma, loss of ARF6 causes resistance to immune checkpoint blockade (ICB). Likewise, low expression of ARF6 in patient tumors correlates with inferior outcomes with ICB. Our data provide new mechanistic insights into tumor immune escape, defined by ARF6-dependent AIR, and support that ARF6-dependent endomembrane trafficking of the IFNγ receptor influences outcomes of ICB.
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OBJECTIVE: The objective of this meta-analysis is to evaluate the impact of genetic polymorphisms on platinum-based chemotherapy (PBC)-induced ototoxicity. DATA SOURCES: Systematic searches of PubMed, Embase, Cochrane, and Web of Science were conducted from the inception of the databases to May 31, 2022. Abstracts and presentations from conferences were also reviewed. REVIEW METHODS: Four investigators independently extracted data in adherence to the Preferred Reporting Items for Systematic Reviews and Meta-Analyses guidelines. Differences in the prevalence of PBC-induced ototoxicity between reference and variant (i) genotypes and (ii) alleles were analyzed. The overall effect size was presented using the random-effects model as an odds ratio (OR) with a 95% confidence interval (CI). RESULTS: From 32 included articles, 59 single nucleotide polymorphisms on 28 genes were identified, with 4406 total unique participants. For allele frequency analysis, the A allele in ACYP2 rs1872328 was positively associated with ototoxicity (OR: 2.61; 95% CI: 1.06-6.43; n = 2518). Upon limiting to cisplatin use only, the T allele of COMT rs4646316 and COMT rs9332377 revealed significant results. For genotype frequency analysis, the CT/TT genotype in ERCC2 rs1799793 demonstrated an otoprotective effect (OR: 0.50; 95% CI: 0.27-0.94; n = 176). Excluding studies using carboplatin or concomitant radiotherapy revealed significant effects with COMT rs4646316, GSTP1 rs1965, and XPC rs2228001. Major sources of variations between studies include differences in patient demographics, ototoxicity grading systems, and treatment protocols. CONCLUSION: Our meta-analysis presents polymorphisms that exert ototoxic or otoprotective effects in patients undergoing PBC. Importantly, several of these alleles are observed at high frequencies globally, highlighting the potential for polygenic screening and cumulative risk evaluation for personalized care.
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Antineoplásicos , Ototoxicidad , Humanos , Antineoplásicos/uso terapéutico , Ototoxicidad/tratamiento farmacológico , Platino (Metal) , Cisplatino , Polimorfismo de Nucleótido Simple , Proteína de la Xerodermia Pigmentosa del Grupo D/genética , Ácido Anhídrido Hidrolasas/genéticaRESUMEN
The effects of the COVID-19 pandemic continue to constrain health-care staff and resources worldwide, despite the availability of effective vaccines. Aerosol-generating procedures such as endoscopy, a common investigation tool for nasopharyngeal carcinoma, are recognised as a likely cause of SARS-CoV-2 spread in hospitals. Plasma Epstein-Barr virus (EBV) DNA is considered the most accurate biomarker for the routine management of nasopharyngeal carcinoma. A consensus statement on whether plasma EBV DNA can minimise the need for or replace aerosol-generating procedures, imaging methods, and face-to-face consultations in managing nasopharyngeal carcinoma is urgently needed amid the current pandemic and potentially for future highly contagious airborne diseases or natural disasters. We completed a modified Delphi consensus process of three rounds with 33 international experts in otorhinolaryngology or head and neck surgery, radiation oncology, medical oncology, and clinical oncology with vast experience in managing nasopharyngeal carcinoma, representing 51 international professional societies and national clinical trial groups. These consensus recommendations aim to enhance consistency in clinical practice, reduce ambiguity in delivering care, and offer advice for clinicians worldwide who work in endemic and non-endemic regions of nasopharyngeal carcinoma, in the context of COVID-19 and other airborne pandemics, and in future unexpected settings of severe resource constraints and insufficiency of personal protective equipment.
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COVID-19 , Infecciones por Virus de Epstein-Barr , Neoplasias Nasofaríngeas , Humanos , Pandemias/prevención & control , Herpesvirus Humano 4 , SARS-CoV-2 , Carcinoma Nasofaríngeo/terapia , ADN , Neoplasias Nasofaríngeas/diagnóstico , Neoplasias Nasofaríngeas/terapiaRESUMEN
The current understanding of genetic susceptibility factors for nasopharyngeal carcinoma (NPC) is still incomplete. To identify novel germline variants associated with NPC predisposition, we analysed whole-exome sequencing data from 119 NPC patients from Singapore with a family history of NPC and/or with early-onset NPC, together with 1337 Singaporean participants without NPC. Variants were prioritised and filtered by selecting variants with minor allele frequencies of <1% in both local control (n = 1337) and gnomAD non-cancer (EAS) (n = 9626) cohorts and a high pathogenicity prediction (CADD score > 20). Using single-variant testing, we identified 17 rare pathogenic variants in 17 genes that were associated with NPC. Consistent evidence of enrichment in NPC patients was observed for five of these variants (in JAK2, PRDM16, LRP1B, NIN, and NKX2-1) from an independent case-control comparison of 156 NPC patients and 9770 unaffected individuals. In a family with five siblings, a FANCE variant (p. P445S) was detected in two affected members, but not in three unaffected members. Gene-based burden testing recapitulated variants in NKX2-1 and FANCE as being associated with NPC risk. Using pathway analysis, endocytosis and immune-modulating pathways were found to be enriched for mutation burden. This study has identified NPC-predisposing variants and genes which could shed new insights into the genetic predisposition of NPC.
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OBJECTIVES: Evidence to support Epstein-Barr virus (EBV)-directed population nasopharyngeal carcinoma (NPC) screening has been growing. Familial aggregation is a well-recognized phenomenon in endemic regions. This systematic review summarizes the role of EBV-directed screening in individuals with a positive family history (FH+) of NPC. METHODS: We searched four electronic databases from their inception to October 2021. We included studies on individuals with FH+ of NPC who had undergone EBV-directed investigations, with no restriction in the testing methods or analytic techniques. The primary and secondary outcomes were EBV positivity rates and NPC incidence rates, respectively. Meta-analyses were performed using the random-effect model. RESULTS: Ten cross-sectional studies (n = 7436) and three cohort studies (n = 4306) were included. The pooled relative risk (RR) of EBV positivity between individuals with and without FH+ of NPC were 2.79 (95 % CI 1.37-5.68, p = 0.005) for viral capsid antigen (VCA) IgA, 3.09 (95 % CI 0.65-14.83, p = 0.16) for Epstein-Barr nuclear antigen (EBNA1) IgA, and 1.76 (95 % CI 1.04-2.96, p = 0.03) for combined EBNA1/VCA IgA. In the three cohort studies, the NPC incidence rates ranged from 90.2 to 266 per 100 000 person-years with high proportions of early-stage diseases. FH+ individuals who were EBV-positive had a 2.5 to 30.7-fold risk of NPC development compared to their EBV-negative counterparts. CONCLUSION: Family members of NPC patients had significantly higher EBV positivity rates than the general population. FH+ individuals who are EBV-positive had high risks of developing NPC. Familial screening using EBV serology may facilitate early NPC detection in endemic areas.
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Infecciones por Virus de Epstein-Barr , Neoplasias Nasofaríngeas , Anticuerpos Antivirales , Estudios Transversales , Infecciones por Virus de Epstein-Barr/complicaciones , Herpesvirus Humano 4/genética , Humanos , Inmunoglobulina A , Carcinoma Nasofaríngeo/complicacionesRESUMEN
The detection of serum Epstein-Barr virus antibodies by immunofluorescence assay (IFA) is considered the gold standard screening test for nasopharyngeal cancer (NPC) in high-risk populations. Given the high survival rate after early detection in asymptomatic patients, compared to the poor prognosis in patients with late-stage NPC, screening using IFA has tremendous potential for saving lives in the general population. However, IFA requires visual interpretation of cellular staining patterns by trained pathology staff, making it labor intensive and hence nonscalable. In this study, an automated fuzzy inference (FI) system achieved high agreement with a human IFA expert in identifying cellular patterns associated with NPC (κ = 0.82). The integration of a deep learning module into FI further improved the performance of FI (κ = 0.90) and reduced the number of uncertain cases that required manual evaluation. The performance of the resulting hybrid model, termed deep learning FI (DeLFI), was then evaluated with a separate testing set of clinical samples. In this clinical validation, DeLFI outperformed human evaluation on the area under the curve (0.926 versus 0.821) and closely matched human performance on Youden J index (0.81 versus 0.80). Data from this study indicate that the combination of deep learning with FI in DeLFI has the potential to improve the scalability and accuracy of NPC detection.
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Aprendizaje Profundo , Infecciones por Virus de Epstein-Barr , Neoplasias Nasofaríngeas , Técnica del Anticuerpo Fluorescente Indirecta , Herpesvirus Humano 4 , Humanos , Carcinoma Nasofaríngeo , Neoplasias Nasofaríngeas/diagnósticoRESUMEN
Nasopharyngeal cancer (NPC) is an Epstein-Barr virus (EBV)-positive epithelial malignancy with an extensive inflammatory infiltrate. Traditional RNA-sequencing techniques uncovered only microenvironment signatures, while the gene expression of the tumor epithelial compartment has remained a mystery. Here, we use Smart-3SEQ to prepare transcriptome-wide gene expression profiles from microdissected NPC tumors, dysplasia, and normal controls. We describe changes in biological pathways across the normal to tumor spectrum and show that fibroblast growth factor (FGF) ligands are overexpressed in NPC tumors, while negative regulators of FGF signaling, including SPRY1, SPRY2, and LGALS3, are down-regulated early in carcinogenesis. Within the NF-κB signaling pathway, the critical noncanonical transcription factors, RELB and NFKB2, are enriched in the majority of NPC tumors. We confirm the responsiveness of EBV-positive NPC cell lines to targeted inhibition of these pathways, reflecting the heterogeneity in NPC patient tumors. Our data comprehensively describe the gene expression landscape of NPC and unravel the mysteries of receptor tyrosine kinase and NF-κB pathways in NPC.
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Infecciones por Virus de Epstein-Barr , Neoplasias Nasofaríngeas , Línea Celular Tumoral , Infecciones por Virus de Epstein-Barr/genética , Infecciones por Virus de Epstein-Barr/patología , Factores de Crecimiento de Fibroblastos/metabolismo , Expresión Génica , Herpesvirus Humano 4/genética , Herpesvirus Humano 4/metabolismo , Humanos , Péptidos y Proteínas de Señalización Intracelular/genética , Proteínas de la Membrana/metabolismo , FN-kappa B/metabolismo , Carcinoma Nasofaríngeo/genética , Neoplasias Nasofaríngeas/genética , Neoplasias Nasofaríngeas/metabolismo , Neoplasias Nasofaríngeas/patología , Transducción de Señal , Microambiente TumoralRESUMEN
Epstein-Barr virus (EBV) is a ubiquitous oncovirus associated with specific epithelial and lymphoid cancers. Among the epithelial cancers, nasopharyngeal carcinoma (NPC), lymphoepithelioma-like carcinoma (LELC), and EBV-associated gastric cancers (EBVaGC) are the most common. The role of EBV in the pathogenesis of NPC and in the modulation of its tumour immune microenvironment (TIME) has been increasingly well described. Much less is known about the pathogenesis and tumour-microenvironment interactions in other EBV-associated epithelial cancers. Despite the expression of EBV-related viral oncoproteins and a generally immune-inflamed cancer subtype, EBV-associated epithelial cancers have limited systemic therapeutic options beyond conventional chemotherapy. Immune checkpoint inhibitors are effective only in a minority of these patients and even less efficacious with molecular targeting drugs. Here, we examine the key similarities and differences of NPC, LELC, and EBVaGC and comprehensively describe the clinical, pathological, and molecular characteristics of these cancers. A deeper comparative understanding of these EBV-driven cancers can potentially uncover targets in the tumour, TIME, and stroma, which may guide future drug development and cast light on resistance to immunotherapy.
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Infecciones por Virus de Epstein-Barr/complicaciones , Infecciones por Virus de Epstein-Barr/inmunología , Herpesvirus Humano 4/inmunología , Enfermedad de Hodgkin/complicaciones , Enfermedad de Hodgkin/inmunología , Carcinoma Nasofaríngeo/complicaciones , Carcinoma Nasofaríngeo/inmunología , Neoplasias Nasofaríngeas/complicaciones , Neoplasias Nasofaríngeas/inmunología , Neoplasias Gástricas/complicaciones , Neoplasias Gástricas/inmunología , Adolescente , Adulto , Animales , Infecciones por Virus de Epstein-Barr/virología , Femenino , Enfermedad de Hodgkin/virología , Humanos , Masculino , Carcinoma Nasofaríngeo/virología , Neoplasias Nasofaríngeas/virología , Factores Sexuales , Neoplasias Gástricas/virología , Microambiente Tumoral/inmunologíaRESUMEN
Natural killer (NK) cells comprise one subset of the innate lymphoid cell (ILC) family. Despite reported antitumor functions of NK cells, their tangible contribution to tumor control in humans remains controversial. This is due to incomplete understanding of the NK cell states within the tumor microenvironment (TME). Here, we demonstrate that peripheral circulating NK cells differentiate down two divergent pathways within the TME, resulting in different end states. One resembles intraepithelial ILC1s (ieILC1) and possesses potent in vivo antitumor activity. The other expresses genes associated with immune hyporesponsiveness and has poor antitumor functional capacity. Interleukin-15 (IL-15) and direct contact between the tumor cells and NK cells are required for the differentiation into CD49a+CD103+ cells, resembling ieILC1s. These data explain the similarity between ieILC1s and tissue-resident NK cells, provide insight into the origin of ieILC1s, and identify the ieILC1-like cell state within the TME to be the NK cell phenotype with the greatest antitumor activity. Because the proportions of the different ILC states vary between tumors, these findings provide a resource for the clinical study of innate immune responses against tumors and the design of novel therapy.
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Neoplasias de Cabeza y Cuello/inmunología , Inmunidad Innata/inmunología , Células Asesinas Naturales/inmunología , Linfocitos/inmunología , Microambiente Tumoral/inmunología , Anciano , Anciano de 80 o más Años , Antígenos CD/metabolismo , Antineoplásicos/metabolismo , Diferenciación Celular/inmunología , Línea Celular Tumoral , Femenino , Neoplasias de Cabeza y Cuello/patología , Humanos , Interleucina-15/metabolismo , Activación de Linfocitos/inmunología , Masculino , Persona de Mediana Edad , Miembro 1 del Grupo A de la Subfamilia 4 de Receptores Nucleares , Fenotipo , Carcinoma de Células Escamosas de Cabeza y Cuello/inmunología , Carcinoma de Células Escamosas de Cabeza y Cuello/patologíaRESUMEN
Rapid diagnostics of adventitious agents in biopharmaceutical/cell manufacturing release testing and the fight against viral infection have become critical. Quantitative real-time PCR and CRISPR-based methods rapidly detect DNA/RNA in 1 h but suffer from inter-site variability. Absolute quantification of DNA/RNA by methods such as digital PCR reduce this variability but are currently too slow for wider application. Here, we report a RApid DIgital Crispr Approach (RADICA) for absolute quantification of nucleic acids in 40-60 min. Using SARS-CoV-2 as a proof-of-concept target, RADICA allows for absolute quantification with a linear dynamic range of 0.6-2027 copies/µL (R2 value > 0.99), high accuracy and low variability, no cross-reactivity to similar targets, and high tolerance to human background DNA. RADICA's versatility is validated against other targets such as Epstein-Barr virus (EBV) from human B cells and patients' serum. RADICA can accurately detect and absolutely quantify EBV DNA with similar dynamic range of 0.5-2100 copies/µL (R2 value > 0.98) in 1 h without thermal cycling, providing a 4-fold faster alternative to digital PCR-based detection. RADICA therefore enables rapid and sensitive absolute quantification of nucleic acids which can be widely applied across clinical, research, and biomanufacturing areas.
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COVID-19 , Infecciones por Virus de Epstein-Barr , Ácidos Nucleicos , Repeticiones Palindrómicas Cortas Agrupadas y Regularmente Espaciadas , Herpesvirus Humano 4/genética , Humanos , SARS-CoV-2 , Sensibilidad y EspecificidadRESUMEN
Importance: Three-dimensionally printed nasopharyngeal swabs (3DP swabs) have been used to mitigate swab shortages during the coronavirus disease 2019 (COVID-19) pandemic. Clinical validation for diagnostic accuracy and consistency, as well as patient acceptability, is crucial to evaluate the swab's performance. Objective: To determine the accuracy and acceptability of the 3DP swab for identifying severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Design, Setting, and Participants: A diagnostic study was conducted from May to July 2020 at 2 tertiary care centers in Singapore with different reference swabs (FLOQSwab [COPAN Diagnostics] or Dacron swab [Deltalab]) and swab processing techniques (wet or dry) to evaluate the performance of the 3DP swab compared with traditional, standard-of-care nasopharyngeal swabs used in health care institutions. The participants were patients with COVID-19 in the first 2 weeks of illness and controls with acute respiratory illness with negative test results for SARS-CoV-2. Paired nasopharyngeal swabs were obtained from the same nostril and tested for SARS-CoV-2 by reverse-transcriptase polymerase chain reaction. The sequence of swabs was randomized based on odd and even participant numbers. Main Outcomes and Measures: Primary outcome measures were overall agreement (OA), positive percentage agreement (PPA), and negative percentage agreement of the 3DP swab compared with reference swabs. Secondary outcome measures were the correlation of cycle threshold (Ct) values of both swabs. Results: The mean (SD) age of participants was 45.4 (13.1) years, and most participants were men (87 of 89 [97.8%]), in keeping with the epidemiology of the COVID-19 pandemic in Singapore. A total of 79 patients with COVID-19 and 10 controls were recruited. Among the patients with COVID-19, the overall agreement and PPA of the 3DP swab was 91.1% and 93.5%, respectively, compared with reference swabs. The PPA was 100% for patients with COVID-19 who were tested within the first week of illness. All controls tested negative. The reverse-transcriptase polymerase chain reaction Ct values for the ORF1ab and E-gene targets showed a strong correlation (intraclass correlations coefficient, 0.869-0.920) between the 3DP and reference swab on independent testing at each institution despite differences in sample processing. Discordant results for both gene targets were observed only at high Ct values. Conclusions and Relevance: In this diagnostic study of 79 patients with COVID-19 and 10 controls, the 3DP swab performed accurately and consistently across health care institutions and could help mitigate strained resources in the escalating COVID-19 pandemic.
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Prueba de Ácido Nucleico para COVID-19/instrumentación , COVID-19/diagnóstico , Nasofaringe/virología , Impresión Tridimensional , Adulto , Diseño de Equipo , Humanos , Persona de Mediana Edad , Pandemias , SARS-CoV-2RESUMEN
INTRODUCTION: The gold standard for COVID-19 diagnosis is currently a real-time reverse transcriptase polymerase chain reaction (RT-PCR) to detect SARS-CoV-2. This is most commonly performed on respiratory secretions obtained via a nasopharyngeal swab. Due to supply chain limitations and high demand worldwide because of the COVID-19 pandemic, access to commercial nasopharyngeal swabs has not been assured. 3D printing methods have been used to meet the shortfall. For longer-term considerations, 3D printing may not compare well with injection molding as a production method due to the challenging scalability and greater production costs of 3D printing. METHODS: To secure sufficient nasopharyngeal swab availability for our national healthcare system, we designed a novel injection molded nasopharyngeal swab (the IM2 swab). We performed a clinical diagnostic study comparing the IM2 swab to the Copan FLOQSwab. Forty patients with a known diagnosis of COVID-19 and 10 healthy controls were recruited. Paired nasopharyngeal swabs were obtained from the same nostril of each participant and tested for SARS-CoV-2 by RT-PCR. RESULTS: When compared to the Copan FLOQswab, results from the IM2 swab displayed excellent overall agreement and positive percent agreement of 96.0% and 94.9%, respectively. There was no significant difference in mean RT-PCR cycle threshold values for the ORF1ab (28.05 vs. 28.03, p = 0.97) and E-gene (29.72 vs. 29.37, p = 0.64) targets, respectively. We did not observe any significant adverse events and there was no significant difference in patient-reported pain. CONCLUSION: In summary, the IM2 nasopharyngeal swab is a clinically safe, highly accurate option to commercial nasopharyngeal swabs.
RESUMEN
OBJECTIVE: The coronavirus disease 2019 (COVID-19) pandemic has led to a global surge in critically ill patients requiring invasive mechanical ventilation, some of whom may benefit from tracheostomy. Decisions on if, when, and how to perform tracheostomy in patients with COVID-19 have major implications for patients, clinicians, and hospitals. We investigated the tracheostomy protocols and practices that institutions around the world have put into place in response to the COVID-19 pandemic. DATA SOURCES: Protocols for tracheostomy in patients with severe acute respiratory syndrome coronavirus 2 infection from individual institutions (n = 59) were obtained from the United States and 25 other countries, including data from several low- and middle-income countries, 23 published or society-endorsed protocols, and 36 institutional protocols. REVIEW METHODS: The comparative document analysis involved cross-sectional review of institutional protocols and practices. Data sources were analyzed for timing of tracheostomy, contraindications, preoperative testing, personal protective equipment (PPE), surgical technique, and postoperative management. CONCLUSIONS: Timing of tracheostomy varied from 3 to >21 days, with over 90% of protocols recommending 14 days of intubation prior to tracheostomy. Most protocols advocate delaying tracheostomy until COVID-19 testing was negative. All protocols involved use of N95 or higher PPE. Both open and percutaneous techniques were reported. Timing of tracheostomy changes ranged from 5 to >30 days postoperatively, sometimes contingent on negative COVID-19 test results. IMPLICATIONS FOR PRACTICE: Wide variation exists in tracheostomy protocols, reflecting geographical variation, different resource constraints, and limited data to drive evidence-based care standards. Findings presented herein may provide reference points and a framework for evolving care standards.