Combining Transoral Nasopharyngeal Brush and Plasma Epstein-Barr Virus DNA in Detecting Locally Recurrent Nasopharyngeal Carcinoma.

OBJECTIVE: To evaluate the sensitivities and specificities of Epstein-Barr virus (EBV) DNA in the detection of locally recurrent or persistent nasopharyngeal carcinoma (NPC) through nasopharyngeal (NP) brush biopsy and plasma, respectively, and whether a combination of both would be superior to the individual tests. STUDY DESIGN: A case-control study was conducted from September 2016 to June 2022. SETTING: A multicentre study at 3 tertiary referral centers in Hong Kong was conducted by the Department of Otorhinolaryngology, Head and Neck Surgery, The Chinese University of Hong Kong. METHODS: Twenty-seven patients with biopsy-confirmed locally recurrent NPC were recruited as study subjects. Magnetic resonance imaging was performed to rule out regional recurrence. The control group consisted of 58 patients with a prior history of NPC who were now disease-free based on endoscopic and imaging findings. Patients underwent both the transoral NP brush (NP Screen®) and blood for plasma Epstein-Barr DNA levels. RESULTS: The sensitivity and specificity of the combined modalities were 84.62% and 85.19%, respectively. The positive predictive value was 73.33% and the negative predictive value was 92.0%. CONCLUSION: The combination of NP brush biopsy and plasma EBV DNA is potentially an additional surveillance modality in detecting the local recurrence of NPC. Further study with a larger sample size would be required to validate the cutoff values.

Nasopharyngeal carcinoma: an evolving paradigm.

The past three decades have borne witness to many advances in the understanding of the molecular biology and treatment of nasopharyngeal carcinoma (NPC), an Epstein-Barr virus (EBV)-associated cancer endemic to southern China, southeast Asia and north Africa. In this Review, we provide a comprehensive, interdisciplinary overview of key research findings regarding NPC pathogenesis, treatment, screening and biomarker development. We describe how technological advances have led to the advent of proton therapy and other contemporary radiotherapy approaches, and emphasize the relentless efforts to identify the optimal sequencing of chemotherapy with radiotherapy through decades of clinical trials. Basic research into the pathogenic role of EBV and the genomic, epigenomic and immune landscape of NPC has laid the foundations of translational research. The latter, in turn, has led to the development of new biomarkers and therapeutic targets and of improved approaches for individualizing immunotherapy and targeted therapies for patients with NPC. We provide historical context to illustrate the effect of these advances on treatment outcomes at present. We describe current preclinical and clinical challenges and controversies in the hope of providing insights for future investigation.

Droplet digital PCR of tumor suppressor gene methylation in serial oral rinses of patients with head and neck squamous cell carcinoma.

BACKGROUND: Head and neck squamous cell carcinoma (HNSCC) currently lacks sensitive approaches to detect cancer-related traits in body fluid. METHODS: Methylation of tumor suppressor genes (TSGs) (PAX5, EDNRB, and DCC) were measured in the oral rinses from 50 HNSCC and 58 control subjects using droplet digital PCR (ddPCR). Diagnostic accuracies in detecting HNSCC and the detection rate of recurrence in the post-treatment monitoring were analyzed. RESULTS: ddPCR TSG methylation detection in oral rinses for diagnosis of HNSCC had an AUC of 0.892 for PAX5, 0.753 for EDNRB, and 0.729 for DCC. Significant drop of TSG methylation was observed after completion of surgery (p < 0.01). 76.9% of the relapse cases had a pre-emptive rebound of methylation above presurgery levels in at least one of the tested markers before confirmed recurrence. CONCLUSIONS: Utilizing ddPCR for TSG methylation detection in oral rinses shows potential for detection and monitoring of HNSCC.

Development of a Novel Inflammation-Based Index for Hepatocellular Carcinoma.

BACKGROUND: The aim of current study was to (1) construct and validate a novel hepatocellular carcinoma (HCC)-specific inflammatory index; (2) compare the performances of the Integrated Liver Inflammatory Score (ILIS) to existing 4 inflammatory indices in HCC; (3) explore the association between the inflammatory indices and systemic/intratumoral inflammatory markers. METHODS: Two cohorts from Hong Kong (HK; n = 1,315) and Newcastle (n = 574) were studied. A novel index was constructed from the HK training set (n = 627). The index was constructed from the training set by combing independent prognostic circulating parameters, followed by validating in the validation set of HK cohort (n = 688) and the Newcastle cohort. Its prognostic performance was compared to 4 inflammatory indices, namely, the neutrophil to lymphocyte ratio, platelet-to-lymphocyte ratio, prognostic nutrition index, and systemic immune-inflammation index, were compared in the HK cohort. Circulating cytokines and intratumoral gene expression were analyzed in a subset of patients with available samples and correlated with the inflammatory indices. RESULTS: In the training set of the HK cohort, the ILIS, was generated: -0.057 × albumin (g/L) + 0.978 × log (Bilirubin, µmol/L) + 1.341 × log (alkaline phosphatase, IU/L) + 0.086 × Neutrophil (109/L) + 0.301 × log (alpha-fetoprotein, µg/L). With cutoff of 2.60 and 3.87, the ILIS could categorize patients into 3 risk groups in the both validation cohorts. ILIS outperforms other inflammatory indices and remains an independent prognosticator for overall survival after adjustment with Barcelona Clinic Liver Cancer (hazard ratio 31.90, p < 0.001). The ILIS had the best prognostic performances as compared to other inflammatory indices. In exploratory analyses, the ILIS correlated with circulating inflammatory cytokines (e.g., IL-8) but not with any intratumoral inflammatory gene expression. CONCLUSIONS: ILIS is an HCC-specific prognostic index built on 5 readily available blood parameters. Its versatility is validated both Eastern and Western population of HCC. The score is correlated with levels of circulating cytokines.

Plasma Epstein-Barr virus DNA as an archetypal circulating tumour DNA marker.

Analysis of circulating tumour DNA (ctDNA), as one type of 'liquid biopsy', has recently attracted great attention. Researchers are exploring many potential applications of liquid biopsy in many different types of cancer. In particular, it is of biological interest and clinical relevance to study the molecular characteristics of ctDNA. For such purposes, plasma Epstein-Barr virus (EBV) DNA from patients with nasopharyngeal carcinoma (NPC) would provide a good model to understand the biological properties and clinical applications of ctDNA in general. The strong association between EBV and NPC in endemic regions has made plasma EBV DNA a robust biomarker for this cancer. There are many clinical utilities of plasma EBV DNA analysis in NPC diagnostics. Its role in prognostication and surveillance of recurrence is well established. Plasma EBV DNA has also been validated for screening NPC in a recent large-scale prospective study. Indeed, plasma EBV DNA could be regarded as an archetypal ctDNA marker. In this review, we discuss the biological properties of plasma EBV DNA from NPC samples and also the clinical applications of plasma EBV DNA analysis in the management of NPC. Of note, the recently reported size analysis of plasma EBV DNA in patients with NPC has highlighted size as an important analytical parameter of ctDNA and demonstrated clinical value in improving the diagnostic performance of an EBV DNA-based NPC screening test. Such insights into ctDNA analysis (including size profiling) may help its full potential in cancer diagnostics for other types of cancer to be realised. © 2019 The Authors. The Journal of Pathology published by John Wiley & Sons Ltd on behalf of Pathological Society of Great Britain and Ireland.

Clinical utility of circulating Epstein-Barr virus DNA analysis for the management of nasopharyngeal carcinoma.

Nasopharyngeal carcinoma (NPC) is one of the commonest cancers in Southern China. The carcinogenesis is closely associated with Epstein-Barr virus (EBV) infection. In endemic regions with high incidence of NPC, EBV genome can be detected in virtually all NPC tumor tissues. Over the last decade, circulating cell-free EBV DNA has been developed as a tumor marker for NPC. Plasma EBV DNA analysis using real-time PCR has been shown to be useful for early detection, prognostication and monitoring of treatment response of NPC. In this review, the clinical applications of EBV DNA analysis in the management of NPC would be discussed.

FetalQuantSD: accurate quantification of fetal DNA fraction by shallow-depth sequencing of maternal plasma DNA.

Noninvasive prenatal testing using massively parallel sequencing of maternal plasma DNA has been rapidly adopted in clinical use worldwide. Fetal DNA fraction in a maternal plasma sample is an important parameter for accurate interpretations of these tests. However, there is a lack of methods involving low-sequencing depth and yet would allow a robust and accurate determination of fetal DNA fraction in maternal plasma for all pregnancies. In this study, we have developed a new method to accurately quantify the fetal DNA fraction by analysing the maternal genotypes and sequencing data of maternal plasma DNA. Fetal DNA fraction was calculated based on the proportion of non-maternal alleles at single-nucleotide polymorphisms where the mother is homozygous. This new approach achieves a median deviation of 0.6% between predicted fetal DNA fraction and the actual fetal DNA fraction using as low as 0.03-fold sequencing coverage of the human genome. We believe that this method will further enhance the clinical interpretations of noninvasive prenatal testing using genome-wide random sequencing.

Noninvasive prenatal molecular karyotyping from maternal plasma.

Fetal DNA is present in the plasma of pregnant women. Massively parallel sequencing of maternal plasma DNA has been used to detect fetal trisomies 21, 18, 13 and selected sex chromosomal aneuploidies noninvasively. Case reports describing the detection of fetal microdeletions from maternal plasma using massively parallel sequencing have been reported. However, these previous reports were either polymorphism-dependent or used statistical analyses which were confined to one or a small number of selected parts of the genome. In this report, we reported a procedure for performing noninvasive prenatal karyotyping at 3 Mb resolution across the whole genome through the massively parallel sequencing of maternal plasma DNA. This method has been used to analyze the plasma obtained from 6 cases. In three cases, fetal microdeletions have been detected successfully from maternal plasma. In two cases, fetal microduplications have been detected successfully from maternal plasma. In the remaining case, the plasma DNA sequencing result was consistent with the pregnant mother being a carrier of a microduplication. Simulation analyses were performed for determining the number of plasma DNA molecules that would need to be sequenced and aligned for enhancing the diagnostic resolution of noninvasive prenatal karyotyping to 2 Mb and 1 Mb. In conclusion, noninvasive prenatal molecular karyotyping from maternal plasma by massively parallel sequencing is feasible and would enhance the diagnostic spectrum of noninvasive prenatal testing.

Noninvasive fetal trisomy 21 detection using chromosome-selective sequencing: a variation of the molecular counting theme.

Fetal DNA is present in the plasma of pregnant women. A fetus with trisomy of a chromosome will release an increased amount of DNA from that chromosome into maternal plasma. Such an increase has previously been measured using methods that allow individual DNA molecules to be counted. One such method involves the use of random massively parallel sequencing of maternal plasma DNA. As the sequencing process is random, sequence tags from a potentially aneuploid chromosome only represent a fraction of the sequencing data. The performance of selective amplification and sequencing of specific genomic regions is a recently reported approach for focusing the sequencing power onto genomic regions of diagnostic interest. This article provides a critical analysis of this approach and puts this method in the perspective of other recent works in the field.

Risk of vitamin A toxicity from candy-like chewable vitamin supplements for children.

Numerous vitamin supplements are available over-the-counter to the general public. Some such supplements are available as candy-like chewable preparations to encourage consumption by children. We report 3 cases of overdose of such preparations. Each patient had taken an estimated 200,000 to 300,000 IU of vitamin A. Their circulating vitamin A (retinol and retinyl palmitate) concentrations were monitored over a 6-month period. There were no clinical or biochemical complications noted. However, there were marked increases in both retinol and retinyl palmitate concentrations above age-related reference ranges. In particular, it took 1 to 3 weeks for the serum retinol concentrations to peak and many months for them to normalize. Parents should be warned about the dangers of excessive vitamin consumption. Clinicians should be aware of the late peak in serum retinol concentrations, which may lead to late complications of vitamin A overdose.