Cell-free circulating tumor RNAs in plasma as the potential prognostic biomarkers in colorectal cancer.

Background: Cell free RNA (cfRNA) contains transcript fragments from multiple cell types, making it useful for cancer detection in clinical settings. However, the pathophysiological origins of cfRNAs in plasma from colorectal cancer (CRC) patients remain unclear. Methods: To identify the tissue-specific contributions of cfRNAs transcriptomic profile, we used a published single-cell transcriptomics profile to deconvolute cell type abundance among paired plasma samples from CRC patients who underwent tumor-ablative surgery. We further validated the differentially expressed cfRNAs in 5 pairs of CRC tumor samples and adjacent tissue samples as well as 3 additional CRC tumor samples using RNA-sequencing. Results: The transcriptomic component from intestinal secretory cells was significantly decreased in the in-house post-surgical cfRNA. The HPGD, PACS1, and TDP2 expression was consistent across cfRNA and tissue samples. Using the Cancer Genome Atlas (TCGA) CRC datasets, we were able to classify the patients into two groups with significantly different survival outcomes. Conclusions: The three-gene signature holds promise in applying minimal residual disease (MRD) testing, which involves profiling remnants of cancer cells after or during treatment. Biomarkers identified in the present study need to be validated in a larger cohort of samples in order to ascertain their possible use in early diagnosis of CRC.

The diagnostic significance of CDH17-positive circulating tumor cells in patients with colorectal cancer.

BACKGROUND: Colorectal cancer (CRC) is the second leading cause of cancer deaths in Hong Kong. We tested the hypothesis that circulating tumor cell (CTC) analysis by ARB101 antibody could be used as a tool for CRC detection, progression, and therapy response. RESEARCH METHODS: ARB101 antibody was used for investigation of CDH17 expression in formalin-fixed, paraffin-embedded (FFPE) tissue sections and circulating tumor cells (CTCs) of CRC patients. RESULTS: Using ARB101, highest sensitivity was observed in 98/100 (98%) colorectal cancer tissue compared to 72/100 gastric cancer (72%) and 27/32 pancreatic cancer (84%). Immunoreactivity of CDH17 was significantly higher in distant metastatic (tumor-node-metastasis [TNM] stage IV) than non-distant metastatic (TNM stage I to III) CRC. ARB101 antibody also manifested the higher sensitivity than c-erbB2 (8%) and epidermal growth factor receptor (EGFR)-targeting antibodies (37%) with the significance (p < 0.0001). ARB101 positive CTCs were detected in 64/83 (77%) TNM stage I to IV CRC patients. Furthermore, ARB101 positive CTCs detected in TNM stage I to III CRC patients before and after surgical operation are statistically significant (p < 0.0001). CONCLUSIONS: CTC detection by ARB101 antibody could serve as a potential non-invasive approach for CRC detection, progression, and monitoring of treatment response.

The non-invasive diagnosis of colorectal cancer via a SOX9-based gene panel.

Colorectal cancer (CRC) threatens human health seriously. Early diagnosis of CRC is critical to improving patient survival. Meanwhile, non-invasive detection through tumor-circulating markers can be an important auxiliary diagnosis. In this study, we performed targeted RNA sequencing in paired tumor and adjacent normal fresh frozen tissues from 68 patients, and we also measured circulating mRNA levels in 4 time-point plasma samples collected before and after operation or chemotherapy. Our results showed that SOX9 (6.73-fold with adjusted p value < 1 × 10-45), MYC (20.59-fold with adjusted p value < 1 × 10-57), and MMP7 (131.94-fold with adjusted p value < 1 × 10-78) highly expressed in tumor compared with adjacent normal tissues. Besides, the circulating mRNA of SOX9 (41.14-fold with adjusted p value < 1 × 10-13) in CRC was significantly higher than in the normal control as well. Moreover, a SOX9-based 9-gene panel (SOX9, GSK3A, FZD4, LEF1, DVL1, FZD7, NFATC1, KRT19, and RUVBL1) showed the non-invasive diagnostic value of CRC (AUC: 0.863 (0.766-0.960), TPR: 0.92, TNR: 0.87). In summary, SOX9 expression consistently increases in tumor and plasma samples from CRC patients, which indicates the important role of SOX9 in CRC progression and its potential in non-invasive diagnosis of CRC.

The current and future applications of in situ hybridization technologies in anatomical pathology.

INTRODUCTION: In situ hybridization (ISH) plays an important role in the field of molecular diagnostics, especially in an anatomical pathology laboratory. ISH is a technique that can detect the targeted DNA or RNA sequences in tissue sections from frozen or fixed materials with labeled DNA or RNA probes. Radioactive and non-radioactive probes are the two major probes that can be used to label the targeted nucleic acids. AREAS COVERED: Two decades after the Human Genome Project, ISH has not only simply been applied to identify the chromosomal location of a human gene but has also been extensively applied to gene expressions studies and utilized for clinical diagnosis, especially for the determination of biomarkers for breast and ovarian cancers - human epidermal growth factor receptor 2. Duchenne muscular dystrophy, Cri-du-chat syndrome, Angelman syndrome, PraderWilli syndrome, cystic fibrosis, and trisomy are diseases that can also be detected by ISH. In this review, the basic principles, historical development, advantages and disadvantages, enhancement in reporting molecules and probes, advancement in detection methods, in situ PCR, clinical applications and novel applications of ISH will be discussed. EXPERT OPINION: With the advancement in ISH technologies and appropriate training, diagnosis can be improved in Anatomical Pathology.

Whole genome amplicon sequencing and phylogenetic analysis of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) from lineage B.1.36.27 isolated in Hong Kong.

BACKGROUND: The import of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) lineage B.1.36.27 has sparked the fourth wave of COVID-19 outbreak in Hong Kong. This strain has been circulating in Hong Kong since September 2020 but rarely found in other countries (<1%). RESEARCH DESIGN AND METHODS: A total of 14 SARS-CoV-2 genome sequences collected from patients in Hong Kong between July 2020 and March 2021 were determined by whole viral genome sequencing using Illumina next-generation sequencing platform, followed by phylogenetic analysis. RESULTS: Of the 14 SARS-CoV-2 genome sequences analyzed, 9 strains belonged to the PANGO lineage B.1.36.27, GISAID clade GH, and Nextclade clade 20A. Compared to the reference genome, 31 nucleotide differences and 11 amino acid differences were identified in the genome of the SARS-CoV-2 from PANGO lineage B.1.36.27. CONCLUSIONS: We reported the nucleotides and amino acids mutations identified in the SARS-CoV-2 from PANGO lineage B.1.36.27. Our viral genome sequences enriched the understanding of SARS-CoV-2 mutational landscape and improved the repertoire of known SARS-CoV-2 variants for tracking and tracing. From this study, we found no evidence to show that SARS-CoV-2 from lineage B.1.36.27 can compromise existing vaccines and antibody therapies.

Implementation and new insights in molecular diagnostics for HIV infection.

INTRODUCTION: Acquired immunodeficiency syndrome (AIDS) is a kind of acquired disease that breaks down the immune system. Human immunodeficiency virus (HIV) is the causative agent of AIDS. By the end of 2016, there were 36.7 million people living with HIV worldwide. Early diagnosis can alert infected individuals to risk behaviors in order to control HIV transmission. Infected individuals are also benefited from proper treatment and management upon early diagnosis. Thanks to the public awareness of the disease, the annual increase of new HIV infections has been slowly declining over the past decades. The advent of molecular diagnostics has allowed early detection and better management of HIV infected patients. Areas covered: In this review, the authors summarized and discussed the current and future technologies in molecular diagnosis as well as the biomarkers developed for HIV infection. Expert Commentary: A simple and rapid detection of viral load is important for patients and doctors to monitor HIV progression and antiretroviral treatment efficiency. In the near future, it is expected that new technologies such as digital PCR and CRISPR-based technology will play more important role in HIV detection and patient management.

Clinical significance of frizzled homolog 3 protein in colorectal cancer patients.

Frizzled homolog 3 receptor was up-regulated in several gastrointestinal cancers such as esophageal and gastric cancers. Moreover, frizzled homolog 3 has recently reported to be expressed in colorectal adenoma specimens. In the present study, we investigated the clinical significance of frizzled homolog 3 protein in colorectal cancer patients. Using immunocytochemical staining, frizzled homolog 3 expression was examined in 186 colorectal cancer specimens, 79 colorectal adenoma specimens, 133 colorectal polyp specimens, 127 colorectal cancer specimens with lymph node and/or distant metastasis, 310 specimens of various non-colorectal cancer metastatic carcinomas and 40 specimens with simultaneous occurrence of colorectal cancer, colorectal adenoma and colorectal polyp. Statistical analysis was used to correlate frizzled homolog 3 protein expression to the clinicohistopathological factors, recurrence/metastasis and survival after follow-up for 42 months in colorectal cancer patients. Frizzled homolog 3 protein was expressed in 100% colorectal cancer specimens, 89% colorectal adenoma specimens, 75% colorectal polyp specimens and 69% normal colorectal epithelial tissues. Moreover, frizzled homolog 3 immunocytochemical scores were highly correlated with colorectal cancer progression. Furthermore, frizzled homolog 3 was expressed in a comparatively lower percentage of metastatic hepatocellular carcinoma and metastatic renal clear cell carcinoma with focal and very weak staining than other metastatic tumor types. On the other hand, the frizzled homolog 3 immunocytochemical scores of colorectal adenomas with synchronous colorectal carcinomas were significantly higher than those of pure colorectal adenomas. Statistical analysis showed that frizzled homolog 3 immunocytochemical scores were associated with Dukes stage and lymph node status. Finally, stratified groups of colorectal cancer patients had significant differences in their recurrence/metastasis and survival. In conclusion, the present large-scale study has clearly showed that frizzled homolog 3 protein can generate clinically important information for colorectal cancer patients.