James A. Radosevich

Objective: The discovery and characterization of tumor associated antigens is increasingly important to advance the field of immuno-oncology. In this regard, labyrinthin has been implicated as a neoantigen found on the cell surface of adenocarcinomas. Data on the (1) topology, (2) amino acid (a.a.) homology analyses and (3) cell surface localization of labyrinthin by fluorescent activated cell sorter (FACS) are studied in support of labyrinthin as a novel, pan-adenocarcinoma marker. Results: Bioinformatics analyses predict labyrinthin as a type II protein with calcium binding domain(s), N-myristoylation sites, and kinase II phosphorylation sites. Sequence homologies for labyrinthin (255 a.a.) were found vs. the intracellular aspartyl/asparaginyl beta-hydroxylase (ASPH; 758 a.a.) and the ASPH-gene related protein junctate (299 a.a.), which are both type II proteins. Labyrinthin was detected by FACS on only non-permeablized A549 human lung adenocarcinoma cells, but not on normal WI-38 human lung fibroblasts nor primary cultures of normal human glandular-related cells. Microscopic images of immunofluorescent labelled MCA 44-3A6 binding to A549 cells at random cell cycle stages complement the FACS results by further showing that labyrinthin persisted on the cell surfaces along with some cell internalization for greater than 20 min.

To determine Labyrinthin (LAB) expression in non-small-cell lung cancer (NSCLC), we immunostained and scored for LAB immunohistochemistry (IHC) expression on sections of tissue microarrays (TMAs) prepared from 256 archival tissue blocks of NSCLC. Propensity-score-weighted Kaplan–Meier curves and weighted Cox models were used to associate LAB expression with overall survival. LAB mRNA expression was assessed in The Cancer Genome Atlas (TCGA) and correlated with clinical phenotype and outcome. Positive LAB IHC expression (>5% of tumor cells) was detected in 208/256 (81.3%) of NSCLC samples, and found in both lung adenocarcinomas (LUAD) and lung squamous cell cancer (LUSC). LAB positivity was associated with poor overall survival (HR = 3.56, 95% CI: 2.3–5.4; p < 0.0001) and high tumor differentiation grade or metastasis compared with negative LAB expression. Univariant and multivariate survival analyses demonstrated LAB expression as an independent prognostic factor for NSCLC patients. LAB RNA expression in TCGA-LUAD was higher in primary and advanced-stage tumors than in normal tissue, and was associated with poorer overall survival. No significant differences or associations were found with LAB RNA expression in TCGA-LUSC. The LAB IHC assay is being used to identify candidate cancer patients for the first-in-human phase I trial evaluating the LAB vaccines (UCDCC#296, NCT051013560).

Autophagy is an evolutionarily conserved pathway that eliminates unwanted proteins out of the cell and increases cell survival. However, dysfunctional autophagy is associated with cancer progression, cellular adaptation, cancer metastasis and makes it an attractive therapeutic target. MicroRNAs (miRNAs) are small single-stranded non-coding RNA molecules that usually bind to 3’UTR of mRNAs. This interaction eventually inhibits protein synthesis by repressing translation and/or by degrading mRNAs. miRNAs play a crucial role in the regulation of autophagy and also behave as both tumor suppressors and promoters in colorectal cancer. This paper defines an overall molecular view of how miRNAs regulate the dual role of autophagy in colorectal cancer. It also highlights how long noncoding RNAs modulate miRNAs expression to regulate autophagy in colorectal cancer. Thus, targeting autophagy by miRNAs seems to be a potential therapeutic strategy for colorectal cancer.

Background: Nitric oxide (NO) exposure has been suggested to cause alterations in DNA methylation in breast cancer. We investigated the effect of NO on DNA methylation of promoters in cell lines of breast cancer. Material and Methods: The methylation status of the promoters of breast cancer 1 (BRCA1), deleted in colon cancer (DCC), Ras-association domain family 1A (RASSF1A), O⁶-methylguanine-DNA methyltransferase (MGMT), and secreted frizzled related protein 1 (SFRP1) were analyzed in the parental and high nitric oxide-adapted cell lines of breast cancer using Illumina MiSequencing. Results: Methylation of RASSF1A promoter in BT-20-HNO (74.7%) was significantly higher than that in BT-20 cells (72%) (p<0.05), whereas in MCF-7-HNO cells, methylation of MGMT promoter was found to have significantly decreased as compared to its parental cell line (45.1% versus 50.1%; p<0.0001). Promoter methylation of SFRP and DCC was elevated in T-47D-HNO relative to its parent cell line (p<0.05). Conclusion: Similarly to the double-edged effects of NO on tumorigenesis, its epigenetic effects through DNA methylation are diverse and contradictory in breast cancer.

A paradigm shift is currently underway on the relationship between cancer treatment markers and therapies. Labyrinthin is a prime example of such a marker because it is a pan-cancer target for adenocarcinomas. This movement supports the idea that we must change our thinking from various cancer types (eg, lung, breast, colon) to “cancer arising” in a given tissue or organ. In doing so, this would further support the efforts toward pan-treatments rather than organ-specific treatments.