Chromatin accessibility dynamics in colorectal cancer liver metastasis: Uncovering the liver tropism at single cell resolution.

Tumor metastasis causes over 90% of cancer related death and no currently available therapies target it. However, there is limited understanding regarding the epigenetic regulation of genes during this complex process. Here by integrating single-cell ATAC-seq (scATAC-seq), single-cell RNA-seq (scRNA-seq), microarray, bulk RNA-seq, immunohistochemistry (IHC) staining, as well as proteomics datasets from paired primary and liver metastatic colorectal cancer (CRC) patient-derived xenograft (PDX) model and patients, we discovered that liver metastatic CRC cells lose their colon-specific chromatin accessible sites yet gain liver-specific ones. Importantly, we observed elevated accessibility of HNF4A, a liver-specific transcription factor, in liver metastatic CRC cells. Subsequently, we performed clustering analysis of liver metastatic CRC cells together with cells involved in liver development, revealing significant heterogeneity among the liver metastatic CRC cells. Over 50% of the liver metastatic CRC cells exhibited characteristics similar to those of erythroid progenitors and hepatocytes, showing increased expression of genes involved in oxidative phosphorylation and glycolysis. Moreover, our discovery further revealed that the MHC and IFN response genes in these cells exhibit moderate epigenetic activity, which is significantly associated with the low objective response rates in checkpoint blockade immunotherapy. Our findings uncovered the critical roles of HNF4A and the cell populations within liver metastatic CRC cells might serve as crucial therapeutic targets for addressing liver metastasis and improving the immunotherapy response in patients with CRC.

Long non-coding RNA DARS-AS1 promotes tumor progression by directly suppressing PACT-mediated cellular stress.

Cancer cells evolve various mechanisms to overcome cellular stresses and maintain progression. Protein kinase R (PKR) and its protein activator (PACT) are the initial responders in monitoring diverse stress signals and lead to inhibition of cell proliferation and cell apoptosis in consequence. However, the regulation of PACT-PKR pathway in cancer cells remains largely unknown. Herein, we identify that the long non-coding RNA (lncRNA) aspartyl-tRNA synthetase antisense RNA 1 (DARS-AS1) is directly involved in the inhibition of the PACT-PKR pathway and promotes the proliferation of cancer cells. Using large-scale CRISPRi functional screening of 971 cancer-associated lncRNAs, we find that DARS-AS1 is associated with significantly enhanced proliferation of cancer cells. Accordingly, knocking down DARS-AS1 inhibits cell proliferation of multiple cancer cell lines and promotes cancer cell apoptosis in vitro and significantly reduces tumor growth in vivo. Mechanistically, DARS-AS1 directly binds to the activator domain of PACT and prevents PACT-PKR interaction, thereby decreasing PKR activation, eIF2α phosphorylation and inhibiting apoptotic cell death. Clinically, DARS-AS1 is broadly expressed across multiple cancers and the increased expression of this lncRNA indicates poor prognosis. This study elucidates the lncRNA DARS-AS1 directed cancer-specific modulation of the PACT-PKR pathway and provides another target for cancer prognosis and therapeutic treatment.

Tissue-specific transcription reprogramming promotes liver metastasis of colorectal cancer.

Metastasis, the development of secondary malignant growths at a distance from a primary tumor, is the cause of death for 90% of cancer patients, but little is known about how metastatic cancer cells adapt to and colonize new tissue environments. Here, using clinical samples, patient-derived xenograft (PDX) samples, PDX cells, and primary/metastatic cell lines, we discovered that liver metastatic colorectal cancer (CRC) cells lose their colon-specific gene transcription program yet gain a liver-specific gene transcription program. We showed that this transcription reprogramming is driven by a reshaped epigenetic landscape of both typical enhancers and super-enhancers. Further, we identified that the liver-specific transcription factors FOXA2 and HNF1A can bind to the gained enhancers and activate the liver-specific gene transcription, thereby driving CRC liver metastasis. Importantly, similar transcription reprogramming can be observed in multiple cancer types. Our data suggest that reprogrammed tissue-specific transcription promotes metastasis and should be targeted therapeutically.

A comprehensive evaluation of connectivity methods for L1000 data.

The methodologies for evaluating similarities between gene expression profiles of different perturbagens are the key to understanding mechanisms of actions (MoAs) of unknown compounds and finding new indications for existing drugs. L1000-based next-generation Connectivity Map (CMap) data is more than a thousand-fold scale-up of the CMap pilot dataset. Although several systematic evaluations have been performed individually to assess the accuracy of the methodologies for the CMap pilot study, the performance of these methodologies needs to be re-evaluated for the L1000 data. Here, using the drug-drug similarities from the Drug Repurposing Hub database as a benchmark standard, we evaluated six popular published methods for the prediction performance of drug-drug relationships based on the partial area under the receiver operating characteristic (ROC) curve at false positive rates of 0.001, 0.005 and 0.01 (AUC0.001, AUC0.005 and AUC0.01). The similarity evaluating algorithm called ZhangScore was generally superior to other methods and exhibited the highest accuracy at the gene signature sizes ranging from 10 to 200. Further, we tested these methods with an experimentally derived gene signature related to estrogen in breast cancer cells, and the results confirmed that ZhangScore was more accurate than other methods. Moreover, based on scoring results of ZhangScore for the gene signature of TOP2A knockdown, in addition to well-known TOP2A inhibitors, we identified a number of potential inhibitors and at least two of them were the subject of previous investigation. Our studies provide potential guidelines for researchers to choose the suitable connectivity method. The six connectivity methods used in this report have been implemented in R package (https://github.com/Jasonlinchina/RCSM).

Microarray is an efficient tool for circRNA profiling.

Circular RNAs (circRNAs) are emerging as a new class of endogenous and regulatory noncoding RNAs in latest years. With the widespread application of RNA sequencing (RNA-seq) technology and bioinformatics prediction, large numbers of circRNAs have been identified. However, at present, we lack a comprehensive characterization of all these circRNAs in interested samples. In this study, we integrated 87 935 circRNAs sequences that cover most of circRNAs identified till now represented in circBase to design microarray probes targeting back-splice site of each circRNA to profile expression of those circRNAs. By comparing the circRNA detection efficiency of RNA-seq with this circRNA microarray, we revealed that microarray is more efficient than RNA-seq for circRNA profiling. Then, we found ∼80 000 circRNAs were expressed in cervical tumors and matched normal tissues, and ∼25 000 of them were differently expressed. Notably, many of these circRNAs detected by this microarray can be validated by quantitative reverse transcription polymerase chain reaction (RT-qPCR) or RNA-seq. Strikingly, as many as ∼18 000 circRNAs could be robustly detected in cell-free plasma samples, and the expression of ∼2700 of them differed after surgery for tumor removal. Our findings provided a comprehensive and genome-wide characterization of circRNAs in paired normal tissues and tumors and plasma samples from multiple individuals. In addition, we also provide a rich resource with 41 microarray data sets and 10 RNA-seq data sets and strong evidences for circRNA expression in cervical cancer. In conclusion, circRNAs could be efficiently profiled by circRNA microarray to target their reported back-splice sites in interested samples.