The epithelial-to-mesenchymal transition (EMT) plays a critical role during normal development and in cancer progression. EMT is induced by various signaling pathways, including TGF-β, BMP, Wnt–β-catenin, NOTCH, Shh, and receptor tyrosine kinases. In this study, we performed single-cell RNA sequencing on MCF10A cells undergoing EMT by TGF-β1 stimulation. Our comprehensive analysis revealed that cells progress through EMT at different paces. Using pseudotime clustering reconstruction of gene-expression profiles during EMT, we found sequential and parallel activation of EMT signaling pathways. We also observed various transitional cellular states during EMT. We identified regulatory signaling nodes that drive EMT with the expression of important microRNAs and transcription factors. Using a random circuit perturbation methodology, we demonstrate that the NOTCH signaling pathway acts as a key driver of TGF-β–induced EMT. Furthermore, we demonstrate that the gene signatures of pseudotime clusters corresponding to the intermediate hybrid EMT state are associated with poor patient outcome. Overall, this study provides insight into context-specific drivers of cancer progression and highlights the complexities of the EMT process.

SUMMARY β-hydroxybutyrate (β-OHB) is an essential metabolic energy source during fasting and functions as a chromatin regulator by lysine β-hydroxybutyrylation (Kbhb) modification of the core histones H3 and H4. We report that Kbhb on histone H3 (H3K9bhb) is enriched at proximal promoters of critical gene subsets associated with lipolytic and ketogenic metabolic pathways in small intestine (SI) crypts during fasting. Similar Kbhb enrichment is observed in Lgr5 + stem cell-enriched epithelial spheroids treated with β-OHB in vitro . Combinatorial chromatin state analysis reveals that H3K9bhb is associated with active chromatin states and that fasting enriches for an H3K9bhb-H3K27ac signature at active metabolic gene promoters and distal enhancer elements. Intestinal knockout of Hmgcs2 results in marked loss of H3K9bhb-associated loci, suggesting that local production of β-OHB is responsible for chromatin reprogramming within the SI crypt. We conclude that modulation of H3K9bhb in SI crypts is a key gene regulatory event in response to fasting.

ObjectiveEnhancer aberrations are beginning to emerge as a key epigenetic feature of colorectal cancers (CRC), however, a comprehensive knowledge of chromatin state patterns in tumour progression, heterogeneity of these patterns and imparted therapeutic opportunities remain poorly described.DesignWe performed comprehensive epigenomic characterisation by mapping 222 chromatin profiles from 69 samples (33 colorectal adenocarcinomas, 4 adenomas, 21 matched normal tissues and 11 colon cancer cell lines) for six histone modification marks: H3K4me3 for Pol II-bound and CpG-rich promoters, H3K4me1 for poised enhancers, H3K27ac for enhancers and transcriptionally active promoters, H3K79me2 for transcribed regions, H3K27me3 for polycomb repressed regions and H3K9me3 for heterochromatin.ResultsWe demonstrate that H3K27ac-marked active enhancer state could distinguish between different stages of CRC progression. By epigenomic editing, we present evidence that gains of tumour-specific enhancers for crucial oncogenes, such as ASCL2 and FZD10, was required for excessive proliferation. Consistently, combination of MEK plus bromodomain inhibition was found to have synergistic effects in CRC patient-derived xenograft models. Probing intertumour heterogeneity, we identified four distinct enhancer subtypes (EPIgenome-based Classification, EpiC), three of which correlate well with previously defined transcriptomic subtypes (consensus molecular subtypes, CMSs). Importantly, CMS2 can be divided into two EpiC subgroups with significant survival differences. Leveraging such correlation, we devised a combinatorial therapeutic strategy of enhancer-blocking bromodomain inhibitors with pathway-specific inhibitors (PARPi, EGFRi, TGFβi, mTORi and SRCi) for EpiC groups.ConclusionOur data suggest that the dynamics of active enhancer underlies CRC progression and the patient-specific enhancer patterns can be leveraged for precision combination therapy.

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A new in-situ generated HCl acid was developed to overcome the fast reaction rate and high corrosion rates of 15 wt% regular HCl acidizing system. The objectives of this work are to: (1) examine the reaction rate of the new in-situ generated HCl with calcite at 100, 150, and 200℉, and (2) compare the reaction rate of 15 wt% regular HCl with the new in-situ generated HCl. The rate of the reaction of 15 wt% HCl and the new in-situ generated HCl was measured using the rotating disk apparatus (RDA). Calcite disks were used with the specifications of 1.5 in. the diameter and 0.65 in. thickness. The effects of disk-rotational speed (200-1,200 rpm) and temperature (100-200℉) were investigated. Calcium concentrations were measured in the samples collected from the RDA, which were used to calculate the rate of dissolution. The disk surface after the tests was analyzed using Scanning-Electron-Microscope–Energy-Dispersive Spectroscopy (SEM-EDS). Experimental results showed that the rate of dissolution at 100 and 150°F was controlled mainly by the rate of mass transfer of the acid to the surface. By increasing the temperature to 200℉, the overall rate of reaction for the in-situ generated HCl was mass transfer limited up to 800 rpm and surface limited above 800 rpm. Based on the dissolution rate results, the diffusion coefficient, the activation energy, and the reaction rate constant at 100, 150, and 200°F were determined for the new developed in-situ generated HCl and were compared to 15 wt% regular HCl. This study will assist in developing a more cost-effective and efficient design of acid treatments through a slower reaction rate of the in-situ generated HCl. This new in-situ generated acid system reacts slower and more efficient compared to regular HCl in carbonate and sandstone reservoirs.