Functional analysis reveals driver cooperativity and novel mechanisms in endometrial carcinogenesis.

High-risk endometrial cancer has poor prognosis and is increasing in incidence. However, understanding of the molecular mechanisms which drive this disease is limited. We used genetically engineered mouse models (GEMM) to determine the functional consequences of missense and loss of function mutations in Fbxw7, Pten and Tp53, which collectively occur in nearly 90% of high-risk endometrial cancers. We show that Trp53 deletion and missense mutation cause different phenotypes, with the latter a substantially stronger driver of endometrial carcinogenesis. We also show that Fbxw7 missense mutation does not cause endometrial neoplasia on its own, but potently accelerates carcinogenesis caused by Pten loss or Trp53 missense mutation. By transcriptomic analysis, we identify LEF1 signalling as upregulated in Fbxw7/FBXW7-mutant mouse and human endometrial cancers, and in human isogenic cell lines carrying FBXW7 mutation, and validate LEF1 and the additional Wnt pathway effector TCF7L2 as novel FBXW7 substrates. Our study provides new insights into the biology of high-risk endometrial cancer and suggests that targeting LEF1 may be worthy of investigation in this treatment-resistant cancer subgroup.

Discordant prognosis of mismatch repair deficiency in colorectal and endometrial cancer reflects variation in antitumour immune response and immune escape.

Defective DNA mismatch repair (dMMR) causes elevated tumour mutational burden (TMB) and microsatellite instability (MSI) in multiple cancer types. dMMR/MSI colorectal cancers (CRCs) have enhanced T-cell infiltrate and favourable outcome; however, this association has not been reliably detected in other tumour types, including endometrial cancer (EC). We sought to confirm this and explore the underpinning mechanisms. We first meta-analysed CRC and EC trials that have examined the prognostic value of dMMR/MSI and confirmed that dMMR/MSI predicts better prognosis in CRC, but not EC, with statistically significant variation between cancers (hazard ratio [HR] = 0.63, 95% confidence interval [CI] = 0.54-0.73 versus HR = 1.15, 95% CI = 0.72-1.58; PINT = 0.02). Next, we studied intratumoural immune infiltrate in CRCs and ECs of defined MMR status and found that while dMMR was associated with increased density of tumour-infiltrating CD3+ and CD8+ T-cells in both cancer types, the increases were substantially greater in CRC and significant only in this group (PINT = 4.3e-04 and 7.3e-03, respectively). Analysis of CRC and EC from the independent Cancer Genome Atlas (TCGA) series revealed similar variation and significant interactions in proportions of tumour-infiltrating lymphocytes, CD8+ , CD4+ , NK cells and immune checkpoint expression, confirming a more vigorous immune response to dMMR/MSI in CRC than EC. Agnostic analysis identified the IFNγ pathway activity as strongly upregulated by dMMR/MSI in CRC, but downregulated in EC by frequent JAK1 mutations, the impact of which on IFNγ response was confirmed by functional analyses. Collectively, our results confirm the discordant prognosis of dMMR/MSI in CRC and EC and suggest that this relates to differences in intratumoural immune infiltrate and tumour genome. Our study underscores the need for tissue-specific analysis of cancer biomarkers and may help inform immunotherapy use. © 2022 The Authors. The Journal of Pathology published by John Wiley & Sons Ltd on behalf of The Pathological Society of Great Britain and Ireland.

Multi-omics analyses of early liver injury reveals cell-type-specific transcriptional and epigenomic shift.

BACKGROUND: Liver fibrosis is a wound-healing response to tissue injury and inflammation hallmarked by the extracellular matrix (ECM) protein deposition in the liver parenchyma and tissue remodelling. Different cell types of the liver are known to play distinct roles in liver injury response. Hepatocytes and liver endothelial cells receive molecular signals indicating tissue injury and activate hepatic stellate cells which produce ECM proteins upon their activation. Despite the growing knowledge on the molecular mechanism underlying hepatic fibrosis in general, the cell-type-specific gene regulatory network associated with the initial response to hepatotoxic injury is still poorly characterized. RESULTS: In this study, we used thioacetamide (TAA) to induce hepatic injury in adult zebrafish. We isolated three major liver cell types - hepatocytes, endothelial cells and hepatic stellate cells - and identified cell-type-specific chromatin accessibility and transcriptional changes in an early stage of liver injury. We found that TAA induced transcriptional shifts in all three cell types hallmarked by significant alterations in the expression of genes related to fatty acid and carbohydrate metabolism, as well as immune response-associated and vascular-specific genes. Interestingly, liver endothelial cells exhibit the most pronounced response to liver injury at the transcriptome and chromatin level, hallmarked by the loss of their angiogenic phenotype. CONCLUSION: Our results uncovered cell-type-specific transcriptome and epigenome responses to early stage liver injury, which provide valuable insights into understanding the molecular mechanism implicated in the early response of the liver to pro-fibrotic signals.

Influence of Vibroacoustic Therapy on Local Status and Rehabilitation of Post-TKR and Post-THR patients.

BACKGROUND: Rehabilitation following major joint replacement surgery is an extremely important part of complementary treatment after the surgical procedure. To obtain the best final treatment outcome possible, apart from improvements in surgical technique, new methods of rehabilitation are still being sought and implemented to increase its effectiveness, safety and patient comfort. One such technique is vibroacoustic therapy, a method of treatment based on the direct transmission of audible sound vibrations through a sound transmitter to the patient's body. The present study assessed the effectiveness of vibroacoustic therapy in early postoperative rehabilitation of post-TKR and post-THR patients. MATERIAL AND METHODS: The study involved 60 post-TKR and post-THR patients divided into two groups depending on the type of surgery. Each group was further divided into an experimental group treated with the VITAFON-2 apparatus and a control group treated with a placebo apparatus; all patients also underwent routine kinesiotherapy that is offered at the hospital. Vibroa-coustic treatment was carried out and evaluated during the first 4 days following surgery. Symptom intensity, range of motion of the operated joints, oedema in the operated limb and haematoma absorption were assessed. RESULTS: Postoperative vibroacoustic therapy performed during the first 4 post-operative days reduced pain more significantly than kinesiotherapy alone. It also clinically improved the absorption of post-traumatic haematomas in post-TKR patients insignificantly better than kinesiotherapy. In post-THR patients who had undergone an anterolateral approach procedure, phoning (vibroacoustic stimulation) significantly sped up the rate of recovery of normal internal rotation angles in the operated joints. CONCLUSIONS: 1. Vibroacoustic therapy applied topically in post-TKR patients exerts a significant positive effect on reducing pain intensity. 2. Topical vibroacoustic therapy does not significantly reduce the oedema around the operated hip and knee joints or increase their range of motion. 3. Vibroacoustic therapy accelerates the absorption of postoperative subcutaneous haematomas in a clinically visible manner.

The MLH1 polymorphism rs1800734 and risk of endometrial cancer with microsatellite instability.

Both colorectal (CRC, 15%) and endometrial cancers (EC, 30%) exhibit microsatellite instability (MSI) due to MLH1 hypermethylation and silencing. The MLH1 promoter polymorphism, rs1800734 is associated with MSI CRC risk, increased methylation and reduced MLH1 expression. In EC samples, we investigated rs1800734 risk using MSI and MSS cases and controls. We found no evidence that rs1800734 or other MLH1 SNPs were associated with the risk of MSI EC. We found the rs1800734 risk allele had no effect on MLH1 methylation or expression in ECs. We propose that MLH1 hypermethylation occurs by different mechanisms in CRC and EC.

Dynamics of cardiomyocyte transcriptome and chromatin landscape demarcates key events of heart development.

Organogenesis involves dynamic regulation of gene transcription and complex multipathway interactions. Despite our knowledge of key factors regulating various steps of heart morphogenesis, considerable challenges in understanding its mechanism still exist because little is known about their downstream targets and interactive regulatory network. To better understand transcriptional regulatory mechanism driving heart development and the consequences of its disruption in vivo, we performed time-series analyses of the transcriptome and genome-wide chromatin accessibility in isolated cardiomyocytes (CMs) from wild-type zebrafish embryos at developmental stages corresponding to heart tube morphogenesis, looping, and maturation. We identified genetic regulatory modules driving crucial events of heart development that contained key cardiac TFs and are associated with open chromatin regions enriched for DNA sequence motifs belonging to the family of the corresponding TFs. Loss of function of cardiac TFs Gata5, Tbx5a, and Hand2 affected the cardiac regulatory networks and caused global changes in chromatin accessibility profile, indicating their role in heart development. Among regions with differential chromatin accessibility in mutants were highly conserved noncoding elements that represent putative enhancers driving heart development. The most prominent gene expression changes, which correlated with chromatin accessibility modifications within their proximal promoter regions, occurred between heart tube morphogenesis and looping, and were associated with metabolic shift and hematopoietic/cardiac fate switch during CM maturation. Our results revealed the dynamic regulatory landscape throughout heart development and identified interactive molecular networks driving key events of heart morphogenesis.

Genomic analysis of DNA repair genes and androgen signaling in prostate cancer.

BACKGROUND: The cellular effects of androgen are transduced through the androgen receptor, which controls the expression of genes that regulate biosynthetic processes, cell growth, and metabolism. Androgen signaling also impacts DNA damage signaling through mechanisms involving gene expression and transcription-associated DNA damaging events. Defining the contributions of androgen signaling to DNA repair is important for understanding androgen receptor function, and it also has translational implications. METHODS: We generated RNA-seq data from multiple prostate cancer lines and used bioinformatic analyses to characterize androgen-regulated gene expression. We compared the results from cell lines with gene expression data from prostate cancer xenografts, and patient samples, to query how androgen signaling and prostate cancer progression influences the expression of DNA repair genes. We performed whole genome sequencing to help characterize the status of the DNA repair machinery in widely used prostate cancer lines. Finally, we tested a DNA repair enzyme inhibitor for effects on androgen-dependent transcription. RESULTS: Our data indicates that androgen signaling regulates a subset of DNA repair genes that are largely specific to the respective model system and disease state. We identified deleterious mutations in the DNA repair genes RAD50 and CHEK2. We found that inhibition of the DNA repair enzyme MRE11 with the small molecule mirin inhibits androgen-dependent transcription and growth of prostate cancer cells. CONCLUSIONS: Our data supports the view that crosstalk between androgen signaling and DNA repair occurs at multiple levels, and that DNA repair enzymes in addition to PARPs, could be actionable targets in prostate cancer.

SONiCS: PCR stutter noise correction in genome-scale microsatellites.

Motivation: Massively parallel capture of short tandem repeats (STRs, or microsatellites) provides a strategy for population genomic and demographic analyses at high resolution with or without a reference genome. However, the high Polymerase Chain Reaction (PCR) cycle numbers needed for target capture experiments create genotyping noise through polymerase slippage known as PCR stutter. Results: We developed SONiCS-Stutter mONte Carlo Simulation-a solution for stutter correction based on dense forward simulations of PCR and capture experimental conditions. To test SONiCS, we genotyped a 2499-marker STR panel in 22 humpback dolphins (Sousa sahulensis) using target capture, and generated capillary-based genotypes to validate five of these markers. In these 110 comparisons, SONiCS showed a 99.1% accuracy rate and a 98.2% genotyping success rate, miscalling a single allele in a marker with low sequence coverage and rejecting another as un-callable. Availability and implementation: Source code and documentation for SONiCS is freely available at https://github.com/kzkedzierska/sonics. Raw read data used in experimental validation of SONiCS have been deposited in the Sequence Read Archive under accession number SRP135756. Supplementary information: Supplementary data are available at Bioinformatics online.

[18F]fluorothymidine PET imaging in the diagnosis of leptomeningeal involvement with diffuse large B-cell lymphoma.

The diagnosis of leptomeningeal carcinomatosis remains difficult despite improvement in central nervous system (CNS) imaging and cytologic examination of the cerebral spinal fluid. False-negative results are common, providing obstacles in assessing both prophylactic and therapeutic efforts. As a result of increased survival of patients with a variety of systemic neoplasms it is likely that central nervous involvement will need to be addressed more often. This article presents a patient with a diffuse large B-cell lymphoma with polymorphic features. Imaging using 18F-labeled fluorodeoxythymidine (FLT) proved useful in demonstrating both parenchymal and leptomeningeal CNS involvement. The potential for FLT to identify proliferative tissue may make it uniquely suitable for detection of CNS malignant disease.