Metastatic site influences driver gene function in pancreatic cancer.

Driver gene mutations can increase the metastatic potential of the primary tumor1-3, but their role in sustaining tumor growth at metastatic sites is poorly understood. A paradigm of such mutations is inactivation of SMAD4 - a transcriptional effector of TGFß signaling - which is a hallmark of multiple gastrointestinal malignancies4,5. SMAD4 inactivation mediates TGFß's remarkable anti- to pro-tumorigenic switch during cancer progression and can thus influence both tumor initiation and metastasis6-14. To determine whether metastatic tumors remain dependent on SMAD4 inactivation, we developed a mouse model of pancreatic ductal adenocarcinoma (PDAC) that enables Smad4 depletion in the pre-malignant pancreas and subsequent Smad4 reactivation in established metastases. As expected, Smad4 inactivation facilitated the formation of primary tumors that eventually colonized the liver and lungs. By contrast, Smad4 reactivation in metastatic disease had strikingly opposite effects depending on the tumor's organ of residence: suppression of liver metastases and promotion of lung metastases. Integrative multiomic analysis revealed organ-specific differences in the tumor cells' epigenomic state, whereby the liver and lungs harbored chromatin programs respectively dominated by the KLF and RUNX developmental transcription factors, with Klf4 depletion being sufficient to reverse Smad4's tumor-suppressive activity in liver metastases. Our results show how epigenetic states favored by the organ of residence can influence the function of driver genes in metastatic tumors. This organ-specific gene-chromatin interplay invites consideration of anatomical site in the interpretation of tumor genetics, with implications for the therapeutic targeting of metastatic disease.

Host response during unresolved urinary tract infection alters female mammary tissue homeostasis through collagen deposition and TIMP1.

Exposure to pathogens throughout a lifetime influences immunity and organ function. Here, we explore how the systemic host-response to bacterial urinary tract infection (UTI) induces tissue-specific alterations to the mammary gland. Utilizing a combination of histological tissue analysis, single cell transcriptomics, and flow cytometry, we identify that mammary tissue from UTI-bearing mice displays collagen deposition, enlarged ductal structures, ductal hyperplasia with atypical epithelial transcriptomes and altered immune composition. Bacterial cells are absent in the mammary tissue and blood of UTI-bearing mice, therefore, alterations to the distal mammary tissue are mediated by the systemic host response to local infection. Furthermore, broad spectrum antibiotic treatment resolves the infection and restores mammary cellular and tissue homeostasis. Systemically, unresolved UTI correlates with increased plasma levels of the metalloproteinase inhibitor, TIMP1, which controls extracellular matrix remodeling and neutrophil function. Treatment of nulliparous and post-lactation UTI-bearing female mice with a TIMP1 neutralizing antibody, restores mammary tissue normal homeostasis, thus providing evidence for a link between the systemic host response during UTI and mammary gland alterations.

Somatic mouse models of gastric cancer reveal genotype-specific features of metastatic disease.

Metastatic gastric carcinoma is a highly lethal cancer that responds poorly to conventional and molecularly targeted therapies. Despite its clinical relevance, the mechanisms underlying the behavior and therapeutic response of this disease are poorly understood owing, in part, to a paucity of tractable models. Here we developed methods to somatically introduce different oncogenic lesions directly into the murine gastric epithelium. Genotypic configurations observed in patients produced metastatic gastric cancers that recapitulated the histological, molecular and clinical features of all nonviral molecular subtypes of the human disease. Applying this platform to both wild-type and immunodeficient mice revealed previously unappreciated links between the genotype, organotropism and immune surveillance of metastatic cells, which produced distinct patterns of metastasis that were mirrored in patients. Our results establish a highly portable platform for generating autochthonous cancer models with flexible genotypes and host backgrounds, which can unravel mechanisms of gastric tumorigenesis or test new therapeutic concepts.

ENOX2 inhibition enhances infiltration of effector memory T-cell and mediates response to chemotherapy in immune-quiescent nasopharyngeal carcinoma.

INTRODUCTION: The immunosuppressive tumor microenvironment is a major barrier for chemotherapy. Different chemosensitization approaches to reinstate immunological surveillance for cancers that are immune quiescent at the outset, have thus been devised. Cancer-specific ENOX2 expression is correlated with abnormal cell growth and has been proposed as a cellular target for anti-cancer activity. However, the potential effects of ENOX2 on the interaction between immune system and tumor cells remain elusive. OBJECTIVES: To understand the mechanisms by which tumor-intrinsic ENOX2-mediated alterations in anti-tumor activity of T-cells and response to chemotherapy. METHODS: In situ multiplexed immunohistochemistry with single cell and bulk RNA sequencing data from nasopharyngeal carcinoma (NPC) human tissues were used to define tumor phenotypes. Two NPC cell lines, with distinct ENOX2 expression, were used in a co-culture platform to study tumor-immune interactions between cancer cells/spheroids and T-cells. The effect of cisplatin treatment with ENOX2 inhibition by idronoxil (IDX) were tested in vitro and in vivo. Multi-parametric flow cytometry was used to characterize T-cell infiltrates in an NPC tumor humanized mouse model treated with combined treatment. RESULTS: NPC predominantly displayed an immune-excluded profile. This "cold-phenotype" was shown to exhibit higher ENOX2 expression and was associate with poorer progression-free survival (PFS). The therapeutic combination of IDX with cisplatin was effective in promoting CD8+ effector memory T cell (Tem) differentiation and mobilization. This Tem signature was highly cytotoxic, with Tem-mediated preferential lysis of higher ENOX2-expressing NPC cells. A combination-treated humanized mouse model showing dramatic shrinkage in tumors, were intra-tumoral Tem-enriched. CONCLUSION: Tumor-intrinsic ENOX2 expression is associated with tumor phenotype and PFS in NPC. Targeting ENOX2 with IDX and cisplatin impose qualitative control of T-cell response by preferentially increasing immune cells infiltration, Tem differentiation and tumor suppression. We suggest that ENOX2 inhibition may be a promising therapeutic strategy to enhance the effects of chemotherapy.

Pharmaceuticals and Personal Care Products in the Aquatic Environment: How Can Regions at Risk be Identified in the Future?

Pharmaceuticals and personal care products (PPCPs) are an indispensable component of a healthy society. However, they are well-established environmental contaminants, and many can elicit biological disruption in exposed organisms. It is now a decade since the landmark review covering the top 20 questions on PPCPs in the environment (Boxall et al., 2012). In the present study we discuss key research priorities for the next 10 years with a focus on how regions where PPCPs pose the greatest risk to environmental and human health, either now or in the future, can be identified. Specifically, we discuss why this problem is of importance and review our current understanding of PPCPs in the aquatic environment. Foci include PPCP occurrence and what drives their environmental emission as well as our ability to both quantify and model their distribution. We highlight critical areas for future research including the involvement of citizen science for environmental monitoring and using modeling techniques to bridge the gap between research capacity and needs. Because prioritization of regions in need of environmental monitoring is needed to assess future/current risks, we also propose four criteria with which this may be achieved. By applying these criteria to available monitoring data, we narrow the focus on where monitoring efforts for PPCPs are most urgent. Specifically, we highlight 19 cities across Africa, Central America, the Caribbean, and Asia as priorities for future environmental monitoring and risk characterization and define four priority research questions for the next 10 years. Environ Toxicol Chem 2024;43:575-588. © 2023 The Authors. Environmental Toxicology and Chemistry published by Wiley Periodicals LLC on behalf of SETAC.

A reversible epigenetic memory of inflammatory injury controls lineage plasticity and tumor initiation in the mouse pancreas.

Inflammation is essential to the disruption of tissue homeostasis and can destabilize the identity of lineage-committed epithelial cells. Here, we employ lineage-traced mouse models, single-cell transcriptomic and chromatin analyses, and CUT&TAG to identify an epigenetic memory of inflammatory injury in the pancreatic acinar cell compartment. Despite resolution of pancreatitis, our data show that acinar cells fail to return to their molecular baseline, with retention of elevated chromatin accessibility and H3K4me1 at metaplasia genes, such that memory represents an incomplete cell fate decision. In vivo, we find this epigenetic memory controls lineage plasticity, with diminished metaplasia in response to a second insult but increased tumorigenesis with an oncogenic Kras mutation. The lowered threshold for oncogenic transformation, in turn, can be restored by blockade of MAPK signaling. Together, we define the chromatin dynamics, molecular encoding, and recall of a prolonged epigenetic memory of inflammatory injury that impacts future responses but remains reversible.

A review of complex in vitro cell culture stressing the importance of fluid flow and illustrated by organ on a chip liver models.

The translation of new technology from development into widespread commercial use is a complex and time-consuming process that requires significant investment. This review looks at some important market needs for more complex in vitro models, the technical difficulties that must be overcome, particularly those connected with introducing fluid flow using microfluidics, and also illustrates the economic benefits of more accurate models for drug toxicity. Beyond the strong ethical arguments for replacing the use of animals in drug safety testing and medical research, the author believes that financial benefits of adopting the new in vitro technology are becoming clear and will drive the adoption by industry.

Antisense oligonucleotide therapy for H3.3K27M diffuse midline glioma.

Diffuse midline gliomas (DMGs) are pediatric high-grade brain tumors in the thalamus, midbrain, or pons; the latter subgroup are termed diffuse intrinsic pontine gliomas (DIPG). The brain stem location of these tumors limits the clinical management of DIPG, resulting in poor outcomes for patients. A heterozygous, somatic point mutation in one of two genes coding for the noncanonical histone H3.3 is present in most DIPG tumors. This dominant mutation in the H3-3A gene results in replacement of lysine 27 with methionine (K27M) and causes a global reduction of trimethylation on K27 of all wild-type histone H3 proteins, which is thought to be a driving event in gliomagenesis. In this study, we designed and systematically screened 2'-O-methoxyethyl phosphorothioate antisense oligonucleotides (ASOs) that direct RNase H-mediated knockdown of H3-3A mRNA. We identified a lead ASO that effectively reduced H3-3A mRNA and H3.3K27M protein and restored global H3K27 trimethylation in patient-derived neurospheres. We then tested the lead ASO in two mouse models of DIPG: an immunocompetent mouse model using transduced mutant human H3-3A cDNA and an orthotopic xenograft with patient-derived cells. In both models, ASO treatment restored K27 trimethylation of histone H3 proteins and reduced tumor growth, promoted neural stem cell differentiation into astrocytes, neurons, and oligodendrocytes, and increased survival. These results demonstrate the involvement of the H3.3K27M oncohistone in tumor maintenance, confirm the reversibility of the aberrant epigenetic changes it promotes, and provide preclinical proof of concept for DMG antisense therapy.

Splicing Factor SRSF1 Promotes Pancreatitis and KRASG12D-Mediated Pancreatic Cancer.

Inflammation is strongly associated with pancreatic ductal adenocarcinoma (PDAC), a highly lethal malignancy. Dysregulated RNA splicing factors have been widely reported in tumorigenesis, but their involvement in pancreatitis and PDAC is not well understood. Here, we report that the splicing factor SRSF1 is highly expressed in pancreatitis, PDAC precursor lesions, and tumors. Increased SRSF1 is sufficient to induce pancreatitis and accelerate KRASG12D-mediated PDAC. Mechanistically, SRSF1 activates MAPK signaling-partly by upregulating interleukin 1 receptor type 1 (IL1R1) through alternative-splicing-regulated mRNA stability. Additionally, SRSF1 protein is destabilized through a negative feedback mechanism in phenotypically normal epithelial cells expressing KRASG12D in mouse pancreas and in pancreas organoids acutely expressing KRASG12D, buffering MAPK signaling and maintaining pancreas cell homeostasis. This negative feedback regulation of SRSF1 is overcome by hyperactive MYC, facilitating PDAC tumorigenesis. Our findings implicate SRSF1 in the etiology of pancreatitis and PDAC, and point to SRSF1-misregulated alternative splicing as a potential therapeutic target. SIGNIFICANCE: We describe the regulation of splicing factor SRSF1 expression in the context of pancreas cell identity, plasticity, and inflammation. SRSF1 protein downregulation is involved in a negative feedback cellular response to KRASG12D expression, contributing to pancreas cell homeostasis. Conversely, upregulated SRSF1 promotes pancreatitis and accelerates KRASG12D-mediated tumorigenesis through enhanced IL1 and MAPK signaling. This article is highlighted in the In This Issue feature, p. 1501.

CRISPR-induced exon skipping of ß-catenin reveals tumorigenic mutants driving distinct subtypes of liver cancer.

CRISPR/Cas9-driven cancer modeling studies are based on the disruption of tumor suppressor genes by small insertions or deletions (indels) that lead to frame-shift mutations. In addition, CRISPR/Cas9 is widely used to define the significance of cancer oncogenes and genetic dependencies in loss-of-function studies. However, how CRISPR/Cas9 influences gain-of-function oncogenic mutations is elusive. Here, we demonstrate that single guide RNA targeting exon 3 of Ctnnb1 (encoding ß-catenin) results in exon skipping and generates gain-of-function isoforms in vivo. CRISPR/Cas9-mediated exon skipping of Ctnnb1 induces liver tumor formation in synergy with YAPS127A in mice. We define two distinct exon skipping-induced tumor subtypes with different histological and transcriptional features. Notably, ectopic expression of two exon-skipped ß-catenin transcript isoforms together with YAPS127A phenocopies the two distinct subtypes of liver cancer. Moreover, we identify similar CTNNB1 exon-skipping events in patients with hepatocellular carcinoma. Collectively, our findings advance our understanding of ß-catenin-related tumorigenesis and reveal that CRISPR/Cas9 can be repurposed, in vivo, to study gain-of-function mutations of oncogenes in cancer. © 2023 The Authors. The Journal of Pathology published by John Wiley & Sons Ltd on behalf of The Pathological Society of Great Britain and Ireland.

Regulatory Risk Assessment of Pharmaceuticals in the Environment: Current Practice and Future Priorities.

How can data on the occurrence of pharmaceuticals and personal care products (PPCPs) in the environment and the quality of ecosystems exposed to PPCPs be used to determine whether current regulatory risk assessment schemes are effective? This is one of 20 "big questions" concerning PPCPs in the environment posed in a landmark review paper in 2012. Ten years later, we review the developments around this question, focusing on the first P in PPCPs, that is, pharmaceuticals, or more specifically the active ingredients included in them (active pharmaceutical ingredients, APIs). We illustrate how extensive data on both the occurrence of APIs and the ecotoxicological sensitivity of aquatic species to them can be used in a retrospective risk assessment. In the Netherlands, current regulatory risk assessment schemes offer insufficient protection against direct ecotoxicological effects from APIs: the toxic pressure exerted by the 39 APIs included in our study exceeds the policy-related protective threshold of 0.05 (the "95%-protection level") in at least 13% of sampled surface waters. In general, anti-inflammatory and antirheumatic products (e.g., diclofenac, ibuprofen) contributed most to the overall toxic pressure, followed by sex hormones and modulators of the genital system (e.g., ethinylestradiol) and psychoanaleptics (e.g., caffeine). We formulated three open questions for future research. The first relates to improving the availability and accessibility of good-quality ecotoxicity data on pharmaceuticals for the global scientific, regulatory, and general public. The second relates to the adaptation of regulatory risk assessment frameworks for developing regions of the world. The third relates to the integration of effect-based and ecological approaches into regulatory risk assessment practice. Environ Toxicol Chem 2023;00:1-12. © 2022 The Authors. Environmental Toxicology and Chemistry published by Wiley Periodicals LLC on behalf of SETAC.

Platr4 is an early embryonic lncRNA that exerts its function downstream on cardiogenic mesodermal lineage commitment.

The mammalian genome encodes thousands of long non-coding RNAs (lncRNAs), many of which are developmentally regulated and differentially expressed across tissues, suggesting their potential roles in cellular differentiation. Despite this expression pattern, little is known about how lncRNAs influence lineage commitment at the molecular level. Here, we demonstrate that perturbation of an embryonic stem cell/early embryonic lncRNA, pluripotency-associated transcript 4 (Platr4), directly influences the specification of cardiac-mesoderm-lineage differentiation. We show that Platr4 acts as a molecular scaffold or chaperone interacting with the Hippo-signaling pathway molecules Yap and Tead4 to regulate the expression of a downstream target gene, Ctgf, which is crucial to the cardiac-lineage program. Importantly, Platr4 knockout mice exhibit myocardial atrophy and valve mucinous degeneration, which are both associated with reduced cardiac output and sudden heart failure. Together, our findings provide evidence that Platr4 is required in cardiac-lineage specification and adult heart function in mice.

Entanglement between Muon and I>1/2 Nuclear Spins as a Probe of Charge Environment.

We report on the first example of quantum coherence between the spins of muons and quadrupolar nuclei. We reveal that these entangled states are highly sensitive to a local charge environment and thus, can be deployed as a functional quantum sensor of that environment. The quantum coherence effect was observed in vanadium intermetallic compounds which adopt the A15 crystal structure, and whose members include all technologically pertinent superconductors. Furthermore, the extreme sensitivity of the entangled states to the local structural and electronic environments emerges through the quadrupolar interaction with the electric field gradient due to the charge distribution at the nuclear (I>1/2) sites. This case study demonstrates that positive muons can be used as a quantum sensing tool to also probe structural and charge-related phenomena in materials, even in the absence of magnetic degrees of freedom.

Excitation function of 54Fe(p,α)51Mn from 9.5 MeV to 18 MeV.

Excitation function of the 54Fe(p,α)51Mn reaction was measured from 9.5 to 18 MeV E 0 , p + by activating a foil stack of 54Fe electrodeposited on copper substrates. Residual radionuclides were quantified by HPGe gamma ray spectrometry. Both 51Mn (t 1/2 = 46.2 min, 〈 E ß + 〉 = 963.7 keV , I ß + = 97 % ; E γ = 749.1 keV, I γ = 0.265%) and its radioactive daughter, 51Cr (t 1/2 = 27.704d, E γ = 320.1 keV, I γ = 9.91%), were used to indirectly quantify formation of 51Mn. Results agree within uncertainty to the only other measurement in literature and predictions of default TALYS theoretical code. Final relative uncertainties are within ±12%.

Toward A Regulatory Pathway for the Use of in Silico Trials in the CE Marking of Medical Devices.

In Silico Trials methodologies will play a growing and fundamental role in the development and de-risking of new medical devices in the future. While the regulatory pathway for Digital Patient and Personal Health Forecasting solutions is clear, it is more complex for In Silico Trials solutions, and therefore deserves a deeper analysis. In this position paper, we investigate the current state of the art towards the regulatory system for in silico trials applied to medical devices while exploring the European regulatory system toward this topic. We suggest that the European regulatory system should start a process of innovation: in principle to limit distorted quality by different internal processes within notified bodies, hence avoiding that the more innovative and competitive companies focus their attention on the needs of other large markets, like the USA, where the use of such radical innovations is already rapidly developing.

PTP1B inhibitors protect against acute lung injury and regulate CXCR4 signaling in neutrophils.

Acute lung injury (ALI) can cause acute respiratory distress syndrome (ARDS), a lethal condition with limited treatment options and currently a common global cause of death due to COVID-19. ARDS secondary to transfusion-related ALI (TRALI) has been recapitulated preclinically by anti-MHC-I antibody administration to LPS-primed mice. In this model, we demonstrate that inhibitors of PTP1B, a protein tyrosine phosphatase that regulates signaling pathways of fundamental importance to homeostasis and inflammation, prevented lung injury and increased survival. Treatment with PTP1B inhibitors attenuated the aberrant neutrophil function that drives ALI and was associated with release of myeloperoxidase, suppression of neutrophil extracellular trap (NET) formation, and inhibition of neutrophil migration. Mechanistically, reduced signaling through the CXCR4 chemokine receptor, particularly to the activation of PI3Kγ/AKT/mTOR, was essential for these effects, linking PTP1B inhibition to promoting an aged-neutrophil phenotype. Considering that dysregulated activation of neutrophils has been implicated in sepsis and causes collateral tissue damage, we demonstrate that PTP1B inhibitors improved survival and ameliorated lung injury in an LPS-induced sepsis model and improved survival in the cecal ligation and puncture-induced (CLP-induced) sepsis model. Our data highlight the potential for PTP1B inhibition to prevent ALI and ARDS from multiple etiologies.

Costs of Physician Continuous Professional Development: A Systematic Review.

PURPOSE: An essential yet oft-neglected step in cost evaluations is the selection of resources (ingredients) to include in cost estimates. The ingredients that most influence the cost of physician continuous professional development (CPD) are unknown, as are the relative costs of instructional modalities. This study's purpose was to estimate the costs of cost ingredients and instructional modalities in physician CPD. METHOD: The authors conducted a systematic review in April 2020, searching MEDLINE, Embase, PsycInfo, and the Cochrane Library for comparative cost evaluations of CPD for practicing physicians. Two reviewers, working independently, screened articles for inclusion and extracted information on costs (converted to 2021 U.S. dollars) for each intervention overall, each ingredient, and each modality. RESULTS: Of 3,338 eligible studies, 62 were included, enumerating costs for 86 discrete training interventions or instructional modalities. The most frequently reported ingredients were faculty time (25 of 86 interventions), materials (24), administrator/staff time (23), and travel (20). Ingredient costs varied widely, ranging from a per-physician median of $4 for postage (10 interventions) to $525 for learner time (13); equipment (9) and faculty time were also relatively expensive (median > $170). Among instructional modalities (≤ 11 interventions per modality), audit and feedback performed by physician learners, computer-based modules, computer-based virtual patients, in-person lectures, and experiences with real patients were relatively expensive (median > $1,000 per physician). Mailed paper materials, video clips, and audit and feedback performed by others were relatively inexpensive (median ≤ $62 per physician). Details regarding ingredient selection (10 of 62 studies), quantitation (10), and pricing (26) were reported infrequently. CONCLUSIONS: Some ingredients, including time, are more important (i.e., contribute more to total costs) than others and should be prioritized in cost evaluations. Data on the relative costs of instructional modalities are insightful but limited. The methods and reporting of cost valuations merit improvement.

Assessment of the Potential Ecotoxicological Effects of Pharmaceuticals in the World's Rivers.

During their production, use, and disposal, active pharmaceutical ingredients (APIs) are released into aquatic systems. Because they are biologically active molecules, APIs have the potential to adversely affect nontarget organisms. We used the results of a global monitoring study of 61 APIs alongside available ecotoxicological and pharmacological data to assess the potential ecotoxicological effects of APIs in rivers across the world. Approximately 43.5% (461 sites) of the 1052 sampling locations monitored across 104 countries in a recent global study had concentrations of APIs of concern based on apical, nonapical, and mode of action-related endpoints. Approximately 34.1% of the 137 sampling campaigns had at least one location where concentrations were of ecotoxicological concern. Twenty-three APIs occurred at concentrations exceeding "safe" concentrations, including substances from the antidepressant, antimicrobial, antihistamine, ß-blocker, anticonvulsant, antihyperglycemic, antimalarial, antifungal, calcium channel blocker, benzodiazepine, painkiller, progestin, and lifestyle compound classes. At the most polluted sites, effects are predicted on different trophic levels and on different endpoint types. Overall, the results show that API pollution is a global problem that is likely negatively affecting the health of the world's rivers. To meet the United Nations' Sustainable Development Goals, work is urgently needed to tackle the problem and bring concentrations down to an acceptable level. Environ Toxicol Chem 2022;41:2008-2020. © 2022 The Authors. Environmental Toxicology and Chemistry published by Wiley Periodicals LLC on behalf of SETAC.