Metformin Pharmacokinetics in Mouse Tumors: Implications for Human Therapy

‘Prime-editing’ proposes to replace traditional programmable nucleases (CRISPR-Cas9) using a catalytically impaired Cas9 (dCas9) connected to a engineered reverse transcriptase, and a guide RNA encoding both the target site and the desired change. With just a ‘nick’ on one strand, it is hypothe- sized, the negative, uncontrollable effects arising from double-strand DNA breaks (DSBs) - translocations, complex proteins, integrations and p53 activation - will be eliminated. However, sequencing data pro- vided (Accid:PRJNA565979) reveal plasmid integration, indicating that DSBs occur. Also, looking at only 16 off-targets is inadequate to assert that Prime-editing is more precise. Integration of plasmid occurs in all three versions (PE1/2/3). Interestingly, dCas9 which is known to be toxic in E. coli and yeast, is shown to have residual endonuclease activity. This also affects studies that use dCas9, like base- editors and de/methylations systems. Previous work using hRad51–Cas9 nickases also show significant integration in on-targets, as well as off-target integration [1]. Thus, we show that cellular response to nicking involves DSBs, and subsequent plasmid/Cas9 integration. This is an unacceptable outcome for any in vivo application in human therapy.

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Tackling Pristinamycin IIB Problems: Synthetic Studies toward Some Fluorinated Analogs

Chemistry Proceedings ◽

10.3390/ecsoc-24-08388 ◽

2020 ◽

Vol 3 (1) ◽

pp. 107

Author(s):

Assia Chebieb ◽

Chewki Ziani-Cherif

Keyword(s):

Chemical Structure ◽

Wittig Reaction ◽

Antimicrobial Agents ◽

Synthetic Approach ◽

Solubility In Water ◽

Fluorinated Analogs ◽

Human Therapy ◽

Primary Structures ◽

Synthetic Studies

Streptogramins are potent antibiotics against numerous highly resistant pathogens and therefore are used in last-resort human therapy. These antibiotics are formed of both A- and B-group compounds named pristinamycins that differ in their basic primary structures. Although pristinamycin IIB is among the most interesting antibiotics in this family, it presents numerous problems related to its chemical structure, such as instability at most pH levels, weak solubility in water, and resistance by bacteria. As a response to the need for developing new antimicrobial agents, we have designed a new analog of pristinamycin IIB, based most importantly on the introduction of fluorine atoms. We conjectured indeed that the introduced modifications may solve the above-mentioned problems exhibited by pristinamycin IIB. Our multistep synthetic approach relies on few key reactions, namely a Wittig reaction, a Grubbs reaction, and dihydroxy, -difluoro API (Advanced Pharmaceutical Intermediate) synthesis

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TAMI-23. NEUTROPHIL-TRIGGERED FERROPTOSIS PROMOTES TUMOR NECROSIS IN GLIOBLASTOMA PROGRESSION

Neuro-Oncology ◽

10.1093/neuonc/noaa215.912 ◽

2020 ◽

Vol 22 (Supplement_2) ◽

pp. ii218-ii218

Author(s):

Patricia Yee ◽

Yiju Wei ◽

Soo Yeon Kim ◽

Tong Lu ◽

Cynthia Lawson ◽

...

Keyword(s):

Tumor Cells ◽

Tumor Necrosis ◽

Treatment Resistance ◽

Binding Motif ◽

Lipid Hydroperoxides ◽

Orthotopic Mouse Model ◽

Mouse Tumors ◽

Downstream Effector ◽

Human Gbm ◽

Early Tumor

Abstract Tumor necrosis indicates poor prognoses in many cancers, including glioblastomas (GBMs). Although thought to result from chronic ischemia, the underlying nature and mechanisms driving the involved cell death remain obscured by lack of animal models recapitulating the extent of necrosis in human GBMs. The molecular and clinical heterogeneity of GBMs adds further complexity. Not all GBMs contain necrosis. Mesenchymal (MES)-GBM, the subtype correlated with worst prognosis and highest treatment resistance, is most closely associated with necrosis. MES-GBM exhibits hyperactivity of transcriptional coactivator with PDZ-binding motif (TAZ), a Hippo tumor suppressive pathway effector whose expression in human GBMs predicts short survival. To elucidate mechanisms driving GBM necrosis, we devised a novel orthotopic mouse model recapitulating human MES-GBM phenotypically and histopathologically by expressing a constitutively-active TAZ mutant (TAZ4SA) in three human GBM cell lines (LN229, U87, and LN18) lacking MES signatures (GBM4SA). GBM4SA mice lived significantly shorter than mice implanted with GBMvector or mutant TAZ unable to bind its downstream effector, TEAD (GBM4SA-S51A). Extensive (≥30% of tumor volume) necrosis was present in GBM4SA mice but not GBMvector or GBM4SA-S51A. In GBM4SA tumors, neutrophils coincide with necrosis temporally and spatially. Neutrophil depletion dampens necrosis. Neutrophils isolated from mouse tumors killed co-cultured tumor cells. Neutrophils induce iron-dependent accumulation of lipid hydroperoxides within tumor cells by transferring myeloperoxidase-containing granules into tumor cells. Inhibiting myeloperoxidase suppresses neutrophil-induced tumor cytotoxicity. Intratumoral glutathione peroxidase 4 (GPX4) overexpression or acyl-CoA synthetase 4 (ACSL4) depletion diminishes necrosis and aggressiveness of tumors. Human GBM analysis indicates neutrophils and ferroptosis are associated with necrosis and predict poor survival. Together, we propose that certain tumor damage(s) during early tumor progression (i.e. ischemia) recruits neutrophils to damaged tissue and results in a positive feedback loop, amplifying GBM necrosis development. We show GBM necrosis involves neutrophil-triggered ferroptosis and reveal an unprecedented pro-tumorigenic role of ferroptosis.

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Antibiotic resistance of bacteria isolated from Pagellus erythrinus microplastics and public health

European Journal of Public Health ◽

10.1093/eurpub/ckaa166.253 ◽

2020 ◽

Vol 30 (Supplement_5) ◽

Author(s):

P Laganà ◽

E Fazio ◽

N Spanò ◽

M Bonsignore

Keyword(s):

Antibiotic Resistance ◽

Petri Dish ◽

Anthropogenic Pollution ◽

Normal Structure ◽

Multidrug Resistant ◽

Morphological Properties ◽

Marine Environments ◽

Pagellus Erythrinus ◽

Human Therapy ◽

The Moment

Abstract Microplastics (MPs) are widespread in the aquatic environmental due to anthropogenic pollution and are ingested indifferently by fishes that confuse them with food. In aquatic environment, mechanical stress, UV radiations, chemical and biological actions cause a constant degradation and breakdown of plastic objects into smaller fragments. Mediterranean Sea is characterized by the highest densities of plastics in the world being a closed basin with a complex hydrodynamics. The aim of this work is to evidence the occurrence of microbial adhesions on MPs found in edible Pagellus erythrinus (Linnaeus, 1758) bought in local Sicilian supermarkets and therefore highlight the potential role of microplastics in conveying antibiotic resistance by ingestion of food by humans. The composition and structural-morphological properties of MPs found in the excised gastrointestinal tract (GIT) and gills are identified using different techniques. In particular, microparticles of different nature (plastics, organic components, cellulose-based materials) have been determined following specific Raman signals on a large spectral range (300-3500 cm-1). The Pagellus was dissected, open longitudinally, within a sterile glass Petri dish. The components deemed exogenous to the normal structure of the gills and GIT were collected. Optical microscopy images showed that both the GIT and the gill of Pagellus erythrinus contain MPs of different colors (black, dark blu) and mainly with a fibrous shape. At the moment, microbial assays show the adhesion of Citrobacter and E. coli in some fibers extracted by gills while Vibrio spp was mainly detected in the fibers present in GIT. Bacterial isolates were screened for susceptibility to antibiotics using the Kirby-Bauer test, choosing the molecules most used in human therapy. The results obtained suggest that plastics may contribute to the spread of multiple antibiotic resistance in marine environments underline the relevance of future studies on this topic. Key messages Plastics can serve as vectors for the spread of multiple resistances to antibiotics across marine environments. Further studies on possible vehicles of multidrug-resistant germs carried by food of various kinds are desirable.

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Metastasis is altered through multiple processes regulated by the E2F1 transcription factor

Scientific Reports ◽

10.1038/s41598-021-88924-y ◽

2021 ◽

Vol 11 (1) ◽

Author(s):

Matthew R. Swiatnicki ◽

Eran R. Andrechek

Keyword(s):

Breast Cancer ◽

Transgenic Mouse Models ◽

Cellular Processes ◽

Mouse Tumors ◽

Mutation Burden ◽

Multiple Processes ◽

Metastatic Process ◽

Cycle Regulation ◽

Bioinformatic Approach ◽

Complicated Nature

AbstractThe E2F family of transcription factors is important for many cellular processes, from their canonical role in cell cycle regulation to other roles in angiogenesis and metastasis. Alteration of the Rb/E2F pathway occurs in various forms of cancer, including breast cancer. E2F1 ablation has been shown to decrease metastasis in MMTV-Neu and MMTV-PyMT transgenic mouse models of breast cancer. Here we take a bioinformatic approach to determine the E2F1 regulated genomic alterations involved in the metastatic cascade, in both Neu and PyMT models. Through gene expression analysis, we reveal few transcriptome changes in non-metastatic E2F1−/− tumors relative to transgenic tumor controls. However investigation of these models through whole genome sequencing found numerous differences between the models, including differences in the proposed tumor etiology between E2F1−/− and E2F1+/+ tumors induced by Neu or PyMT. For example, loss of E2F1 within the Neu model led to an increased contribution of the inefficient double stranded break repair signature to the proposed etiology of the tumors. While the SNV mutation burden was higher in PyMT mouse tumors than Neu mouse tumors, there was no statistically significant differences between E2F WT and E2F1 KO mice. Investigating mutated genes through gene set analysis also found a significant number of genes mutated in the cell adhesion pathway in E2F1−/− tumors, indicating this may be a route for disruption of metastasis in E2F1−/− tumors. Overall, these findings illustrate the complicated nature of uncovering drivers of the metastatic process.

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Irradiation of subcutaneous mouse tumors with a clinical linear accelerator validated by alanine dosimetry

Radiation Measurements ◽

10.1016/j.radmeas.2021.106636 ◽

2021 ◽

pp. 106636

Author(s):

C. Ankjærgaard ◽

A.Z. Johansen ◽

M.M.K. von Staffeldt ◽

C.E. Andersen ◽

D.H. Madsen ◽

...

Keyword(s):

Linear Accelerator ◽

Mouse Tumors

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In Vitro and In Vivo Multispectral Photoacoustic Imaging for the Evaluation of Chromophore Concentration

Sensors ◽

10.3390/s21103366 ◽

2021 ◽

Vol 21 (10) ◽

pp. 3366

Author(s):

Aneline Dolet ◽

Rita Ammanouil ◽

Virginie Petrilli ◽

Cédric Richard ◽

Piero Tortoli ◽

...

Keyword(s):

Remote Sensing ◽

Photoacoustic Imaging ◽

Hyperspectral Remote Sensing ◽

Reference Spectrum ◽

In Vivo Experiments ◽

Mouse Tumors ◽

Chromophore Concentration ◽

Saturation Rate

Multispectral photoacoustic imaging is a powerful noninvasive medical imaging technique that provides access to functional information. In this study, a set of methods is proposed and validated, with experimental multispectral photoacoustic images used to estimate the concentration of chromophores. The unmixing techniques used in this paper consist of two steps: (1) automatic extraction of the reference spectrum of each pure chromophore; and (2) abundance calculation of each pure chromophore from the estimated reference spectra. The compared strategies bring positivity and sum-to-one constraints, from the hyperspectral remote sensing field to multispectral photoacoustic, to evaluate chromophore concentration. Particularly, the study extracts the endmembers and compares the algorithms from the hyperspectral remote sensing domain and a dedicated algorithm for segmentation of multispectral photoacoustic data to this end. First, these strategies are tested with dilution and mixing of chromophores on colored 4% agar phantom data. Then, some preliminary in vivo experiments are performed. These consist of estimations of the oxygen saturation rate (sO2) in mouse tumors. This article proposes then a proof-of-concept of the interest to bring hyperspectral remote sensing algorithms to multispectral photoacoustic imaging for the estimation of chromophore concentration.

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