Press Release | Dosimetry and toxicity studies of a sulfonamide derivative of Sulforhodamine 101

 

The article by Dr. E Savio et al. is published in Current Radiopharmaceuticals, 2018

BACKGROUND

The SR101 N-(3-[18F]Fluoropropyl) sulfonamide ([18F]SRF101) is a Sulforhodamine 101 (SR101) derivative that was previously synthesised by our group. The fluorescent dye SR101 has been reported as a specific marker of astroglia in the neocortex of rodents in vivo. Some limitations of SR101 with respect to cell specificity have been identified. This derivative was labelled to study its biological in vivo behaviour through PET imaging.

As some recent investigations have shown, astroglia is affected in the early stages of different neurologic diseases, and reactive astrocytes contribute to neuroinflammation at later stages. This investigation focused on reactive astrocytosis in Alzheimer’s disease (AD). It is postulated that [18F]SRF101 is a potential PET radiotracer for astrocytosis detection. A biological characterisation of the novel tracer in a triple-transgenic mice model of AD (3xTg) was performed, and the results were compared with those of healthy Black C57BL6J mice. The biodistribution studies showed a hepatobiliary metabolisation of the compound. PET imaging with [18F]SRF101 revealed that the novel tracer could be a marker of astrocytosis in this animal model. These results suggested that [18F]SRF101 is a promising candidate for a more extensive evaluation as a PET astrocyte tracer.

To conclude the preclinical characterisation of [18F]SRF101 and to allow for drafting a pilot clinical evaluation in patients, radiation dosimetry studies of [18F]SRF101 injections were conducted on mice considering the following issues: i) the absorbed dose assessment can be based on biokinetic data obtained in small animals and ii) to develop new radiotracers for brain imaging, studies have been carried out on time-activity curves in different sections of the brain and in other organs.

A single-dose toxicity study was also carried out considering the guidelines on toxicology studies applicable to radiopharmaceuticals. The toxicological limit chosen in this case was <100 μg based on the concept of microdosing. The study was carried out with a dose set by allometric scaling with a safety factor of 100. Because [18F]SRF101 is a radiopharmaceutical prepared by a not-quantitative radiolabelling reaction and its synthetic procedure involves a purification step to separate the desired radioactive compound from the reaction mixture, toxicity studies were performed for unlabelled SRF101.

In summary, the aim of this study was to perform a toxicological evaluation of [18F]SRF101 and to estimate the human radiation dosimetry of this tracer based on preclinical studies. Obtaining this data will allow for moving forward to assess its potential as a PET imaging radiopharmaceutical for clinical use.

RESULTS

All animals survived until the end of the study with no systemic signs of toxicity throughout the entire observation period. No drug related changes were noted in the parameters examined during the 14-day study, including body weight, food consumption, eyes changes, clinical pathology parameters, gross necropsy findings, absolute and relative organ weights, histopathology findings, or microscopic lesions.

The absorbed and effective doses were estimated by OLINDA/EXM V2.0. Both dosimetric models–male and female–presented the same tendency. The highest total absorbed dose values were for the different sections of the intestines (left colon, small intestine, right colon and rectum). For the male dosimetric model, other organs exhibiting a high total absorbed dose included the liver, kidneys, gallbladder wall and pancreas. For the female dosimetric model, the uterus was added in addition to the mentioned organs.

The effective dose for male and female dosimetric models was 4.03E-03mSv/MBq and 5.08E-03mSv/MBq, respectively. For an administrated activity of 350MBq this would correspond to 1.41mSv and 1.78mSv for the same dosimetric models.

CONCLUSIONS

A toxicological evaluation of SRF101 was performed, which verified the biosafety of the new compound for single-intravenous injections in humans. The dosimetry calculations from the animal data revealed that the radiation-associated risk of [18F]SRF101 would be of the same order as other 18F radiopharmaceuticals used in clinical applications. In summary, the data generated through these studies confirm that the novel radiotracer would be safe for use in human PET imaging. This would allow for drafting a pilot clinical evaluation of patients.

Browse the Article at: http://www.eurekaselect.com/165008

Press Release | Modified LDL particles activate inflammatory pathways in monocyte-derived macrophages

The article by Dr. . Alexander N. Orekhov et al. is published in Current Pharmaceutical Design, 2018

 

One of the main characteristics of atherosclerosis is the accumulation of lipids in the intimal layer of the arterial wall. In atherosclerotic plaques, phagocytic cells, such as macrophages, engulf atherogenic low-density lipoprotein (LDL) particles, but are unable to process them, and thus become foam cells, having cytoplasm packed with lipid droplets. Foam cells are characterized by several typical features: they have decreased ability to migrate, while displaying enhanced production of pro-inflammatory cytokines. Therefore foam cells participate in maintaining chronic inflammation in the lesion. Such changes of phenotype in comparison to normal macrophages should be based on changes in gene expression patterns of these cells. The study of foam cell formation is of key importance to our understanding of atherosclerosis pathogenesis and for the development of novel diagnostic and therapeutic tools. However, little is known so far on gene expression changes that take place during conversion of macrophages to foam cells.

Previous studies have shown several clusters of genes up- or down-regulated in macrophages in response to oxidized LDL, which is known to be atherogenic. Among the up-regulated genes were scavenger receptors SCA and CD36, nuclear receptors PPARγ, LXRα and RXRγ, and cholesterol efflux protein ABCA1. Regarding the inflammatory response, modified LDL appeared to trigger up-regulation of genes with anti-inflammatory activities, such as IL1-RA, DSCR1, annexin 1, and the Burton’s tyrosine kinase repressor SH3 protein, and down-regulation of a number of pro-inflammatory genes, including leukotriene A4 hydrolase, cathepsin G, elastase 2, RNase A family 2 and 3 proteins, cytochromeb-245, and CD64. However, modern powerful tools, such as transcriptome analysis, may provide more detailed data on change of gene expression patterns during atherosclerotic plaque development and reveal causative relationships between gene expression patterns and pathologic phenotypic alterations.

We performed a transcriptome analysis of macrophages treated with atherogenic LDL that causes intracellular cholesterol accumulation. We used the strategy of upstream analysis for causal interpretation of the expression changes. This strategy has three major steps: (1) analysis of promoters and enhancers of identified differentially expressed genes to identify transcription factors involved in the process under study; (2) reconstruction of signaling pathways that activate these transcription factors; and (3) identification of master-regulators of these pathways.

In this study, we used human monocyte-derived macrophages treated with different lipoprotein-containing samples : high-density lipoprotein (HDL), native LDL, which does not induce cholesterol accumulation in cultured cells, and 3 types of modified atherogenic LDL (oxidized LDL, acetylated LDL and desialylated LDL). In this experiment, low concentrations of native LDL and HDL did not increase the total or esterified cholesterol content in cultured macrophages. After incubation with the substances, mRNA was isolated from the cells and analysed using high-throughput sequencing on HiSeq 1500.

In this study, we discovered 27 transcription factors, including c-Ets, GR-alpha, BRCA1, E2F-1, E2F-6 and EGR-1, that were potentially responsible for the changes in gene expression induced by modified atherogenic LDL. These transcription factors were used for identifying the master-regulators (genes and proteins) responsible for regulation of large cascades of differentially expressed genes. The most reliable of identified master-regulators were IL7R, TIGIT, CXCL8, F2RL1, EIF2AK3, IL7, TSPYL2, ANXA1, DUSP1 and IL15. In the Discussion section of our paper, we give more detail on each of these master-regulators. In general, the genes that were up-regulated in response to lipid accumulation in macrophages induced by atherogenic LDL were mostly involved in inflammation and immune response, and not in cholesterol metabolism. Our results suggest a possibility that it is not cholesterol accumulation that causes an innate immunity response, but rather the immune response is a consequence of a cellular reaction to modified LDL. These results highlight the importance of the inflammatory component in the pathogenesis of atherosclerosis.

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