Chemical Composition, Functional and Anticancer Properties of Carrot.

Plants are a valuable source of drugs for cancer treatment. Daucus carota has been investigated for its health properties. In particular, Daucus carota L. subsp. Sativus, the common edible carrot root, has been found to be rich in bioactive compounds such as carotenoids and dietary fiber and contains many other functional components with significant health-promoting features, while Daucus carota L. subsp. Carrot (Apiacae), also known as wild carrot, has been usually used for gastric ulcer therapy, diabetes, and muscle pain in Lebanon. Here, we review the chemical composition of Daucus carota L. and the functional properties of both edible and wild carrot subspecies. Then, we focus on compounds with anticancer characteristics identified in both Daucus carota subspecies, and we discuss their potential use in the development of novel anticancer therapeutic strategies.

Different Extraction Procedures Revealed the Anti-Proliferation Activity from Vegetable Semi-Purified Sources on Breast Cancer Cell Lines.

Breast cancer (BC) remains the leading cause of mortality in women, despite significant advancements in diagnosis. Thus, the identification of new compounds for its treatment is critical. Phytochemicals are known to exhibit anti-cancer properties. Here, we investigated the anti-proliferation potential of extracts from carrot, Calendula officinalis flower, and Aloe vera on breast cancer vs. epithelial cell lines. Various extraction methods were used, and the proliferative effect of the resulting extracts was assessed by proliferation assay on breast cancer and epithelial cell lines. Carrot, Aloe leaf, and Calendula flower extracts were extracted by hexane and methanol methods, and their semi-purified extracts were able to specifically inhibit the proliferation of breast cancer cell lines. The extract composition was investigated by colorimetric assays, UHPLC-HRMS, and MS/MS analysis. All the extracts contained monogalactosyl-monoacylglycerol (MGMG), while digalactosyl-monoacylglycerol (DGMG) and aloe-emodin were found in Aloe, and glycerophosphocholine (GPC) derivatives were identified in Calendula, except for the isomer 2 detected in carrot, suggesting that their observed different anti-proliferative properties may be associated with the different lipid compounds. Interestingly, Calendula extract was able to strongly inhibit the triple negative breast cancer MDA-MB-231 cell line proliferation (about 20% cell survival), supporting MGMG and GPC derivatives as potential drugs for this BC subtype treatment.

Development of a Qualitative Test to Detect the Presence of Organophosphate Pesticides on Fruits and Vegetables.

BACKGROUND: In recent decades, the use of pesticides in agriculture has increased at a fast pace, highlighting safety problems for the environment and human health, which in turn has made it necessary to develop new detection and decontamination systems for pesticides. METHODS: A new qualitative test capable of detecting the presence of pesticides on fruits and vegetables by using thermostable enzymes was discovered, and the test was carried out on apples and aubergines. The contaminating pesticides were extracted from fruits with acetonitrile and analyzed with a biosensor system based on the thermostable esterase EST2 immobilized on a nitrocellulose filter. This enzyme is irreversibly inhibited mainly in the presence of organophosphates pesticides. Therefore, by observing esterase activity inhibition, we revealed the presence of residual pesticides on the fruits and vegetables. RESULTS: By analyzing the rate of esterase activity inhibition, we predicted that residual pesticides are present on the surface of the fruits. When we cleaned the fruits by washing them in the presence of the phosphotriesterase SsoPox before the detection of the esterase activity on filters, we observed a full recovery of the activity for apples and 30% for aubergines, indicating that the enzymatic decontamination of organophosphates pesticides took place. CONCLUSIONS: The reported method permitted us to assess the pesticides present on the vegetables and their decontamination.

Melanoma Immunotherapy and Precision Medicine in the Era of Tumor Micro-Tissue Engineering: Where Are We Now and Where Are We Going?

Malignant melanoma still remains a cancer with very poor survival rates, although it is at the forefront of personalized medicine. Most patients show partial responses and disease progressed due to adaptative resistance mechanisms, preventing long-lasting clinical benefits to the current treatments. The response to therapies can be shaped by not only taking into account cancer cell heterogeneity and plasticity, but also by its structural context as well as the cellular component of the tumor microenvironment (TME). Here, we review the recent development in the field of immunotherapy and target-based therapy and how, in the era of tumor micro-tissue engineering, ex-vivo assays could help to enhance our melanoma biology knowledge in its complexity, translating it in the development of successful therapeutic strategies, as well as in the prediction of therapeutic benefits.

Golgi maturation-dependent glycoenzyme recycling controls glycosphingolipid biosynthesis and cell growth via GOLPH3.

Glycosphingolipids are important components of the plasma membrane where they modulate the activities of membrane proteins including signalling receptors. Glycosphingolipid synthesis relies on competing reactions catalysed by Golgi-resident enzymes during the passage of substrates through the Golgi cisternae. The glycosphingolipid metabolic output is determined by the position and levels of the enzymes within the Golgi stack, but the mechanisms that coordinate the intra-Golgi localisation of the enzymes are poorly understood. Here, we show that a group of sequentially-acting enzymes operating at the branchpoint among glycosphingolipid synthetic pathways binds the Golgi-localised oncoprotein GOLPH3. GOLPH3 sorts these enzymes into vesicles for intra-Golgi retro-transport, acting as a component of the cisternal maturation mechanism. Through these effects, GOLPH3 controls the sub-Golgi localisation and the lysosomal degradation rate of specific enzymes. Increased GOLPH3 levels, as those observed in tumours, alter glycosphingolipid synthesis and plasma membrane composition thereby promoting mitogenic signalling and cell proliferation. These data have medical implications as they outline a novel oncogenic mechanism of action for GOLPH3 based on glycosphingolipid metabolism.

WTAP and BIRC3 are involved in the posttranscriptional mechanisms that impact on the expression and activity of the human lactonase PON2.

The activity of human paraoxonase 2 (PON2) is rapidly reduced in cells incubated with the bacterial quorormone 3-Oxo-dodecanoyl Homoserine Lactone (3OC12HSL), an observation that led to hypothesize a fast PON2 post-translational modification (PTM). Recently, we detected a 3OC12HSL-induced PTM in a cell-free system in which a crude extract from 3OC12HSL-treated HeLa cells was able to inactivate and ubiquitinate at position 144 a recombinant PON2. Here we show the occurrence of this and new PTMs on PON2 in HeLa cells. PTMs were found to gather nearby the two SNPs, A148G, and S311C, that are related to type-2 diabetes and its complications. Furthermore, we detected a PTM nearby a 12 amino acids region that is deleted in PON2 Isoform 2. An in vitro mutation analysis showed that the SNPs and the deletion are involved in PON2 activity and suggested a role of PTMs on its modulation, while a SAXS analysis pointed to Isoform 2 as being largely unstructured, compared to the wild type. Besides, we discovered a control of PON2 expression via a putative mRNA operon involving the Wilms tumor 1 associated protein (WTAP) and the E3 ubiquitin ligase (E3UbL) baculoviral IAP repeat-containing 3 (BIRC3).

Brassicaceae-Derived Anticancer Agents: Towards a Green Approach to Beat Cancer.

Cancer is the main cause of mortality and morbidity worldwide. Although a large variety of therapeutic approaches have been developed and translated into clinical protocols, the toxic side effects of cancer treatments negatively impact patients, allowing cancer to grow. Brassica metabolites are emerging as new weapons for anti-cancer therapeutics. The beneficial role of the consumption of brassica vegetables, the most-used vegetables in the Mediterranean diet, particularly broccoli, in the prevention of chronic diseases, including cardiovascular diseases, diabetes, and obesity, has been well-documented. In this review, we discuss the anti-tumor effects of the bioactive compounds from Brassica vegetables with regard to the compounds and types of cancer against which they show activity, providing current knowledge on the anti-cancer effects of Brassica metabolites against major types of tumors. In addition, we discuss the impacts of industrial and domestic processing on the compounds' functional properties before their consumption as well as the main strategies used to increase the content of health-promoting metabolites in Brassica plants through biofortification. Finally, the impacts of microbiota on the compounds' bioactivity are considered. This information will be helpful for the further development of efficacious anti-cancer drugs.

A proteomic approach to study Escherichia coli. Acetyl esterase interactors unveil a sequence motif involved in protein-protein interaction.

E. coli acetyl esterase (Aes) and beta-cysthationase (MalY) interact, probably with the same mechanism, with the N-terminus of transcriptional activator of maltose regulon (MalT). In order to investigate the basic mechanism of this interaction, we used both a proteomic and a bioinformatic approach. Affinity-based mass spectrometry experiments with purified Aes protein as bait allowed to fish twenty-three, apparently specific, interactors from crude extracts of E. coli cells grown in different conditions. The group of interactors appeared quite heterogeneous, comprising Aes itself, some molecular chaperons, metabolic enzymes, and several proteins of unknown function. Among the identified proteins, two are in some way related to the maltose metabolism and two are related to the lipopolysaccharide metabolism. By superposing the structures of the Alicyclobacillus acidocaldarius EST2, an Aes homolog, and MalY, a region of structural similarity was discovered that allowed detecting a short stretch of nine residues with sequence similarity among EST2, AES and MalY. Degenerated sequence consensuses derived from the alignment were used to analyse the E. coli proteome in the Swiss Prot database and permitted to retrieve sequences of Aes interactors already known or detected in this study. Most of these interactors (14 out of 25) contain the expected consensus. A site-directed mutant of Aes R179A made in the consensus sequence resulted in complete loss of interaction. Based on the analysis of the available three-dimensional structures and mutagenic and structural data inferred from literature, we predict a role of this motif in protein-protein interaction.

The Aes protein and the monomeric alpha-galactosidase from Escherichia coli form a non-covalent complex. Implications for the regulation of carbohydrate metabolism.

Aes, a 36-kDa acetylesterase from Escherichia coli, belongs to the hormone-sensitive lipase family, and it is involved in the regulation of MalT, the transcriptional activator of the maltose regulon. The activity of MalT is depressed through a direct protein-protein interaction with Aes. Although the effect is clear-cut, the meaning of this interaction and the conditions that trigger it still remain elusive. To perform a comparative thermodynamic study between the mesophilic Aes protein and two homologous thermostable enzymes, Aes was overexpressed in E. coli and purified. At the last step of the purification procedure the enzyme was eluted from a Mono Q HR 5/5 column as a major form migrating, anomalously, at 56 kDa on a calibrated Superdex 75 column. A minor peak that contains the Aes protein and a polypeptide of 50 kDa was also detected. By a combined analysis of size-exclusion chromatography and surface-enhanced laser desorption ionization-time of flight mass spectrometry, it was possible to demonstrate the presence in this peak of a stable 87-kDa complex, containing the Aes protein itself and the 50-kDa polypeptide in a 1:1 ratio. The homodimeric molecular species of Aes and of the 50-kDa polypeptide were also detected. The esterase activity associated with the 87-kDa complex, when assayed with p-nitrophenyl butanoate as substrate, proved 6-fold higher than the activity of the major Aes form of 56 kDa. Amino-terminal sequencing highlighted that the 50-kDa partner of Aes in the complex was the alpha-galactosidase from E. coli. The E. coli cells harboring plasmid pT7-SCII-aes and, therefore, expressing Aes were hampered in their growth on a minimal medium containing raffinose as a sole carbon source. Because alpha-galactosidase is involved in the metabolism of raffinose, the above findings suggest a potential role of Aes in the regulation of carbohydrate metabolism in E. coli.