DNA Methylation Patterns of a Satellite Non-coding Sequence – FA-SAT in Cancer Cells: Its Expression Cannot Be Explained Solely by DNA Methylation

Satellite ncRNAs are emerging as key players in cell and cancer pathways. Cancer-linked satellite DNA hypomethylation seems to be responsible for the overexpression of satellite non-coding DNAs in several tumors. FA-SAT is the major satellite DNA of Felis catus and recently, its presence and transcription was described across Bilateria genomes. This satellite DNA is GC-rich and includes a CpG island, what is suggestive of transcription regulation via DNA methylation. In this work, it was studied for the first time the FA-SAT methylation profile in cat primary cells, in four passages of the cat tumor cell line FkMTp and in eight feline mammary tumors and the respective disease-free tissues. Contrary to what was expected, we found that in most of the tumor samples analyzed, FA-SAT DNA was not hypomethylated. Furthermore, in these samples the transcription of FA-SAT does not correlate with the methylation status. The use of a global demethylating agent, 5-Azacytidine, in cat primary cells caused an increase in the FA-SAT non-coding RNA levels. However, global demethylation in the tumor FkMTp cells only resulted in the increased levels of the FA-SAT small RNA fraction. Our data suggests that DNA methylation of FA-SAT is involved in the regulation of this satellite DNA, however, other mechanisms are certainly contributing to the transcriptional status of the sequence, specifically in cancer

Tocopherols as bioactive compounds in different cultivars of Gomphrena globosa L.

Tocopherols are lipid-soluble molecules that belong to the group of the Vitamin E compounds, playing an essential role on human nutrition and health [1]. The term "Vitamin E" was first introduced as an important dietary factor for animal reproduction and, more than 40 years later, it was associated to antioxidant properties. Nowadays, most of the researches focus on the fundamental chemistry that explains their antioxidant properties, specific location, role in biological membranes, and particularity on the benefits of these compounds for human health [1]. The antioxidant activity of tocopherols can be explained by two main mechanisms: first as scavengers of lipid peroxyl radicals, produced from polyunsaturated fatty adds of the membrane phospholipids and lipoproteins, through the transfer of a hydrogen atom; and second as a singlet molecular oxygen quencher, which can oxidize membrane lipids, proteins, amino acids, and nucleic acids, among others [2, 3]. In this context, our study focused on the identification and quantification of tocopherols in three different Gomphrena globosa L. cultivars (red, white and pink globe amaranth). The analysis was performed by HPLC and the quantification was based on their fluorescence signal response. White and pink globe amaranth revealed similar amounts of Υ-tocopherol (1. 04 and 1.09 mg/100 g of dry weight) and total tocopherols (1. 37 and 1.38 mg/100 g, respectively). α-Tocopherol was found in higher concentrations in red globe amaranth (0.55 mg/100 g), that also presented Υ-tocopherol (0. 50 mg/100 g), but this was the only sample where α-tocopherol was not detected. This latter isoform was found in low amounts in the other two cultivars in concentrations ranging from 0. 05 to 0.06 mg/100 g (for white and pink globe amaranth, respectively). The results obtained offer prospects for the use of these less studied cultivars of G. globosa as antioxidants, making them suitable to be included in the human diet

Antioxidant and anti-inflammatory properties of red, white and pink globe amaranth hydromethanolic extracts

Medicinal plants have been playing a vital role on human health and healing, representing one of the major sources of drugs in modem and traditional medicine [1]. Plants synthesize and preserve a variety of biochemical products that can be used as pharmaceutical compounds [2], and recently there has been an increasing interest in the therapeutic potential of plants as antioxidants and anti-inflammatories [3]. Oxidative stress and inflammation play critical roles in the pathogenesis of many diseases, such as cancer, cardiovascular disease, arthritis and obesity [4], among others. Thus, the aim of this study was to explore the bioactivity of red, white and pink globe amaranth (different cultivars of Gomphrena globosa L.) hydromethanolic extracts, namely the antioxidant and anti-inflammatory activities. The antioxidant activity was tested through radicals scavenging capacity, reducing power, and lipid peroxidation inhibition, whereas the anti-inflammatory activity was assessed by monitoring the inhibition of nitric oxide (NO) release in the mouse macrophage-like cell line RAW 264.7. The absence of toxicity of the extracts was also confirmed by the sulphorodamine B (SRB) assay applied to a porcine liver primary culture (PLP2) established by the authors. Among the three studied samples, pink globe amaranth showed the highest antioxidant activity, with the lowest ECso values (0.25 to 1.02 mg/mL), followed by red (0.41 to 1.30 mg/mL) and white (0.57 to 1.47 mg/mL) globe amaranth. Regarding the anti-inflammatory activity, pink and red globe amaranth also revealed the lowest ECso values (133 and 136 pg/mL, respectively), with white globe amaranth revealing an ECso value of 198pg/mL. None of the extracts presented cytotoxicity in PLP2 cells up to 400 pg/mL. From the results obtained, we can conclude that the extracts of these plants can be considered good sources of antioxidants and can also be used for anti-inflammatory purposes

Natural compounds: co-delivery strategies with chemotherapeutic agents or nucleic acids using lipid-based nanocarriers

Cancer is one of the leading causes of death, and latest predictions indicate that cancer- related deaths will increase over the next few decades. Despite significant advances in conventional therapies, treatments remain far from ideal due to limitations such as lack of selectivity, non-specific distribution, and multidrug resistance. Current research is focusing on the development of several strategies to improve the efficiency of chemotherapeutic agents and, as a result, overcome the challenges associated with conventional therapies. In this regard, combined therapy with natural compounds and other therapeutic agents, such as chemotherapeutics or nucleic acids, has recently emerged as a new strategy for tackling the drawbacks of conventional therapies. Taking this strategy into consideration, the co-delivery of the above-mentioned agents in lipid-based nanocarriers provides some advantages by improving the potential of the therapeutic agents carried. In this review, we present an analysis of the synergistic anticancer outcomes resulting from the combination of natural compounds and chemotherapeutics or nucleic acids. We also emphasize the importance of these co-delivery strategies when reducing multidrug resistance and adverse toxic effects. Furthermore, the review delves into the challenges and opportunities surrounding the application of these co-delivery strategies towards tangible clinical translation for cancer treatment.This work was supported by the Portuguese Foundation for Science and Technology (FCT) in the framework of the Strategic Funding UIDB/04650/2020 and UIDB/04050/2020 (CF-UM-UP and CBMA), UIDB/04046/2020 and UIDP/04046/2020 (BioISI), UIDP/04378/2020 and UIDB/04378/2020 (UCIBIO), and LA/P/0140/2020 (i4HB). E.F. thanks FCT for the PhD grant (SFRH/BD/147938/2019)

pH-responsive hybrid nanoassemblies for cancer treatment: formulation development, optimization, and in vitro therapeutic performance

Current needs for increased drug delivery carrier efficacy and specificity in cancer necessitate the adoption of intelligent materials that respond to environmental stimuli. Therefore, we developed and optimized pH-triggered drug delivery nanoassemblies that exhibit an increased release of doxorubicin (DOX) in acidic conditions typical of cancer tissues and endosomal vesicles (pH 5.5) while exhibiting significantly lower release under normal physiological conditions (pH 7.5), indicating the potential to reduce cytotoxicity in healthy cells. The hybrid (polymeric/lipid) composition of the lyotropic non-lamellar liquid crystalline (LNLCs) nanoassemblies demonstrated high encapsulation efficiency of the drug (>90%) and high drug loading content (>7%) with colloidal stability lasting at least 4 weeks. Confocal microscopy revealed cancer cellular uptake and DOX-loaded LNLCs accumulation near the nucleus of human hepatocellular carcinoma cells, with a large number of cells appearing to be in apoptosis. DOX-loaded LNLCs have also shown higher citotoxicity in cancer cell lines (MDA-MB 231 and HepG2 cell lines after 24 h and in NCI-H1299 cell line after 48 h) when compared to free drug. After 24 h, free DOX was found to have higher cytotoxicity than DOX-loaded LNLCs and empty LNLCs in the normal cell line. Overall, the results demonstrate that DOX-loaded LNLCs have the potential to be explored in cancer therapy.This research was funded by FCT/MCTES—Foundation for Science and Technology I.P. from the Minister of Science, Technology, and Higher Education (PIDDAC) and European Regional Development Fund (ERDF) by the COMPETE—Programa Operacional Factores de Competitividade (POFC) through the project CONCERT [POCI-01-0145-FEDER-032651 and PTDC/NAN-MAT/326512017]. This work was also supported by the Portuguese Foundation for Science and Technology (FCT) in the framework of the Strategic Funding UIDB/04650/2020 and UIDB/04050/2020 (CF-UM-UP and CBMA). The authors are grateful for the support of the BioISI center grant (UIDB/04046/2020 and UIDP/04046/2020), financed by FCT. R.M. acknowledges FCT I.P. for funding within the Scientific Employment Stimulus instrument (CEECIND/00526/2018)

New potential anti-hepatocellular carcinoma (HCC) agents: in vitro evaluation of anti-HCC activity and hepatotoxicity of 6-substituted methyl 3-aminothieno[3,2-b]pyridine-2-carboxylates and QSAR studies

Hepatocellular carcinoma (HCC) is a major health problem with more than 660,000 new cases per year worldwide [1]. HCC is resistant to commonly used treatments like chemotherapy and radiotherapy and new anti-HCC therapies are urgently needed. Sorafenib was the first approved small molecule against HCC and underlines the importance of identifying potential new anti-HCC drugs [2]. Thirty-two 6-substituted methyl 3-aminothieno[3,2-b]pyridine-2-carboxylates, previously prepared by some of us [3,4], were evaluated as potential new anti-HCC agents by studying their in vitro cell growth inhibition on human HepG2 cells, generally regarded as a good HCC model, and hepatotoxicity using a porcine liver primary cell culture (PLP1). The presence of amino groups linked to a benzene moiety on the substituent of the 6-position emerged as the key element for the anti-HCC activity

A comparison of the bioactivity and phytochemical profile of three different cultivars of globe amaranth: red, white, and pink

The phytochemical profiles and bioactivities of red, white and pink globe amaranth (Gomphrena haageana K., Gomphrena globosa var. albiflora and Gomphrena sp., respectively), much less studied than the purple species (G. globosa L.), were compared. The chemical characterization of the samples included the analysis of macronutrients and individual profiles of sugars, organic acids, fatty acids, tocopherols, and phenolic compounds. Their bioactivity was evaluated by determining the antioxidant and anti-inflammatory activities; the absence of cytotoxicity was also determined. Red and pink samples showed the highest sugar content. Otherwise, the white sample gave the highest level of organic acids, and together with the pink one showed the highest tocopherol and PUFA levels. Quercetin-3-O-rutinoside was the major flavonol in white and pink samples, whereas a tetrahydroxy-methylenedioxyflavone was the major compound in the red variety, which revealed a different phenolic profile. The pink globe amaranth hydromethanolic extract revealed the highest antioxidant activity, followed by those of red and white samples. The anti-inflammatory activity was more relevant in red and pink varieties. None of the samples presented toxicity in liver cells. Overall, these samples can be used in bioactive formulations against inflammatory processes and in free radical production.The authors are grateful to Foundation for Science and Technology (FCT, Portugal) for financial support to the research center CIMO (strategic project PEst-OE/AGR/UI0690/2014), R. Calhelha grant (SFRH/BPD/68344/2010) and L. Barros researcher contract under “Programa Compromisso com Ciência – 2008”