Phosphomonoester Phosphoethanolamine Induces Apoptosis in Human Chronic Myeloid Leukemia Cells

Background:Leukemia is a type of cancer that starts in the blood or blood-forming tissues.It results from the clonal proliferation of hematopoietic cells in the bone marrow and/or lymphoid tissues,which subsequently reach the peripheral circulation and can infiltrate other systems.There are many different kinds of leukemia,and treatments are different for each one.Chronic leukemia is with a slower growing than acute leukemia but could be just as life-threatening.Phospholipids are antitumor analogs,such as synthetic phosphoethanolamine,which is a phosphorylated compound capable of controlling cellular proliferation and inducing apoptosis in several types of tumor cells.Methods:K562 and K562-Lucena(MDR+)human chronic myeloid leukemia cells were treated with synthetic phosphoethanolamine(Pho-s).The viability was evaluated by sulforhodamine B(SRB)assay and cell cycle phases,apoptosis,markers expression,and mitochondrial potential were assessed by flow cytometry.Results:Tumor cells formed clusters in suspension and decreased significantly viability.The concentrations for IC50%were obtained.Pho-s treated were 43.1 mM(K562)and 145.9 mM(K562-Lucena MDR+)in a period of 24 hours.Pho-s induced changes in the distribution of cell population phases of cell cycle which showed an increase in fragmented DNA and increased markers expression envolved apoptosis pathways a decrease in the G1/G0 phase.Discussion:Treatment of K562 and K562-Lucena(MDR+)chronic myeloid leukemia cells with Pho-s showed dose and time dependent cytotoxic effects.This cytotoxicity induced a decrease in proliferative capacity,mitochondrial electrical potential,and consequently release of cytochrome C;inhibition of Bcl-2 family protein expression,increase in pro-apoptotic family members Bad and Bax,dependent on p53 expression.Conclusion:This study presented a significant therapeutic potential of Phos-s in this type of leukemia through the apoptotic effects on tumor cells independently of the molecular resistance profile(MDR+).

Meclizine Chloridrate and Methyl-β-Cyclodextrin Associated with Monophosphoester Synthetic Phosphoethanolamine Modulating Proliferative Potential in Triple-Negative Breast Cancer Cells

Synthetic phosphoethanolamine(Pho-s)is a monophosphoester ester with anti-inflammatory and pro-apoptotic properties.Meclizine chloridrate(MC)is a histamine H1 receptor blocker that is also able to inhibit cellular respiration.However,MC does not inhibit cellular respiration in isolated mitochondria such as antimycin and rotenone.Methyl-β-cyclodextrin(MβCD)belongs to theβ-cyclodextrin family,which is capable of removing cholesterol from the plasma membrane.The aim of this study was to evaluate the proliferative effects of meclizine chloridrate and methyl-β-cyclodextrin compounds associated with synthetic phosphoethanolamine in a triple-negative human breast tumor line,MDA-MB-231 Cell viability of the tumor line and normal cells FN1 was evaluated by MTT colorimetric test;the production of free radicals was determined by lipoperoxidation(LPO)test;and the percentage of cell cycle phases and proliferative index was evaluated by flow cytometry.Cell viability demonstrated a significant decrease with the treatments of MβCD,MC and Pho-s associated with MC.The production of free radicals decreases significantly in all treatments.In addition,a significant increase of DNA fragment and decrease in G0/G1 cell cycle phase were observed in cellular percentage with concentrations of 20 and 30 mM of Pho-s in association with MC and MβCD,respectively.

2-aminoethyl Dihydrogen Phosphate as a Modulator of Proliferative and Apoptotic Effects in Breast Cancer Cell Lines

Background: Breast cancer is a type of cancer that affects more women throughout the world, in developing anddeveloped countries. 2-AEH2P is a phospholipid analog of cellular membrane, which makes it different from existing molecules fortheir absorption, stability and display anti-inflammatory, anti-proliferative and pro-apoptotic properties. Methods: MCF-7 humanbreast adenocarcinoma cells were treated with 2-AEH2P. The viability and adhesion cells were evaluated by MTT assay. Cell cyclephases, apoptosis, markers and mitochondrial potential were assessed by flow cytometry. Morphological ultrastructural analyzeswere performed by laser confocal microscopy. Results: MCF-7 Tumor cells acquired round shapes, lost cytoplasmic expansions,formed clusters in suspension and decreased significantly viability. There were changes in the morphology, membrane fragmentationand loss of cytoplasmic projection. The obtained concentrations for IC50% were 37.2;25.8;1.8 mM for periods of 24, 48 and 72 h,respectively. Changes in the distribution of cell population phases of the cell cycle showed an increase in fragmented DNA and anincrease in the G2/M phase. The expression β-gal showed proliferative reduction induced by 2-AEH2P. Laser confocal microscopyshowed changes in the mitochondrial membrane and alteration in distribution. Proliferative index of MCF-7 tumor cells treated with2-AEH2P decreased significantly when compared to fibroblast normal cells. The compound 2-AEH2P is a phospholipid withantiproliferative potential and apoptosis modulator.

Antitumor Effect of Cationic INKKI Peptide from Bovine <i>β</i>-Casein on Melanoma B16F10

Cationic peptide with the sequence INKKI 41-45 was isolated from bovine β-casein after tryptic hydrolysis and synthetized. The aim of this work was to evaluate the antiproliferative activity in vitro and antitumor effect in animal model. The in vitro cytotoxicity was evaluated on B16F10 melanoma cells by MTT assay. Detection of apoptosis was measured using the annexin V/PI double staining and cell cycle analysis performed flow cytometry. Caspase-3 activity was analyzed with substrate specific fluorogenic DEVD-MCA. In vivo, antitumor activity was evaluated in B16F10 melanoma tumor-bearing C57BL/6J mice. The animals were treated with 55 mg/kg INKKI administered into peritumoral region, while control group received saline solution. The following antitumor parameters were examined: tumor volume, number of metastases, tumor delayed time, tumor doubling time. Histological analyses were performed with H & E staining. The results showed that INKKI induced dose-response cytotoxicity selective for B16F10 melanoma cells (IC50 1.7 μM) and did not present cytotoxic effects for FN1 fibroblast cells. INKKI-induced apoptosis detected trough of annexin V/PI assay and it was accompanied with an increase of sub-G1 apoptotic fractions and significant increase of caspase-3 cleavage. The tumor-bearing mice treated with INKKI showed a significant reduction in tumor volume of 72.62% and decreased of metastasis number loci. In addition, INKKI caused a significant delay in tumor growth and prolonged the tumor doubling time. Histological analysis revealed an increased of necrosis areas and reduction of tumor cells in tumor treated with INKKI, it was a many hallmark of its antitumor effects observed from in vivo experiments. In conclusion, we show that INKKI is a peptide that could be considered a new putative candidate development to anticancer therapy drug.

Response Proliferative Capacity of Undifferentiated Stem Cells of Obtained Human Adult Dental Follicle

Objective: The aim of this study was correlation proliferative activity, markers express stem cells, and lipid peroxides of undifferentiated stem cells of human adult dental follicle (DF) following culture. Methods: For this study, we used 8 samples from DF of impacted third molars to maintain culture conditions and evaluated the growth curve, cell viability, production of lipid peroxidation, cell cycle phases, and proliferative index during 25 days of culture. Results: Cells after culture showed characteristics of fibroblast-like type following 25th day of culture. The results of lipid peroxidation showed that stem cells in culture produce 13 nmoles/ml malondialdehyde at the start of culture, increasing until the 12th day and then began a decline that lasted until the 25th day. We revealed that DFSCs presented a significantly higher percentage of cells in S + G2/M phases by the 15th day of culture compared with cells at the start of culture. Cell surface markers revealed that cell lines were negative for HLA-DR and positive for CD90, CD44, and CD105. The expression of p21 protein, involved in the regulation of the cell cycle, showed a significant increase from the 15th to 25th day of culture. Results of cell division rates show a significant increase between the 6th and 15th day of culture. Conclusions: We conclude that the culture remained stable during the 25 days of culture, presenting the markers of stem cells and markers of control, progression, and cell proliferation that there was an increased production of lipid peroxides between the 6th and 12th days;this increase is related to the increased numbers of cells that also occurs during this period. Then, there is a significantly decline in the production of lipid peroxides and the number of cells, which is accompanied by an increase in cell unviability.

The Antiproliferative and Pro-apoptotic Role of 2-aminoethyl dihydrogen Phosphate in Triple-negative Breast Tumor Cells

Antineoplastic phospholipids are a new class of antitumor agents.These molecules interact with the plasma membrane,changing numerous pathways that induce cell death,with high selectivity for cancer cells.A representative of this class of antineoplastic agents is 2-aminoethyl dihydrogen phosphate(2-AEH2P).It is present in high intracellular concentrations in various tissues and organelles with antitumor,antiproliferative and pro-apoptotic action.Therefore,4T1 triple-negative tumor cells were treated in different concentrations in order to assess the cytotoxic potential and its effects on the modulation of cell death pathways in association with the chemotherapeutic drug Paclitaxel.2-AEH2P promoted cytotoxicity in tumor cells and significant morphological changes,however,it did not cause these effects in normal cells.There was positive regulation of proteins involved in the intrinsic pathway of cell death by apoptosis and regulation of the phases of cell cycle progression.Furthermore,structural and distribution changes in mitochondria,as well as decreased cell density and regression of the cytoskeleton were observed.The 2-AEH2P demonstrated a modulatory potential of apoptotic pathways inducing cell death,being a new compound with antitumor properties.

Clinical Oncology Translational Study Phase-1 with Antitumor Phosphorylethanolamine(2-AEH_(2)P)in Dogs with Neoplasms

The animal has the potential to serve as a clinical model for human disease,due to striking similarities and homologies in diseases.Early clinical development of a new drug may influence its final destination,and a careful and thoughtful approach to Phase-1 clinical trials is essential.Phosphorylethanolamine(2-AEH2P)is a new cytostatic phospholipid agent that,unlike most chemotherapeutic drugs used today,does not target DNA or cytoskeleton,but act on the cell membrane.Studies carried out by our group have shown several effects of 2-AEH2P controlling the progression of the cell cycle,in the alterations of the mitochondrial electrical potential capable of inducing apoptosis and autophagy in several types of human and animal tumor cells.The aim of this study is to establish the recommended safety dose of a 2-AEH2P monophosphoester,in dogs with tumor following the Phase 1 study model proposed by Fibonacci.The Phase-1 study following the Fibonacci model,2-AEH2P was safe at all staggered doses up to 150 mg/Kg for 8 weeks.Intravenous administration in staggered doses according to the Fibonacci model,showed that the drug is safe,no advent of mortality during the study period or acute toxicity were observed.It is not a drug with hemolytic properties or that induces anemia.It does not lead to changes in liver and renal functions and was able to modulate leukocyte production.2-AEH2P is new compound with antitumor potential,being useful for future veterinary and human tumors,as a combination of chemotherapeutic agents.

Tree-Fall Gaps Effects on Spider(Araneae)Assemblages in an Atlantic Forest Landscape in Northeastern Brazil

The study investigates the effect of spatial and temporal tree-fall gaps structure on spiders’ assemblage in an Atlantic forest fragmented area in Brazil. It was conducted in the Michelin Ecological Preserve-REM (Bahia), 190 ha forest remnant. Samples were collected on leaf-litter (50 × 50 cm) at five tree-fall gaps formations (2), within five adjacent primary forest and five inner edge parcels. During 16 months (between May 2009 and October 2012), 480 m2 leaf-litter samples were collected, from which spiders were extracted using mini-Winkler traps. The observed and estimated richness of spider’s species was higher at the edge

An overview of the marine natural products in clinical trials and on the market

The first marine natural products that served as leads or scaffolds for medicines were discovered in the middle of last century:the arabinosyl glycosides from the marine sponge Tectitethya crypta.Synthesis and modifications of the natural molecules generated antiviral and antileukemic drugs developed in the 1970’s and in the following decades,including the first effective treatment against HIV infection.With the improvement of techniques for the elucidation of chemical structure of the molecules,as well as chemical synthesis,especially from the 1990’s,there was an increase in the number of bioactive natural products characterized from marine organisms.New chemical structures with high specificity towards molecular targets in cells allowed the development of new drugs with indication for the treatment of several illnesses,from cancer to new antibiotics,and even neurological disorders.Currently there are at least 13 molecules derived from marine natural products on advanced clinical trials,and nine were approved to be used as medicines.Considering that in the past eight years,more than 1000 new compounds from marine organisms were described,per year,the expectation is that many more drugs will be derived from marine natural products in a near future.

Serological evidence of Toxoplasma gondii infection as potential risk for the development of lepromatous leprosy in an endemic area for both neglected tropical diseases in Brazil

Background:Mycobacterium leprae and Toxoplasma gondii infections are both neglected tropical diseases highly prevalent in Brazil.Infection with certain parasite species can significantly alter susceptibility to other important pathogens,and/or influence the development of pathology.Here we investigated the possible influence of M.leprae/T.gondii co-parasitism on the manifestation of leprosy and its clinical forms.Methods:Participants(n=291)were recruited in Campos dos Goytacazes city,Rio de Janeiro state,southeast Brazil,from August 2015 to December 2019 and clinically diagnosed for leprosy.Participants were selected based on the presence(patients)or absence(healthy controls)of the leprosy disease.Contacts of patients were also recruited for this study.Serum samples from patients(n=199)with leprosy,contacts(n=40)and healthy controls(n=52)were investigated for levels of IgM and IgG anti-phenolic glycolipid-1(PGL-1)by ELISA.Additionally,IgG antibody against soluble Toxoplasma antigen(STAg)was measured in sera samples from leprosy patients,contacts and healthy controls for Toxoplasma gondii serology by ELISA.Anti-PGL-1 IgG and IgM levels were compared using one-way ANOVA Kruskal-Wallis or Mann-Whitney,while Spearman test was used to correlate levels of IgG anti-STAg and IgM/IgG anti-PGL-1 from seropositive and seronegative individuals for T.gondii infection.The risk of T.gondii infection for leprosy disease was assessed using Fisher’s test.Results:Levels of IgM anti-PGL-1 antibodies were significantly higher in multibacillary(MB)patients compared to paucibacillary(PB)patients(P=0.0068).Higher IgM and IgG levels anti-PGL-1 were detected in patients with the lepromatous forms.The serologic prevalence for T.gondii infection was 74.9%.We detected increased anti-STAg antibody levels in leprosy patients(79.4%),reaching 88.8%within those with lepromatous form of this disease.The leprosy risk increase in T.gondii seropositive individuals was two-fold(odds ratio[OR]=2.055;95% confidence intervals[95%CI]:1.18–3.51)higher than those seronegative,and considering the lepromatous leprosy risk this increase was even dramatic(OR=4.33;95%CI:1.76–9.69)in T.gondii seropositive individuals.Moreover the leprosy risk in T.gondii seropositive individuals was weakly correlated to the levels of IgG anti-STAg and IgM/IgG anti-PGL-1.Conclusions:Altogether,our results suggest that T.gondii infection may exert immunomodulatory properties that influence to the susceptibility of leprosy,mainly on its more severe clinical form.A better understanding of parasite immunomodulation can ultimately contribute to the development of medical applications.

Reproduction ecology of the recently invasive snake Hemorrhois hippocrepis on the island of Ilbiza Elba

Knowing the causes of biological invasion success can be relevant to combat future invasive processes.The recent invasion of the horseshoe whip snake Hemorrhois hippocrepis on the island of Ibiza provides the opportunity to compare natural history traits between invasive and source populations,and to unravel what makes this snake a successful invader that is threatening the only endemic vertebrate of the island,Podarcis pityusensis.This study compares the basic reproductive traits of mainland native and invasive populations of the snake.Our results revealed that invasive populations were characterized by female maturity at a smaller size,extended reproductive period,and much lower reproduction frequency compared to the native population.In contrast,some major reproductive traits--the abdominal fat body cycle,clutch size,hatchling body size,and hatchling body condition,did not differ between the two populations.Some of these results must reflect the environmental differences in the recently invaded island with respect to the source area,and overall plasticity of reproductive traits.Plasticity is evolutionarily interesting,and may aid the succesful growth of this species in their invasiveness of Mediterranean islands like Ibiza.The most significant finding is that this expression of phenotypic plasticity occurred rapidly in this invasive population,within a period of 14 years maximum.Our results on the reproduction ecology of the invasive population were not conclusive regarding the factors determining the invasiveness of the snake and pointed to alternative causes.