Human Blood Cancer Research Articles

Cytotoxicity effects of leaf extracts of Ciplukan (Physalis angulata; Solanaceae) on human blood and ovary cancer cell lines

Physalis angulata has identified by molecular phylogenetic analysis and the results showed genetically close related with an anticancer plant, Withania somnifera. The aims of this preliminary study was to evaluate cytotoxicity effects of leaf extract of P. angulata (LEP) either in powder or pasta form on cells viability, cells proliferation, and inhibition on human ovary cancer cell lines (SKOV3) and human blood cancer cell lines (HL60). Cytotoxicity effects was analysed by MTS Cell Proliferation Assay Kit. Doxorubicin was used as positive control in this experiment. Surprisingly, two treatments (powder and pasta) have different toxic effects. We found that IC50 and viability (LC50) for SKOV3 cell lines for powder was between 93 ug/ml and 187 ug/ml, and 187 ug/ml and 375 ug/ml for pasta. IC50 for HL60 cell lines was 23 ug/ml for powder and around 18 ug/ml for pasta. Cell viability (LC50) of HL60 treated with LEP showed that 23 ug/ml for powder or 46 ug/ml for pasta. These results suggested that LEP have antiproliferative and inhibition activities both on SKOV3 or HL60.

Copper chaperone ATOX1 is required for MAPK signaling and growth in BRAF mutation-positive melanoma

Copper (Cu) is a tightly regulated micronutrient that functions as a structural or catalytic cofactor for specific proteins essential for a diverse array of biological processes. While the study of the extremely rare genetic diseases, Menkes and Wilson, has highlighted the requirement for proper Cu acquisition and elimination in biological systems for cellular growth and proliferation, the importance of dedicated Cu transport systems, like the Cu chaperones ATOX1 and CCS, in the pathophysiology of cancer is not well defined. We found that ATOX1 was significantly overexpressed in human blood, breast, and skin cancer samples, while CCS was significantly altered in human brain, liver, ovarian, and prostate cancer when compared to normal tissue. Further analysis of genetic expression data in Cancer Cell Line Encyclopedia (CCLE) revealed that ATOX1 is highly expressed in melanoma cell lines over other cancer cell lines. We previously found that Cu is required for BRAFV600E-driven MAPK signaling and melanomagenesis. Here we show that genetic loss of ATOX1 decreased BRAFV600E-dependent growth and signaling in human melanoma cell lines. Pharmacological inhibition of ATOX1 with a small molecule, DCAC50, decreased the phosphorylation of ERK1/2 and reduced the growth of BRAF mutation-positive melanoma cell lines in a dose-dependent manner. Taken together, these results suggest that targeting the Cu chaperone ATOX1 as a novel therapeutic angle in BRAFV600E-driven melanomas.

Effect of CXCR9-CXCR3 Cytokine Signaling Pathway in Polycytemia Vera

<strong>Objective</strong>: Myeloproliferative neoplasms (MPNs) are among the most common myeloproliferative disorders. MPNs are characterized by excessive production of terminally differentiated blood cells. The expression of CXCR3, responsible for regulating the proliferation of healthy hematopoietic stem cells, is known to be induced by granulocyte-macrophage colony stimulating factor (GM-CSF). The aim of this study is to investigate the role of CXCL9-CXCR3 signaling pathway in the progression of MPN. <strong>Material and Method</strong>: We determined the expression of CXCL9 and two isoforms of the CXCR3 receptor (CXCR3A and CXCR3B) on the surface of human peripheral blood mononuclear cells (PBMC) and cancer stem cells. Real-time polymerase chain reaction (qPCR) was used to measure expression levels, and flow cytometry was used to examine the presence of cell surface receptors. In addition, qPCR was used to quantify CXCR3 expression after GM-CSF application in cell culture. In PBMC and CD34+ cells, the mRNA level of CXCR3A expression was found to be elevated in patients with MPN. <strong>Results</strong>: There was no statistically significant difference in mRNA expression of CXCR3B and CXCL9 between patients and healthy controls.There was significant reduction in cell surface expression of the CXCR3 receptor in PBMC obtained from patients. T <strong>Conclusion</strong>: hus, these results indicate that imbalance in the expression of CXCR3A/CXCR3B isoforms and chemokines CXCL9, CXCL10 and CXCL11 that bind to these receptors, may mediate inflammation and cancer progression in MPN.

ANALYSIS OF FATTY ACIDS FROM OIL OF GREEN TEA (CAMELLIA SINENSIS L) BY GAS CHROMATOGRAPHY COUPLED WITH FLAME IONIZATION DETECTOR AND ITS ANTICANCER AND ANTIBACTERIAL ACTIVITY IN VITRO

Objective: Tea is a widely consumed beverage worldwide. The effect of green tea is mainly due to its high polyphenols-(-) epigallocatechin-3-gallate (EGCG) content in the culture of cancer cell and bacterial cells. The present work was carried out to investigate the efficacy of green tea oil (GTO) against cancer cells and bacterial cells.&#x0D; Methods: In this study green tea oil was prepared from green tea for different experiment and determination of fatty acids profile from green tea oil. In the present study, peripheral blood lymphocyte (PBL) was chosen as human peripheral blood lymphocytes and blood cancer MCF-7 cells were chosen as human cancer cells. To fulfill our aims and also to evaluate the activity of this phytomedicine against normal lymphocytes and cancer cells the cell samples were divided into 26 experimental groups in the following ways. Each Petri dish contains 2 X 105 cells.&#x0D; Results: GTO shows a potent anticancer agent but nontoxic to normal cells. The GTO decreases the reduced glutathione (GSH) level and increase the oxidized glutathione (GSSG) level significantly (P&lt;0.05) in MCF-7 cells. But in lymphocytes the GSH level and GSSG level were almost the same with the control group but doxorubicin (DOX) significantly decreased the GSH and increase the GSSG level. Green tea oil treatment causes generation of reactive oxygen species (ROS) in MCF-7 cells revealed by DCFH2DA staining. Agar diffusion test shows the GTO is effective against multi-drug resistant bacteria.&#x0D; Conclusion: This phytomedicine has a potent anticancer activity without damaging the normal lymphocytes. So, this drug can be used for further treatment of anticancer and antibacterial.

Evaluating the effect of anti–cancer nano drugs dosage and reduced leukemia and polycythemia vera levels on trend of the human blood and bone marrow cancers under synchrotron radiation

Leukemia is a cancer of the blood and bone marrow; there are many types of leukemia, depending on which type of white blood cell is involved. Treatment varies with the type and stage of disease. In fact, it is a blood cancer that begins in the marrow of our bones, the soft center where new blood cells grow. If we have polycythemia vera, our marrow makes too many red blood cells, which causes our blood to get too thick. That can make us more likely to have clots, a stroke, or a heart attack. Treatment of leukemia and polycythemia vera levels on trend of the human blood and bone marrow cancers for optimum usage of anti–cancer Nano drugs dosage is one of the most important issues of sustainable development. Therefore, the only available way to deal with the problem of treatment of leukemia and polycythemia vera levels on trend of the human blood and bone marrow cancers is optimum usage of anti-cancer Nano drugs dosage. In this study, we evaluated the chemical changes of anti–cancer Nano drugs dosage in human blood and bone marrow cancers’ cells such as leukemia, a type of cancer found in our blood and bone marrow cancers is caused by the rapid production of abnormal white blood cells. The high number of abnormal white blood cells is not able to fight infection, and they impair the ability of the bone marrow cancers to produce red blood cells and platelets and determine the treatment trend of these changes.

Automatic Identification of Acute Lymphoblastic Leukemia on Blood Cell An image Using Geometric Features

Acute Lymphoblastic Leukemia (ALL) is a human blood cancer which causes most deaths among other types of blood cancer. It commonly occurs in children and teenagers. ALL is recognised when there is a presence of lymphoblast cell in human blood. Haematologists diagnose ALL using visual microscopic observation from a human periphery blood sample. The inspection takes a long time and is limited to the subjective experience of the haematologist, and therefore, causes differences in the diagnostic results among haematologists. In this study, the researchers proposed automatic ALL Identification system to solve the problem. The method used an image processing technique that analysed a human blood cells morphology. The system identified the differences between normal white blood cells and lymphoblast cells by considering the shape and the size of the nucleus and also the cytoplasm. There are 33 microscopic an image data of a human blood cell that were used in this research. Geometric features used in the study were an area, perimeter, eccentricity, equivDiameter, solidity, roundness and circularity of a human blood the nucleus and cytoplasm and also the ratio of area and the perimeter of the cells and the nuclei. Firstly, the nucleus and cytoplasm were extracted from the other component of a human blood cell using Gram-Schmidt as an orthogonalisation-based segmentation method and then using Otsu threshold. Support Vector Machine was then used to classify whether the white blood cells were lymphoblasts or normal cells. Lastly, ALL identification was evaluated by the parameter of sensitivity and misclassification rate metrics using k-fold cross-validation.

Anti-cancer Parasporin Toxins of New Bacillus thuringiensis Against Human Colon (HCT-116) and Blood (CCRF-CEM) Cancer Cell Lines.

Bacillus thuringiensis is one of the most important microorganisms used against cancer cell lines in latest studies all over the world. This study aims to perform the isolation, molecular identification, and to identify novel B. thuringiensis strains that specifically targeting human cancer cell-killing activities in Iran. A total of 88 B. thuringiensis isolates were recovered from Iran. Upon the treatment of the non-hemolytic crystal proteins by proteinase K, five isolates belonging to three biotypes, thuringiensis, kurstaki and sotto of B. thuringiensis are found to have different cytotoxicity toward HCT-116 and CCRF-CEM cell lines. Digested inclusions of the isolates consisted of one major poly peptide of 34-kDa, as estimated by sodium dodecyl-sulfate polyacrylamide gel electrophoresis. The structure, molecular identification, and functionality of five isolates inclusion proteins have shown to be closely like to parasporin-2 but their size of activated protein is not similar to this parasporin. It is unclear that discovered damaging proteins are parasporin-2. This 34-kD protein exhibited varying degrees of cytocidal activity toward human colon and blood cancer cells and caused cell swelling and the formation of blebs in the surface of the cells or alteration in cytoskeleton. The soil in the humid and temperate climates of Iran is a good reservoir for parasporin producing B. thuringiensis. The isolated B. thuringiensis strains exhibit specific and different cytocidal activities against human colon and blood cancer cells. Parasporin is a novel cytotoxic protein to human cancer cells produced by B. thuringiensis and these toxins appeared to attack an identical target on human cancer cells.

Detection and proteomic characterization of extracellular vesicles in human pancreatic juice

AimsThe prognosis of patients with pancreatic cancer has remained virtually unchanged with a high mortality rate compared to other types of cancers. An earlier detection would provide a time window of opportunity for treatment and prevention of deaths.In the present study we investigated extracellular vesicle (EV)-associated potential biomarkers for pancreatic cancer by directly assessing EV size-based subpopulations in pancreatic juice samples of patients with chronic pancreatitis or pancreatic cancer. In addition, we also studied blood plasma and pancreatic cancer cell line-derived EVs. MethodsComparative proteomic analysis was performed of 102 EV preparations from human pancreatic juices, blood, and pancreatic cancer cell lines Capan-1 and MIA PaCa-2. EV preparations were also characterized by electron microscopy, tunable resistive pulse sensing, and flow cytometry. ResultsHere we describe the presence of EVs in human pancreatic juice samples. Pancreatic juice EV-associated proteins that we identified as possible candidate markers for pancreatic cancer included mucins, such as MUC1, MUC4, MUC5AC, MUC6 and MUC16, CFTR, and MDR1 proteins. These candidate biomarkers could also be detected by flow cytometry in EVs found in pancreatic juice and those secreted by pancreatic cancer cell lines. ConclusionsTogether our data show that detection and characterization of EVs directly in pancreatic juice is feasible and may prove to be a valuable source of potential biomarkers of pancreatic cancer.

Translocation Breakpoints Preferentially Occur in Euchromatin and Acrocentric Chromosomes.

Chromosomal translocations drive the development of many hematological and some solid cancers. Several factors have been identified to explain the non-random occurrence of translocation breakpoints in the genome. These include chromatin density, gene density and CCCTC-binding factor (CTCF)/cohesin binding site density. However, such factors are at least partially interdependent. Using 13,844 and 1563 karyotypes from human blood and solid cancers, respectively, our multiple regression analysis only identified chromatin density as the primary statistically significant predictor. Specifically, translocation breakpoints preferentially occur in open chromatin. Also, blood and solid tumors show markedly distinct translocation signatures. Strikingly, translocation breakpoints occur significantly more frequently in acrocentric chromosomes than in non-acrocentric chromosomes. Thus, translocations are probably often generated around nucleoli in the inner nucleoplasm, away from the nuclear envelope. Importantly, our findings remain true both in multivariate analyses and after removal of highly recurrent translocations. Finally, we applied pairwise probabilistic co-occurrence modeling. In addition to well-known highly prevalent translocations, such as those resulting in BCR-ABL1 (BCR-ABL) and RUNX1-RUNX1T1 (AML1-ETO) fusion genes, we identified significantly underrepresented translocations with putative fusion genes, which are probably subject to strong negative selection during tumor evolution. Taken together, our findings provide novel insights into the generation and selection of translocations during cancer development.

Novel and Transcendental Prevention, Diagnosis and Treatment Strategies for Investigation of Interaction among Human Blood Cancer Cells, Tissues, Tumors and Metastases with Synchrotron Radiation under Anti-Cancer Nano Drugs Delivery Efficacy Using MATLAB Modeling and Simulation

Novel and Transcendental Prevention, Diagnosis and Treatment Strategies for Investigation of Interaction among Human Blood Cancer Cells, Tissues, Tumors and Metastases with Synchrotron Radiation under Anti-Cancer Nano Drugs Delivery Efficacy Using MATLAB Modeling and Simulation

An Introduction to Erythropoiesis Approaches.

Many experimental models have been used to study erythropoiesis. Even prior to the advent of the genetic manipulation of animal models, erythropoiesis was examined in the mouse, chicken, sheep, goat, and rabbit, among other vertebrates. Erythroid cell lines derived from human blood cancers were also very useful, as they could be genetically manipulated more easily than whole animals. Genetic models in the mouse, zebrafish, and frog have provided a plethora of information advancing our understanding of erythropoiesis, and remain gold standards in the field for studies of hemoglobin switching, and experiments to study authentic blood cell development. Mouse and human embryonic stem (ES) and induced pluripotent (iPS) cells can be differentiated to erythroid cells in culture, though their use is somewhat limited by their propensity to express only the embryonic and fetal globin genes. Some very useful cell lines have been developed by manipulating ES or fetal liver erythroid progenitor cells from knockout mouse models. In recent years, our understanding of erythropoiesis has improved, due to the ability to knock down genes in native human hematopoietic stem and progenitor cells derived from umbilical cord blood or bone marrow, and differentiate them ex vivo to the erythroid lineage. These native cells, and cell lines derived from them, are now providing essential information about human erythropoiesis, which is complementary to that obtained from animal studies. This review provides some perspective about the cell and animal models used to study erythropoiesis over the years.

Contextual tumor suppressor function of T cell death-associated gene 8 (TDAG8) in hematological malignancies

BackgroundExtracellular acidosis is a condition found within the tumor microenvironment due to inadequate blood perfusion, hypoxia, and altered tumor cell metabolism. Acidosis has pleiotropic effects on malignant progression; therefore it is essential to understand how acidosis exerts its diverse effects. TDAG8 is a proton-sensing G-protein-coupled receptor that can be activated by extracellular acidosis.MethodsTDAG8 gene expression was analyzed by bioinformatic analyses and quantitative RT-PCR in human hematological malignancies. Retroviral transduction was used to restore TDAG8 expression in U937, Ramos and other blood cancer cells. Multiple in vitro and in vivo tumorigenesis and metastasis assays were employed to evaluate the effects of TDAG8 expression on blood cancer progression. Western blotting, immunohistochemistry and biochemical approaches were applied to elucidate the underlying mechanisms associated with the TDAG8 receptor pathway.ResultsTDAG8 expression is significantly reduced in human blood cancers in comparison to normal blood cells. Severe acidosis, pH 6.4, inhibited U937 cancer cell proliferation while mild acidosis, pH 6.9, stimulated its proliferation. However, restoring TDAG8 gene expression modulated the U937 cell response to mild extracellular acidosis and physiological pH by reducing cell proliferation. Tumor xenograft experiments further revealed that restoring TDAG8 expression in U937 and Ramos cancer cells reduced tumor growth. It was also shown U937 cells with restored TDAG8 expression attached less to Matrigel, migrated slower toward a chemoattractant, and metastasized less in severe combined immunodeficient mice. These effects correlated with a reduction in c-myc oncogene expression. The mechanistic investigation indicated that Gα13/Rho signaling arbitrated the TDAG8-mediated c-myc oncogene repression in response to acidosis.ConclusionsThis study provides data to support the concept that TDAG8 functions as a contextual tumor suppressor down-regulated in hematological malignancies and potentiation of the TDAG8 receptor pathway may be explored as a potential anti-tumorigenic approach in blood cancers.

BAMClipper: removing primers from alignments to minimize false-negative mutations in amplicon next-generation sequencing

Amplicon-based next-generation sequencing (NGS) has been widely adopted for genetic variation detection in human and other organisms. Conventional data analysis paradigm includes primer trimming before read mapping. Here we introduce BAMClipper that removes primer sequences after mapping original sequencing reads by soft-clipping SAM/BAM alignments. Mutation detection accuracy was affected by the choice of primer handling approach based on real NGS datasets of 7 human peripheral blood or breast cancer tissue samples with known BRCA1/BRCA2 mutations and >130000 simulated NGS datasets with unique mutations. BAMClipper approach detected a BRCA1 deletion (c.1620_1636del) that was otherwise missed due to edge effect. Simulation showed high false-negative rate when primers were perfectly trimmed as in conventional practice. Among the other 6 samples, variant allele frequencies of 5 BRCA1/BRCA2 mutations (indel or single-nucleotide variants) were diluted by apparently wild-type primer sequences from an overlapping amplicon (17 to 82% under-estimation). BAMClipper was robust in both situations and all 7 mutations were detected. When compared with Cutadapt, BAMClipper was faster and maintained equally high primer removal effectiveness. BAMClipper is implemented in Perl and is available under an open source MIT license at https://github.com/tommyau/bamclipper.

Hyperthermia enhances bortezomib-induced apoptosis in human white blood cancer cells

At present, the current therapeutic strategy for apoptosis induction mainly relies on the administration of pharmacological apoptotic modulators. Apart from that, apoptosis can be induced by various external stimuli such as hyperthermia, ionizing radiation, and electric fields. Despite advantages, both physical and pharmacological approaches bear some limitations as well. The rationale of this study was to overcome the limitations by combining hyperthermia and apoptotic modulator ‘bortezomib’ (Velcade). Two types of human blood cancer cell lines were utilized: human leukemic monocyte lymphoma cell U937 line and peripheral blood mononuclear cells (PMBCs) derived from the patient diagnosed with acute myeloid leukemia. Prior to apoptosis experiments, cytotoxicity tests were performed at three types of temperature regimes (40°, 42° and 44°C). We observed a gradual inhibition of cell viability correlating with an increase of temperature and drug concentration in both cell lines. However, there was no significant difference between sham group and groups of leukemic PMBCs treated by high temperature (44°C) and bortezomib. In U937 cells, combined treatment by heat shock and bortezomib led to an increase the number of cells underwent the late apoptosis stage. At the same time, similar treatment of PMBCs resulted in the stimulation of early apoptosis. Our data suggest that combination of bortezomib and hyperthermia enhances apoptosis induction in human cancer white blood cells, indicating a therapeutic potential for blood cancer therapy.

Purification and characterization of a peptide from soybean with cancer cell proliferation inhibition

A peptide from protein hydrolysate fraction obtained from a high oleic acid soybean line (N98-4445A) that had shown significant activity against human cancer cell lines was identified and purified. Three peptides showing highest activity were identified by reverse phase HPLC in the 10–50 kDa fraction of the protein and purified using peptide specific affinity chromatography column. The three individual peptides were tested for anti-proliferative activity against blood, colon and liver cancer cell lines using 3-(4,5-dimethyl thiazole-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) cell titer assay. Enhanced colon cancer cell inhibition (80%) was observed after testing a pure peptide (molecular size of 18 kDa with 158 amino acid residues) which also demonstrated a time-response of 96 hr after incubation based on trypan blue dye exclusion study. The impact of this study lies in deriving a pure single peptide with anti-proliferative activity on human colon and blood cancer cells. Practical applications Soybean is cultivated in the United States mostly for extracting the oil leaving the residue (soybean meal) which is commonly used as an animal feed due to its high protein content. This study has shown that soy peptides produced enzymatically from the meal of a high oleic acid soybean line (N98-4445A) were found to have anticancer activity. This study demonstrated an impending value for a pure peptide derived from this soy peptides as an alternative and inexpensive anti-cancer therapeutic agent as well as a value addition to soy meal by-product.

Soybean peptide fractions inhibit human blood, breast and prostate cancer cell proliferation.

In this study, we examined in vitro the bio-activity of peptide fractions obtained from soybeans against blood (CCRF-CEM and Kasumi-3), breast (MCF-7), and prostate (PC-3) cancer cell proliferation. Gastro-intestinal treated peptide fractions (<5, 5-10 and 10-50kDa) prepared from seed proteins of two high oleic acid soybean lines-N98-4445A, S03-543CR and one high protein line-R95-1705, were tested for anticancer activity against human breast, blood and prostate cancer cell lines. Anti-proliferative cell titer assay was conducted to assess the inhibitory effects of the peptide fractions, while trypan blue dye exclusion assay was used to determine the dose response of most effective fractions. Results showed that the peptide fractions inhibited the cancer cell lines up to 68.0% and the minimum concentration to get 50% inhibitory activity (IC50) ranged between 608 and 678µg/mL. This multiple site in vitro cancer inhibition by GI friendly peptides could have the potential use as food ingredients or nutritional supplements in an alternative cancer therapy.

P156: Detection adducts DNA in human blood and lung cancer tissue by a hybrid quadrupole time-of-flight (Q-TOF) mass spectrometer

Background Identification of DNA-based biomarkers of cancer cells is highly promising and rapidly developing direction that can advance early detection and therapy of malignancies. DNA adducts are felicitous markers of cancer, because their chemical structure is significantly different from that of mutated or methylated DNA, that allows to determine them with high precision using mass spectrometry. The aim of this work is to develop the methodology of sample preparation and its mass spectrometric analysis. Samples were prepared from the blood plasma and from the tumor tissue from lung cancer patients and from blood of healthy individuals. Materials and methods DNA was isolated from the blood plasma and tissue by using column method (BioSilica, Russia) The final yield from 1 ml of blood was 100 ng. DNA samples were subjected to acid hydrolysis (1 M HCl) at 70 °C. After 3 h, the hydrolysis was stopped by cooling on ice for 5 min and later on adding an equivalent amount of an alkali and a phosphate buffer solution (pH 7). To assess the extent of hydrolysis of the samples they were analysed by electrophoresis on a 1.2% agarose gel in Tris-acetate buffer. The samples were extracted at cartridge HF Bond Elut-C18 100 mg, 1 ml (Agilent Technologies, USA) and eluted in several fractions with a gradual increase of methanol in the eluent. Stream of nitrogen was applied to dry the extract. The samples were subjected to mass spectrometric analysis after pre-separation by UHLC Ultimate 3000 RS (Dionex, USA) in a column Dionex Acclaim RSLC 120 C18 (2.1 × 50 mm 120 A, 0.2 μm) flow rate of 0.5 ml/min using as eluents 0.1% solution of formic acid in water (A) and 0.1% solution of formic acid in atsetontrile (B). Elution was carried out in gradient mode: (%B): 0–3min (5%), 3–28min (5–95%), 28–30 min (95%), 30–31 min (95–5%), 31–35 min (5%). Mass spectrometry was carried out on ESI-qTOF ultrahigh resolution Maxis 4G (Bruker, Germany) in the positive ion detection mode range 50–1000 m/z, 2 Hz with the following settings electrospray ion source: CV 3800 V, Nebulizer gas 1 bar, Dry Gas 8 l/min, Dry Temp: 200 °C. Results It was found that the DNA which was cleaved with acid hydrolysis in the result contained single DNA bases. The samples were stable at 4 °C for at least 7 days. The optimal eluent for solid phase extraction of DNA is 80% solution of methanol in water. The number of DNA adducts was evaluated by the integrated value of the mass spectrometric response detector. It was shown that most amount of adducts 4-hydroxy-1-(3-pyridyl)-1- butanone and N3-(2-carbamoyl-2-hydroxyethyl)adenine was found in DNA samples derived from tumor tissue. The adduct N7-(2-carbamoyl-2-hydroxyethyl)guanine was found in tumor tissue samples and DNA derived from plasma, as well as in all samples of healthy tissue. Conclusion The established protocol of DNA sample preparation followed by analysis using a mass spectrometer high resolution allows to detect the content of the DNA adducts in small DNA probes (100 ng). We found, that acid hydrolysis is cheaper and more practical in comparison to enzymatic digestion in order to generated samples containing single DNA bases without damaging the structure of adducts. This research was supported by Federal Targeted Programme for Research and Development in Priority Areas of Development of the Russian Scientific and Technological Complex for 2014–2020, “Development of molecular signatures for early detection of lung cancer” (No. 14.575.21.0064 from 05.08.2014, RFMEFI57514X0064) and supported by Tomsk State University, Competitiveness Improvement Program”. Work was conducted with the application of the Tomsk regional common use center technical equipment acquired thanks to a grant of the Russian Ministry of the Agreement No.14.594.21.0001 (RFMEFI59414X0001).

Abstract 2292: The role of immature colon carcinoma transcript 1 during c-myc deregulation in fast-onset mouse plasmacytoma

Abstract Murine plasmacytoma (mPCT) models human blood cancers that involve the deregulation of c-myc by a translocation that juxtaposes c-myc to one of the immunoglobin loci. The fast-developing mPCT does not depend on c-myc translocation; rather infection with v-abl/c-myc retrovirus produces elevated c-myc levels. Chromosome 11 subcytoband E2 is always (100%) found duplicated compared to slow-developing mPCT (7.1%). The gene, immature colon carcinoma transcript1 (ict1) is located within 11E2 and overexpressed as a result of 11E2 duplication. The effect of ict1 overexpression during c-myc deregulation may contribute to the aggressive development of mPCT. Here we look at the tumorigenic potential of ict1 in vitro. The ict1 gene has been cloned into an Ecdysone inducible vector and transiently transfected (Nucleofection) into mouse PreB cells that also have inducible c-myc expression activated by 4-hydroxytamoxifen. This allows us to look at overexpression of ict1 along with c-myc simultaneously or each separately. Immunofluorescence was used to confirm overexpression of the target genes. Proliferation and apoptosis levels were determined for each condition by Edu and TUNEL assay respectively. Metaphase spreads were prepared before and after each experiment. We have confirmed by immunofluorescence overexpression of ict1 using the inducible vector system. A study in these cell lines shows that ict1 overexpression increases cell proliferation levels, and may reduce apoptosis compared to cells that were not induced for ict1/c-myc expression. Metaphase spreads show normal diploid chromosome number of 40 (at time 0, and every 24 hours). Further genetic analysis to identify any chromosomal abnormalities that may account for these changes will be performed (SKY). We have identified a potential role of ict1 overexpression in vitro. In order to further explore these results we will try to restore proliferation and apoptotic levels relative to a normal PreB cell phenotype in plasmacytoma cells in vitro in the MOPC 460D cell line, and ex vivo in plasmacytoma cells isolated from PCT-carrying mice. 11E2 is syntenic to human chromosome 17q25, which has chromosomal abnormalities in many other neoplasms, such as acute and chronic myeloid leukemia, neuroblastoma, breast, ovarian, thyroid cancers. If we can identify the role of overexpression of ict1 during c-myc deregulation in fast-onset mPCT, we can gain insight into the aggressiveness of the disease progression and determine if ict1 could lead to a new therapeutic target for aggressive blood cancers with c-myc deregulation. Citation Format: Amy K. Dahl, Ruedee Sakulratchata, Sabine Mai. The role of immature colon carcinoma transcript 1 during c-myc deregulation in fast-onset mouse plasmacytoma. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 2292. doi:10.1158/1538-7445.AM2015-2292

Interaction of β(3) /β(2) -peptides, consisting of Val-Ala-Leu segments, with POPC giant unilamellar vesicles (GUVs) and white blood cancer cells (U937)--a new type of cell-penetrating peptides, and a surprising chain-length dependence of their vesicle- and cell-lysing activity.

Many years ago, β(2) /β(3) -peptides, consisting of alternatively arranged β(2) - and β(3) h-amino-acid residues, have been found to undergo folding to a unique type of helix, the 10/12-helix, and to exhibit non-polar, lipophilic properties (Helv. Chim. Acta 1997, 80, 2033). We have now synthesized such 'mixed' hexa-, nona-, dodeca-, and octadecapeptides, consisting of Val-Ala-Leu triads, with N-terminal fluorescein (FAM) labels, i.e., 1-4, and studied their interactions with POPC (=1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine) giant unilamellar vesicles (GUVs) and with human white blood cancer cells U937. The methods used were microfluidic technology, fluorescence correlation spectroscopy (FCS), a flow-cytometry assay, a membrane-toxicity assay with the dehydrogenase G6PDH as enzymatic reporter, and visual microscopy observations. All β(3) /β(2) -peptide derivatives penetrate the GUVs and/or the cells. As shown with the isomeric β(3) /β(2) -, β(3) -, and β(2) -nonamers, 2, 5, and 6, respectively, the derivatives 5 and 6 consisting exclusively of β(3) - or β(2) -amino-acid residues, respectively, interact neither with the vesicles nor with the cells. Depending on the method of investigation and on the pretreatment of the cells, the β(3) /β(2) -nonamer and/or the β(3) /β(2) -dodecamer derivative, 2 and/or 3, respectively, cause a surprising disintegration or lysis of the GUVs and cells, comparable with the action of tensides, viral fusion peptides, and host-defense antimicrobial peptides. Possible sources of the chain-length-dependent destructive potential of the β(3) /β(2) -nona- and β(3) /β(2) -dodecapeptide derivatives, and a possible relationship with the phosphate-to-phosphate and hydrocarbon thicknesses of GUVs, and eukaryotic cells are discussed. Further investigations with other types of GUVs and of eukaryotic or prokaryotic cells will be necessary to elucidate the mechanism(s) of interaction of 'mixed' β(3) /β(2) -peptides with membranes and to evaluate possible biomedical applications.

Poly-(allylamine hydrochloride)-coated but not poly(acrylic acid)-coated upconversion nanoparticles induce autophagy and apoptosis in human blood cancer cells.

Upconversion nanoparticles (UCNPs) have gained increased attention due to their various medical applications such as drug delivery, detection, imaging, and photodynamic therapy. But little is known about their direct biological activity. In the present study, we synthesized NaYF4:Yb,Er UCNPs coated with poly-(allylamine hydrochloride) (PAH) or poly(acrylic acid) (PAA) and investigated their effects in human myeloma and leukemia cells. When these cells were incubated with both types of UCNPs, we found that PAH-UCNPs but not PAA-UCNPs increased the expression of LC3-II, a hallmark of autophagy. This effect was confirmed by the accumulation of LC3 puncta as analyzed by immunofluorescence microscopy. Induction of LC3-II could be blocked by 3-methyl adenosine (3-MA), an autophagy inhibitor. Consistent with this observation, PAH-UCNPs also inhibited both AKT and mTOR activation, the key step in autophagy activation. Further studies demonstrated that PAH-UCNPs also decreased Bcl-2 but increased Beclin1 and Atg14 expression, suggesting that PAH-UCNPs-induced autophagy was associated with increased PI3KC3-Beclin1 activity. Moreover, PAH-UCNPs induced apoptosis in myeloma cells and enhanced apoptosis induced by bortezomib and doxorubicin, two major anti-myeloma drugs. Therefore, our study suggested that PAH-UCNPs alone can induce both apoptosis and autophagy in human blood cancer cells by modulating the AKT-mTOR and PI3KC3-Beclin1-Atg14 signaling pathways. This study implies the potential application of PAH-UCNPs in blood cancer-cell killing.