Anti-cancer agents in Saudi Arabian herbals revealed by automated high-content imaging

Natural products have been used for medical applications since ancient times. Commonly, natural products are structurally complex chemical compounds that efficiently interact with their biological targets, making them useful drug candidates in cancer therapy. Here, we used cell-based phenotypic profiling and image-based high-content screening to study the mode of action and potential cellular targets of plants historically used in Saudi Arabia’s traditional medicine. We compared the cytological profiles of fractions taken from Juniperus phoenicea (Arar), Anastatica hierochuntica (Kaff Maryam), and Citrullus colocynthis (Hanzal) with a set of reference compounds with established modes of action. Cluster analyses of the cytological profiles of the tested compounds suggested that these plants contain possible topoisomerase inhibitors that could be effective in cancer treatment. Using histone H2AX phosphorylation as a marker for DNA damage, we discovered that some of the compounds induced double-strand DNA breaks. Furthermore, chemical analysis of the active fraction isolated from Juniperus phoenicea revealed possible anti-cancer compounds. Our results demonstrate the usefulness of cell-based phenotypic screening of natural products to reveal their biological activities.

[1]  V. Mohan,et al.  GC-MS ANALYSIS OF BIOACTIVE COMPONENTS OF AERIAL PARTS OF KIRGANELIA RETICULATA POIR (EUPHORBIACEAE) , 2013 .

[2]  John D. Roberts.  ABCs of FT-NMR , 2000 .

[3]  C. Voolstra,et al.  High-resolution phenotypic profiling of natural products-induced effects on the single-cell level , 2017, Scientific Reports.

[4]  A. Emwas,et al.  Solid state NMR and bioequivalence comparison of the pharmacokinetic parameters of two formulations of clindamycin. , 2011, International journal of clinical pharmacology and therapeutics.

[5]  P. Keifer.  NMR spectroscopy in drug discovery: tools for combinatorial chemistry, natural products, and metabolism research. , 2000, Progress in drug research. Fortschritte der Arzneimittelforschung. Progres des recherches pharmaceutiques.

[6]  E. Ahn,et al.  Sphinganine causes early activation of JNK and p38 MAPK and inhibition of AKT activation in HT-29 human colon cancer cells. , 2006, Anticancer research.

[7]  Jing Zhang,et al.  High-content screening moves to the front of the line. , 2006, Drug discovery today.

[8]  H. Gottlieb,et al.  Growth inhibitory activity of cucurbitacin glucosides isolated from Citrullus colocynthis on human breast cancer cells. , 2007, Biochemical pharmacology.

[9]  S. Sarker,et al.  Citrullus colocynthis (L.) Schrad (bitter apple fruit): a review of its phytochemistry, pharmacology, traditional uses and nutritional potential. , 2014, Journal of ethnopharmacology.

[10]  N. Hail.  Mitochondria: A novel target for the chemoprevention of cancer , 2005, Apoptosis.

[11]  A. El-Osta,et al.  γH2AX: a sensitive molecular marker of DNA damage and repair , 2010, Leukemia.

[12]  H. Kessler,et al.  Applications of NMR in drug discovery. , 2001, Current opinion in chemical biology.

[13]  E. Baydan,et al.  Cytotoxic effects of etephon and maleic hydrazide in Vero, Hep2, HepG2 cells , 2014, Drug and chemical toxicology.

[14]  S. Alwasel,et al.  Investigation of Antiulcer and Antioxidant Activity of Juniperus phoenicea L. (1753) Essential Oil in an Experimental Rat Model. , 2015, Biological & pharmaceutical bulletin.

[15]  P. Taylor,et al.  In vitro activity of extracts and constituents of Pelagonium against rapidly growing mycobacteria. , 2004, International journal of antimicrobial agents.

[16]  Z. Darżynkiewicz,et al.  Induction of ATM Activation, Histone H2AX Phosphorylation and Apoptosis by Etoposide: Relation to Cell Cycle Phase , 2007, Cell cycle.

[17]  M. Rich.  Eicosanoids and Other Bioactive Lipids in Cancer, Inflammation and Radiation Injury , 1993, Developments in Oncology.

[18]  L. Jelinski.  Modern NMR Spectroscopy. , 1984 .

[19]  S. Malek,et al.  Cytotoxic Components of Pereskia bleo (Kunth) DC. (Cactaceae) Leaves , 2009, Molecules.

[20]  D Lansing Taylor,et al.  Past, present, and future of high content screening and the field of cellomics. , 2007, Methods in molecular biology.

[21]  K. Kohn,et al.  Apoptosis induced by topoisomerase inhibitors. , 2003, Current medicinal chemistry. Anti-cancer agents.

[22]  E. Abdallah,et al.  Traditional medicinal plants indigenous to Al-Rass province, Saudi Arabia , 2010 .

[23]  B. Poolman,et al.  Mechanisms of membrane toxicity of hydrocarbons. , 1995, Microbiological reviews.

[24]  Roger S. Macomber,et al.  A Complete Introduction to Modern NMR Spectroscopy , 1997 .

[25]  T. Efferth,et al.  Pharmacogenomics of Cameroonian traditional herbal medicine for cancer therapy. , 2011, Journal of ethnopharmacology.

[26]  K. McManus,et al.  ATM-dependent DNA damage-independent mitotic phosphorylation of H2AX in normally growing mammalian cells. , 2005, Molecular biology of the cell.

[27]  Mark R Viant,et al.  Optimized metabolite extraction from blood serum for 1H nuclear magnetic resonance spectroscopy. , 2008, Analytical biochemistry.

[28]  W. Lee,et al.  Antioxidant effects of quinoline alkaloids and 2,4-di-tert-butylphenol isolated from Scolopendra subspinipes. , 2006, Biological & pharmaceutical bulletin.

[29]  Z. Darżynkiewicz,et al.  DNA Damage Induced by DNA Topoisomerase I- and Topoisomerase II- Inhibitors Detected by Histone H2AXphosphorylation in Relation to the Cell Cycle Phase and Apoptosis , 2003, Cell cycle.

[30]  Rafael Brüschweiler,et al.  Strategy for automated analysis of dynamic metabolic mixtures by NMR. Application to an insect venom. , 2007, Analytical chemistry.

[31]  Anthony Nichols,et al.  High content screening as a screening tool in drug discovery. , 2007, Methods in molecular biology.

[32]  J. White,et al.  Effect of Stearic Acid on Human Cervical Cancer Cell Growth , 1993 .

[33]  Joshua M. Stuart,et al.  "Function-first" lead discovery: mode of action profiling of natural product libraries using image-based screening. , 2013, Chemistry & biology.

[34]  K. Stoeber,et al.  The cell cycle and cancer , 2012, The Journal of pathology.

[35]  Yan Yin,et al.  NF-κB, JNK and p53 pathways are involved in tubeimoside-1-induced apoptosis in HepG2 cells with oxidative stress and G₂/M cell cycle arrest. , 2011, Food and chemical toxicology : an international journal published for the British Industrial Biological Research Association.

[36]  L. Liu,et al.  Tumor cell death induced by topoisomerase-targeting drugs. , 2001, Annual review of pharmacology and toxicology.

[37]  R. Abratt,et al.  The cancer burden in Africa. , 2007, South African medical journal = Suid-Afrikaanse tydskrif vir geneeskunde.

[38]  Mamdouh M. Ali,et al.  Synthesis and anticancer effects of some novel pyrazolo[3,4-d]pyrimidine derivatives by generating reactive oxygen species in human breast adenocarcinoma cells. , 2011, European journal of medicinal chemistry.

[39]  A I Saeed,et al.  TM4: a free, open-source system for microarray data management and analysis. , 2003, BioTechniques.

[40]  P. Sun,et al.  ISOLATION, PURIFICATION AND STRUCTURAL CHARACTERIZATION OF A NOVEL WATER-SOLUBLE GLUCAN FROM THE FRUITING BODIES OF PHELLINUS BAUMII PILÁT , 2010 .

[41]  Sanchita,et al.  Purification, characterization, and in vitro activity of 2,4-Di-tert-butylphenol from Pseudomonas monteilii PsF84: conformational and molecular docking studies. , 2014, Journal of agricultural and food chemistry.

[42]  U. Moll,et al.  p53's mitochondrial translocation and MOMP action is independent of Puma and Bax and severely disrupts mitochondrial membrane integrity , 2008, Cell Research.

[43]  D. Kingston,et al.  Plant anticancer agents. X. Lignans from Juniperus phoenicea. , 1980, Journal of Natural Products.

[44]  G. Agoramoorthy,et al.  Antibacterial and antifungal activities of fatty acid methyl esters of the blind-your-eye mangrove from India , 2007 .

[45]  Oliver Rausch,et al.  High content cellular screening. , 2006, Current opinion in chemical biology.

[46]  Albert Gough,et al.  High-Content Screening: A New Approach to Easing Key Bottlenecks in the Drug Discovery Process , 1997 .

[47]  Oleg Lapets,et al.  A quantitative cell-based high-content screening assay for the epidermal growth factor receptor-specific activation of mitogen-activated protein kinase. , 2004, Assay and drug development technologies.

[48]  S. Mizutani,et al.  Phosphorylation of histone H2AX at M phase in human cells without DNA damage response. , 2005, Biochemical and biophysical research communications.

[49]  I. Malami,et al.  Effects of standardized stem bark extract of Mangifera indica L. in wistar rats with 2,4-dinitrophenylhydrazine-induced haemolytic anaemia , 2015 .

[50]  P. Pacher,et al.  Simple quantitative detection of mitochondrial superoxide production in live cells. , 2007, Biochemical and biophysical research communications.

[51]  I. Cock,et al.  Gas chromatography-mass spectroscopy analysis of bioactive petalostigma extracts: Toxicity, antibacterial and antiviral activities , 2014, Pharmacognosy magazine.

[52]  A. Składanowski,et al.  H2AX Phosphorylation: Its Role in DNA Damage Response and Cancer Therapy , 2010, Journal of nucleic acids.

[53]  Szu-Ying Wu,et al.  NPRL-Z-1, as a New Topoisomerase II Poison, Induces Cell Apoptosis and ROS Generation in Human Renal Carcinoma Cells , 2014, PloS one.

[54]  E. Krausz,et al.  Cell-based high-content screening of small-molecule libraries. , 2007, Current opinion in chemical biology.

[55]  Yves Pommier,et al.  γH2AX and cancer , 2008, Nature Reviews Cancer.

[56]  John A. Tallarico,et al.  Integrating high-content screening and ligand-target prediction to identify mechanism of action. , 2008, Nature chemical biology.

[57]  Christian Bailly,et al.  Contemporary challenges in the design of topoisomerase II inhibitors for cancer chemotherapy. , 2012, Chemical reviews.

[58]  H. Akbari,et al.  Evaluation of Effectiveness of Herbal Medication in Cancer Care: A Review Study , 2012, Iranian journal of cancer prevention.

[59]  R. Radi,et al.  Mitochondrial superoxide radicals mediate programmed cell death in Trypanosoma cruzi: cytoprotective action of mitochondrial iron superoxide dismutase overexpression. , 2007, The Biochemical journal.

[60]  B. Stewart,et al.  World cancer report 2014. , 2014 .

[61]  A. Emwas,et al.  Gas chromatography-mass spectrometry of biofluids and extracts. , 2015, Methods in molecular biology.

[62]  M. Curini,et al.  Diterpenoids and Triterpenoids from the Resin of Bursera microphylla and Their Cytotoxic Activity. , 2015, Journal of natural products.

[63]  G. Zon,et al.  31P- and 13C-n.m.r.-spectral and chemical characterization of the end-group and repeating-unit components of oligosaccharides derived by acid hydrolysis of Haemophilus influenzae type b capsular polysaccharide. , 1983, Carbohydrate research.

[64]  A. Emwas,et al.  Sample collection and preparation of biofluids and extracts for gas chromatography-mass spectrometry. , 2015, Methods in molecular biology.

[65]  Alessandra Montecucco,et al.  Cellular response to etoposide treatment. , 2007, Cancer letters.

[66]  S. Benchimol,et al.  Poly(ADP-ribose) Polymerase-1 Is a Positive Regulator of the p53-mediated G1 Arrest Response following Ionizing Radiation* , 2003, Journal of Biological Chemistry.

[67]  F. Khorshid,et al.  The cytotoxic effect of small and large molecules of PMF fraction extracted from camel urine on cancer cells. , 2015 .

[68]  Chao-Po Lin,et al.  Roles of DNA topoisomerase II isozymes in chemotherapy and secondary malignancies , 2007, Proceedings of the National Academy of Sciences.

[69]  G. Wang,et al.  Use-dependent inhibition of Na+ currents by benzocaine homologs. , 1996, Biophysical journal.