Location: Natural Products Utilization Research
2016 Annual Report
Objectives
Objective 1: Discover new leads from plant-based natural resources with anti-infective and anti-cancer, immunomodulator and anti-inflammatory activities using cell-based screens and mechanistic assays using molecular target-based approaches.
Sub-objective 1A: Source novel natural resources from terrestrial plants, marine organisms and microbes from around the world for biological testing.
Sub-objective 1B: Prepare, maintain and manage Natural Products Repository and Laboratory Information Management System (LIMS).
Sub-objective 1C: Evaluate natural product extracts and natural product derived pure compounds for potential anticancer, anti-infective, anti-inflammatory, neuroprotective and anti-diabetic properties, and for utility in metabolic and immune disorders.
Sub-objective 1D: Isolation and structural elucidation of lead compounds.
Objective 2: For the best candidates, characterize mechanisms of action, selectivity, toxicity, functional activity in secondary assays and in animal models of plant-based anti-infective and anti-cancer, immunomodulator and anti-inflammatory compounds.
Sub-objective 2A: Characterization of mechanisms of action and functional activity of leads.
Sub-objective 2B: Characterization of selectivity and toxicity of lead compounds.
Objective 3: Develop methods for analysis of bioactive or medicinally important plants and quality control of their derived products.
Objective 4: Assess selected medicinal or aromatic plants for cultivation, harvest and processing to optimize yields of biomass and active principles.
Approach
The approach includes a program of: (1) Using cell-based screening and mechanistic assays to discover new pharmaceutical and agrochemical leads from natural sources; (2) Using secondary assays and animal models to characterize mechanisms of action, selectivity, toxicity and functional activity of the best candidate compounds having anti-infective, anti-cancer, immunomodulatory, or anti-inflammatory properties; (3) Selection, agronomics and analysis of bioactive or medicinally important plants and their derived products.
Progress Report
Researchers at the National Center for Natural Products Research (NCNPR) at the University of MS, Oxford, MS maintained basic discovery operations, with emphasis on the discovery of antifungals, anticancer, anti-inflammatory agents and immunomodulating agents. Continued to source plant materials for screening from our own plant collections and from numerous collaborators. Added 1057 plant samples to our inventory this year. Screened over 20,000 natural product crude extracts, semi-purified fractions and purified compounds for biological activities against specific molecular targets and whole cell systems. As part of our continuing effort in the search for anti-infective, anticancer, and immunomodulator/anti-inflammatory leads from natural sources, 128 compounds were isolated, 62 of which are novel. In addition, 53 compounds related to these isolates were synthesized. Of the compounds tested, 36 were found to be biologically active for potential agricultural or medical uses. Many showed potent phytotoxic, antifungal, antibacterial, or antimalarial activities. Approximately 250 of our isolated actives or extracts have been characterized in more detailed follow-up assays to determine their mode of action, pharmaceutical properties, toxicity, and selectivity across a range of assays. In addition to these basic operations we have selected a number of these compounds for more advanced study, whether for characterizing mechanisms of action, determining suitability for further pharmaceutical development, evaluation in disease models in preclinical studies, or in field applications. About 100 plant extracts were screened for anti-inflammatory activities through target based cellular assays including NF-kB, iNOS and oxidative stress and over 100 pure compounds were screened for anti-inflammatory activities through these targets. About 100 plant extracts and 100 pure compounds were screened for anticancer properties against human tumor cell lines. About 50 plant extracts were screened for PPAR activation activity in order to explore their antidiabetic potential. About 80 pure compounds were explored for antidiabetic potential. The actives were selected and were followed further in secondary assays such as western blots and RT-PCR to look into the expression of target proteins and evaluate the mechanism of action. About 25 extracts of medicinal plants and 40 isolated constituents from medicinal plants were also screened for their potential of causing drug interaction in terms of PXR activation and CYPs and P-gp inhibitions. Several were followed in secondary assays of enzymatic activity and gene expression analysis.
In collaboration with ElSohly Laboratories, Inc., continued development of a product shown to be effective in animal models for desensitization to poison ivy dermatitis. The lead compound in the poison ivy project is now being evaluated in a stage 1 clinical trial. The Medicinal Plant Garden at the University of Mississippi continues to expand its renowned collection of living medicinal plants. New demonstration beds and field plots are being developed. Here NCNPR cultivates and processes medicinal plants to be used in the discovery program.
NCNPR finished construction in 2015 of a 96,000 sq. ft. research wing and final stages of occupancy are in progress. The new research wing will expand and enhance the research capabilities of NCNPR with a second plant specimen repository, herbarium, and laboratories for plant tissue cultures, cellular cultures, scale-up isolation and synthetic chemistry.
Accomplishments
1. Continued with development of agents for prevention/treatment of poison ivy dermatitis. Scientists at the NCNPR at the University of Mississippi in collaboration with ElSohly Laboratories, Inc. are developing preventive treatments for poison ivy dermatitis. Development continued on the compound shown to be effective in animal models for desensitization to poison ivy dermatitis and shown to have desirable bioavailability and toxicological properties. This product has attained investigational new drug (IND) status by the US Food and Drug Administration and a phase 1 clinical trial is underway.
2. New scientific technologies that improve laboratory operations. Scientists at the NCNPR invented a chromatographic device which uses centrifugal force to efficiently separate complex mixtures of organic compounds into their various components. A prototype instrument was built and tested. A patent application and research disclosure have been filed. An option agreement has been executed with a company for commercialization of the product. This will provide new solutions to chromatographic challenges faced by scientists and laboratory technicians in many fields, including agriculture, pharmaceutics, and food science.
3. Develop treatments for cancer. A cancer research program requires a drug discovery program in order to explore all avenues of treatment. The Cancer Drug Discovery Core of the University of Mississippi Cancer Institute is a cooperative venture of the School of Pharmacy and the Medical Center. This program takes the most promising lead natural product extracts/pure compounds, identified using our battery of 13 cancer-related luciferase assays, and determines their cytotoxic potential against both bulk and tumor stem-like cells isolated from various types of patient-derived tumor biopsies in vitro. A new faculty member added this year will oversee translation of natural product leads identified by the Cancer Drug Discovery Core by characterizing those that enhance responsiveness of various tumor types to chemotherapy using a chemosensitivity testing approach (ChemoID®) developed and patented by him. During this period, we have been transitioning this assay to a 384 well format and focusing on identification of natural product extracts and pure compounds that impact glioblastoma in vitro. Currently a trial of 100 patients with glioblastoma is underway using ChemoID®; these patient derived tumor cells in culture will be used to evaluate our natural product leads. This represents an approach that could afford us an important and unique niche in the Precision Medicine Initiative - identifying patient/tumor-specific natural product-chemotherapeutic combinations that are effective against both bulk tumor cells and tumor stem-like cells.
4. Discovery of new drugs to overcome resistance in bacteria. Bacterial resistance to currently used antibiotics has become a worldwide health concern, in particular drug resistance has increased dramatically in the ESKAPE pathogens (Enterococcus faecium, Staphylococcus aureus, Klebsiella pneumoniae, Acinetobacter baumanni, Pseudomonas aeruginosa, and Enterobacter). We have developed a primary screen to evaluate the ability of natural products to reverse drug resistance. Resistant organisms are treated with natural products in the presence of sub-inhibitory concentrations of antibiotics. These organisms are resistant because they produce ESBL, NDM and KPC beta lactamases the major causes of clinical resistance and antibiotic failure. Continued screening of our natural product samples in these assays is underway.
5. Continued development of immune enhancing natural products and bioassay methods for their standardization. Scientists at the NCNPR have developed a natural product extract from the cyanobacteria Arthrospira platensis that contains a very potent Toll-like receptor 2 (TLR2) agonist. Published research supports the role of TLR2 agonists in maintaining intestinal epithelial barrier function. Increased permeability of the intestinal epithelium is associated with increased inflammation, depression and other pathophysiological conditions. We are establishing collaborations with scientists at The University of Mississippi Medical Center to perform clinical trials on the cyanobacterial extract for use in preventing cancer therapy-induced GI mucositis and treatment of depression. We are also working with the Department of Health, Exercise Science, and Recreation Management at The University of Mississippi to plan a clinical trial evaluating the effectiveness of the cyanobacterial extract in prevention of delayed onset muscle soreness. In collaboration with Phytochemical Services Incorporated (Oxford, MS) we have set-up a battery of different in vitro bioassay methods capable of quantitating the potency of natural product materials that enhance immune function. These bioassays are expected to overcome the limitations of chemical standardization methods since physiochemical properties cannot fully predict the biological potency of the high molecular weight substances that are typically responsible for the immune enhancing effects of natural product materials. Bioassay standardization methods provide the means to fully characterize immune enhancing natural product materials for use in research, clinical trials and the consumer market.
6. New antifungal natural products for use in agriculture and medicine. Scientists at the NCNPR are working on discovering novel antifungal compounds for treating life-threatening opportunistic fungal infections. In addition, projects are ongoing for discovering compounds that can potentiate the activity of current antifungal drugs, especially in drug-resistant fungal pathogens. This year, scientists in this program evaluated the mechanism behind the synergistic effects of two different natural products in combination with the antifungal drugs amphotericin B and caspofungin. Transcript profiling analysis revealed that these compounds prevented fungal cells from adapting to the specific stress exerted by each drug. In addition, RNA-Seq technology was used to determine that each compound targeted a unique cellular pathway that was different from that targeted by the antifungal drug. These discoveries will allow the development of new therapies that will improve the activity of current antifungal drugs in pathogens that have become resistant to them. This year, the amphotericin B-potentiating compound was advanced to an in vivo evaluation study in a mouse model of fungal infection in our collaborator’s lab at Duke University. Finally, this year, in addition to the analysis of in-house compounds, studies were also conducted on commercially available collections of drug-like molecules. Using a molecular assay, ~12,000 molecules were evaluated. Eight drug-like molecules were identified that potentiated the activity of the antifungal drug caspofungin in fungal pathogens, especially in drug-resistant strains. Further studies are ongoing to evaluate the in vivo efficacy of these compounds in combination with caspofungin and to characterize their drug-potentiating mechanism. These accomplishments may lead to new treatments for numerous diseases of plants, animals, and humans.
None.
Review Publications
Wang, Y., Avula, B., Abe, N., Wei, F., Wang, M., Ma, S., Ali, Z., Elsohly, M.A., Khan, I.A. 2016. Tandem Mass Spectrometry for Structural Identification of Sesquiterpene alkaloids from the stems of dendrobium nobile using LC-QToF3. Planta Medica. 82(7):662-70.
Haron, M.H., Tyler, H.L., Pugh, N.D., Moraes, R.M., Maddox, V.L., Jackson, C.R., Pasco, D.S. 2016. Activities and prevalence of proteobacteria members colonizing Echinacea purpurea fully account for in vitro macrophage activation exhibited by extracts of this botanical. Planta Medica. 89(4-5):451-62. DOI: 10.1007/s11103-015-0381-3
Jain, S., Jacob, M., Walker, L., Tekwani, B. 2016. Screening North American plant extracts in vitro against Trypanosoma brucei, the causative agent for Human African Trypanosomiasis. BMC Complementary and Alternative Medicine. 16(1):131. doi: 10.1186/s12906-016-1122-0.
Bufalo, J., Cantrell, C.L., Jacob, M.R., Schrader, K.K., Tekwani, B.L., Kustova, T.S., Ali, A., Boaro, C.S.F. Antimicrobial and antileishmanial activities of diterpenoids isolated from the roots of Salvia deserta. Planta Medica. 2016;82:131-137.
Nicolau-Goncalves, V., Cantrell, C.L., Wedge, D.E., Ferreira, M.C., Soares, M.A., Jacob, M.R., Oliveira, F., Galante, D., Rodrigues, F., Alves, T., Zani, C., Junior, P., Murta, S., Romanho, A., Barbosa, E., Kroon, E., Oliveira, J., Gomez-Silva, B., Galetovic, A., Rosa, C.A., Rosa, L. 2015. Fungi associated with rocks of the Atacama Desert: taxonomy, distribution, diversity, ecology and bioprospection for bioactive compounds. Environmental Microbiology. 18:232-245.
Avula, B., Chittiboyina, A.G., Wang, Y., Sagi, S.J., Raman, V., Wang, M., Khan, I.A. 2016. Simultaneous determination of aegeline and six coumarins from different parts of the plant Aegle marmelos using UHPLC-PDA-MS and chiral separation of aegeline using HPLC-ToF-MS . Planta Medica. 82(06):580-588.
Meepagala, K.M., Johnson, R.D., Techen, N., Wedge, D.E., Duke, S.O. 2015. Phomalactone from a phytopathogenic fungus infecting Zinnia elegans (Asteraceae) leaves. Journal of Chemical Ecology. 41:602-612.
Wang, M., Carrell, E., Chittiboyina, A.G., Avula, B., Wang, Y., Zhao, J., Parcher, J.F., Khan, I.A. 2016. Concurrent and supercritical fluid chromatographic analysis of Terpene Lactones and ginkolic acids in Ginko biloba. Analytical and Bioanalytical Chemistry. 408(17):4649-4660. doi: 10.1007/s00216-016-9544-6.
Zhang, Z., Zulfiqar, A., Khan, S.I., Khan, I.A. 2016. Cytotoxic monacolins from red yeast rice, a Chinese medicine and food. Journal of Natural Products. 202:262-8. DOI:10.1016/j.foodchem.2015.12.039.
Aumsuawan, P., Khan, S.I., Khan, I.A., Ali, Z., Avula, B., Walker, L.A., Shariat-Madar, Z., Helferich, W.G., Katzenellenbogen, B.S., Dasmahapatra, A.K. 2016. The anticancer potential of steroidal saponin, dioscin, isolated from wild yam (Dioscorea villosa) root extract in invasive human breast cancer cell line MDA-MB-231 in vitro. Archives Of Biochemistry and Biophysics. 591:98-110. doi: 10.1016/j.abb.2015.12.001.
Manda, V.K., Ibrahim, M.A., Dale, O.R., Kumarihamy, M., Cutler, S.J., Khan, I.A., Walker, L.A., Muhammad, I., Khan, S.I. 2016. Modulation of CYPs, P-gp, and PXR by Eschscholzia californica (California poppy) and its alkaloids. Planta Medica. 82:551-558. DOI http://dx.doi.org/10.1055/s-0042-103689
Lata, H., Chandra, S., Wang, Y., Elsohly, M.A., Khan, I.A. 2015. Polyhouse cultivation of invitro raised elite Stevia rebaudiana Bertoni: An assessment of biochemical and photosynthetic characteristics. Journal of Tropical Agriculture. 33(4):1-7. 2015
Dasmahapatra, A.K., Khan, I.A. 2015. DNA methyltransferase expressions in Japanese rice fish (Oryzias latipes) embryogenesis is developmentally regulated and modulated by ethanol and 5-azacytidine. Comparative Biochemistry and Physiology Part C: Toxicology & Pharmacology. 13:176-177 (2015).
Aumsuwan, P., Khan, S.I., Khan, I.A., Walker, L.A., Dasmahapatra, A.K. 2016. Gene expression profiling and pathway analysis in MCF-7 and MDA-MB-231 human breast cancer cell lines treated with dioscin. Data in Brief. (2016)272-279.
Avula, B., Sagi, S.J., Gafner, S., Upton, R., Wang, Y., Wang, M., Khan, I.A. 2015. Identification of Ginkgo biloba supplements adulteration using high performance thin layer chromatography and ultra high performance liquid chromatography-diode array detector-quadrupole time of flight-mass spectometry. Analytical and Bioanalytical Chemistry. 407(25):7733-7746. doi: 10.1007/s00216-015-8938-1.
Kaymakcioglu, B.K., Beyhan, N., Tabanca, N., Ali, A., Wedge, D.E., Duke, S.O., Bernier, U.R., Khan, I.A. 2015. Discovery and structure activity relationships of 2-pyrazolines derived from chalcones from a pest management perspective. Medicinal Chemistry Research. 24:3632-3644.
Parveen, I., Wang, M., Zhao, J., Chittiboyina, A.G., Tabanca, N., Ali, A., Baerson, S.R., Techen, N., Chappell, J., Khan, I.A., Pan, Z. 2015. Investigating sesquiterpene biosynthesis in Ginkgo biloba: molecular cloning and functional characterization of (E,E)-farnesol and a-bisabolene synthases. Plant Molecular Biology. 89:451-462.
Ding, Y., Liu, H., Tekwani, B.L., Nanayakkara, N., Khan, I.A., Walker, L.A., Doerksen, R.J. 2016. Methemoglobinemia hemotoxicity of some antimalarial 8-aminoquinoline analogues and their hydroxylated derivatives: density functional theory computation of ionization potentials. Chemical Research in Toxicology. http://dx.doi.org/acs.orglett.6b00063.
Li, C., Ren, G., Yang, B., Miklossy, G., Turkson, J., Fei, P., Ding, Y., Walker, L.A., Cao, S. 2016. Meroterpenoids with antiproliferative activity from a Hawaiian-plant associated fungus Peyronellaea coffeae-arabicae FT238. Organic Letters. 18(10):2335-8. doi:10.1021/acs.orglett.6b00685.
Li, C., Ding, Y., Yang, B., Miklossy, G., Yin, H., Walker, L.A., Turkson, J., Cao, S. 2015. A New Metabolite with a unique 4-pyranone-ylactam-1,4-thiazine moiety from a Hawaiian-plant associated fungus. Organic Letters. 17(14):3556-9. doi:10.1021/acs.orglett.5b01650.
Tekwani, B.L., Avula, B., Sahu, R., Chaurasiya, N.D., Khan, S.I., Jain, S., Fasinu, P.S., Herath, H.B., Stanford, D., Nanayakkara, N.D., Mcchesney, J.D., Yates, T.W., Elsohly, M.A., Khan, I.A., Walker, L.A. 2015. Enantioselective pharmacokinetics of primaquine in healthy human volunteers. Drug Metabolism and Disposition. 43(4):571-7.
Avonto, C., Chttiboyina, A.G., Rua, D., Khan, I.A. 2015. A fluorescence high throughput screening method for the detection of reactive electrophiles as potential skin sensitizers. Journal of Toxicology and Applied Pharmacology. 289:177-184.
Cimmino, A., Evidente, M., Masi, M., Ali, A., Tabanca, N., Khan, I.A., Evidente, A. 2015. Papyracillic acid and its derivatives as biting deterrents against Aedes aegypti(Diptera: Culicidae): structure–activity relationships. Medical Chemistry Research. 24:3981-3989.
Avonto, C., Chittiboyina, A.G., Wang, M., Vasquez, Y., Rua, D., Khan, I.A. 2016. In chemico evaluation of tea tree essential oils as skin sensitizers: Impact of the chemical composition on aging and generation of reactive species. Chemical Research in Toxicology. 29:1108-1117.
Li, C., Ding, Y., Yang, B., Hoffman, N., Yin, H., Mahmud, T., Turkson, J., Cao, S. 2016. Ermophilane sesquiterpenes from Hawaiian endophytic fungus Chaetoconis sp.FT087. Phytochemistry. 126:41-6. doi:doi:10.1016/j.phytochem.2016.03.005.
Mohamed, S.M., Bachkeet, E.Y., Bayoumi, S.A., Jain, S., Cutler, S., Tekwani, B.L., Ross, S.A. 2015. Potent antitrypanosomal triterpenoid saponins from Mussaenda luteola. Fitoterapia. 107:114-121.
Radwan, M., Wanas, A.S., Fronczek, F.R., Jacob, M.R., Ross, S.A. 2015. Polybrominated diphenyl ethers from the marine organisms Lendenfeldia dendyi and Sinularia dura with anti-MRSa activity. Medical Chemistry Research. 24:3398-3404.
Gorovoi, P.G., Suleimen, E.M., Dudkin, R.V., Wang, M., Khan, I.A., Ross, S.A. 2015. Constituent composition and biological activity of Nepeta manchuriensis essential oil. Chemistry of Natural Compounds. 51(5):989-990.
Fasinu, P.S., Gurley, B.J., Walker, L.A. 2015. Clinically relevant pharmacokinetic herb-drug interactions in antiretroviral therapy. Current Drug Metabolism. 17(1):52-64.
