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ARS Home » Southeast Area » Oxford, Mississippi » Natural Products Utilization Research » Research » Research Project #428996

Research Project: Discovery and Development of Natural Products for Pharmaceutical and Agrochemical Applications II

Location: Natural Products Utilization Research

2018 Annual Report

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.

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
In a cooperative agreement with ARS researchers at Oxford, Mississippi, researchers at the National Center for Natural Products Research (NCNPR) at the University of Mississippi, Oxford, Mississippi, maintained basic discovery operations, with emphasis on the discovery of antifungals, cancer prevention agents, anti-inflammatory, anti-diabetic agents and immunomodulating agents. Continued to source plant materials for screening from our own plant collections and from numerous collaborators. We added 402 plant samples to our inventory this year and screened over 9,801 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, cancer chemopreventive, and immunomodulator/anti-inflammatory leads from natural sources, 110 compounds were isolated. In addition, 85 compounds were synthesized. Of the compounds tested, 16 were found to be biologically active for potential agricultural or medical uses. Many showed potent phytotoxic, antifungal, antibacterial, or antimalarial activities. In order to explore the antidiabetic potential of medicinal plants, about 50 plant extracts and 65 pure compounds were screened for PPAR agonistic activity. The actives were selected and were followed further in secondary assays such as adipogenic effect, antiadipogenic effect, and the agonistic effect towards LXR (which regulates lipid metabolism). 13 extracts were screened to find out if they possessed adipogenic effects (similar to rosiglitazone). A total of 22 (18 extracts and 4 pure compounds) were found to exhibit no adipogenic effects and they were selected to be tested for antiadipogenic effect to evaluate their potential against obesity and metabolic disorder as well as to understand their mechanism of antidiabetic and hypolipidemic action. Hypolipidemic action was determined in terms of activation of LXR pathway for 50 plant extracts and 64 pure compounds. 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 150 plant extracts were screened for anti-inflammatory activities through target based cellular assays including NF-kB, iNOS and oxidative stress and over 140 pure compounds were screened for anti- inflammatory activities through these targets. The cytotoxicity of actives was also determined to find out the selectivity towards the target. About 10 extracts of medicinal plants and 25 isolated pure constituents from medicinal plants were also screened for their potential for causing drug interaction in terms of PXR activation and CYPs and P-glycoprotein inhibitions. Two plants were followed for in vivo anti diabetic efficacy in high fat diet fed animal model for diabetes, obesity and metabolic disorder. 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 has now been 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 were 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 occupancy of the new plant specimen repository is 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.

1. Continued with development of agents for prevention/treatment of poison ivy dermatitis. Through a cooperative agreement with ARS researchers at Oxford, Mississippi, researchers at the National Center for Natural Products Research (NCNPR) at the University of Mississippi, Oxford, 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 U.S. Food and Drug Administration. A phase one trial has been completed by Hapten Sciences, Inc. Preparations are underway to initiate Phase II trial. Meanwhile, another formulation from one of our patented compounds prepared by Elsohly Lab, Inc. has been tested in guinea pigs at the center. This formulation was also found to be effective in preventing Poison Ivy dermatitis.

2. Identify dietary or botanical products with cancer chemopreventive potential. This program takes the most promising lead natural product extracts/pure compounds, identified using our battery of 13 cancer-related signaling pathway 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. During this period, ARS researchers at Oxford, Mississippi, have mostly focused on the mechanism of action of the most promising leads and have identified a potent inhibitor of glycolysis that when used in combination with a mitochondrial complex I inhibitor synergistically kills many types of human cancer cell lines and cells from patients tumors. We have also conducted mouse studies with this combination using the B16 melanoma model and have found significant inhibition of tumor development.

3. Continued development of immune-enhancing natural products and novel bioassays. Through a cooperative agreement with the ARS researchers at Oxford, Mississippi, researchers at the National Center for Natural Products Research (NCNPR) have developed a natural product extract from the cyanobacteria Arthrospira platensis that is enriched for Braun-type lipoproteins (potent toll-like receptor 2 agonists). Continued efforts focus on development of this product as a dietary-based approach for balancing innate immune function and its potential for enhancing the efficacy of immune checkpoint blockade in cancer therapy. A new project was also initiated with a company to help develop a mixture of natural products for treatment of bovine mastitis. During the last 15 years, NCNPR researchers have discovered that bacterial components (from endophytic bacteria) represent the major in vitro and in vivo innate immune cell activation agents within Echinacea and some other botanicals. This research received international recognition as the most innovative research published in Planta Medica during 2016. NCNPR researchers are also continuing to develop novel bioassays. In collaboration with Phytochemical Services Incorporated (Oxford, Mississippi) these researchers continue to offer and develop critically needed testing services for the standardization of immune-enhancing dietary supplements. These testing services are based on the novel concept of using bioassay-based standardization tools to quantitate the immune-enhancing potency within a broad range of supplements. In a separate project, a new bioassay system was set-up to discover natural products that expose beta glucans that are buried within the fungal cell wall, thus enabling the immune system to detect and elicit a response against these organisms.

4. New antifungal natural products for use in agriculture and medicine. Through a collaborative agreement ARS, researchers at Oxford, Mississippi and Agricen Sciences, with National Center for Natural Products Research, have identified a highly efficient microbial strain producing antifungal fusaricidins on a gram scale. A total of 48 fusaricidins including 28 new compounds have been characterized from this strain. The major compounds fusaricidins A and B account for 50% of the total fusaricidins. These compounds have demonstrated strong antifungal activities against a broad array of fungal pathogens including Cryptococcus neoformans. The structural diversity and potency of these fusaricidins makes this microbial strain as a potential bio-control agent for the treatment of fungal diseases in agricultural crops. The purified fusaricidins may serve as candidates for antifungal drug development. In a search for natural products that are potentiators of current antifungal drugs, mechanistic studies were performed. Three compounds were identified as potentiators of the cell wall damaging drug, caspofungin (CAS). The results revealed that a marine-derived cyclic peptide synergized with CAS by inducing the cell wall damage response pathway. These results were used to initiate a new target-based screen to identify compounds that induce this pathway. More than 2000 in-house compounds have been screened so far, and over 15 compounds were identified that strongly induced this pathway. Follow up studies are ongoing to determine if these compounds will synergize with CAS. NCNPR researchers also initiated a new assay to identify natural products that can function as B-glucan exposing agents, i.e. agents that can cause the B-glucans in the fungal cell wall to become exposed at the cell surface, thereby making the fungal cell visible to human immune cells. Once the optimization and validation is completed, several natural products will be screened to identify B-glucan exposing agents.

Review Publications
Parcher, J.F., Wang, M., Chittiboyina, A.G., Khan, I.A. 2018. In-source collision-induced dissociation (IS-CID): Applications, issues and structure elucidation with single-stage mass analyzers. Drug Testing and Analysis Journal. 10:28-36.
Wang, M., Haider, S., Chittiboyina, A.G., Parcher, J.F., Khan, I.A. 2018. 1,5-Dimethylhexylamine (octodrine) in sports and weight loss supplements: Natural constituent or synthetic chemical? Journal of Pharmaceutical and Biomedical Analysis. 152:298-305.
Zhao, J., Wang, M., Avula, B., Khan, I.A. 2018. Detection and quantification of phenethylamines in sports dietary supplements by NMR approach. Journal of Pharmaceutical and Biomedical Analysis. 151:347-355.
Raslan, A., Radwan, M., Ahmed, S., Nafady, A., Wanas, A., Babu, T., Hassanin, H., Elsohly, M. 2017. Evaluation of secondary metabolites from the Red Sea tunicate polyclinum constellatum. Pharmacy & Pharmacology International Journal. 5(3): doi:10.15406/ppij.2017.05.00123
Zhang, J., Zhao, J., Samoylenko, V., Jain, S., Tekwani, B.L., Muhammad, I. 2018. New polyisoprenylated polycyclic phloroglucines from Clusia gundlachii. Natural Product Communications. 13(3):361-365.
Slater, S., Lasonkar, P.B., Haider, S., Alqahtani, M.J., Chittiboyina, A.G., Khan, I.A. 2018. One-step, stereoselective synthesis of octahydrochromanes via the Prins reaction and their cannabinoid activities. Tetrahedron Letters. 59:807-810.
Wang, M., Raman, V., Zhao, J., Avula, B., Wang, Y., Wylie, P.L., Khan, I.A. 2018. Application of GC/Q-ToF combined with advanced data mining and chemometric tools in the characterization and quality control of bay leaves. Planta Medica.