Gajendra Shrestha

Lichens: a promising source of antibiotic and anticancer drugs

Lichens are symbiotic associations between fungi and a photosynthetic alga and/or cyanobacteria. Lichenized fungi have been found to produce a wide array of secondary metabolites, most of which are unique to the lichenized condition. These secondary metabolites have shown an impressive range of biological activities including antibiotics, antifungal, anti-HIV, anticancer, anti-protozoan, etc. This review focuses primarily on the antibiotic and anticancer properties of lichen secondary chemicals. We have reviewed various publications related to antibiotic and anticancer drug therapies emphasizing results about specific lichens and/or lichen compounds, which microbes or cancer cells were involved and the main findings of each study. We found that crude lichen extracts and various isolated lichen compounds often demonstrate significant inhibitory activity against various pathogenic bacteria and cancer cell lines at very low concentrations. There were no studies examining the specific mechanism of action against pathogenic bacteria; however, we did find a limited number of studies where the mechanism of action against cancer cell lines had been explored. The molecular mechanism of cell death by lichen compounds includes cell cycle arrest, apoptosis, necrosis, and inhibition of angiogenesis. Although lichens are a reservoir for various biologically active compounds, only a limited number have been tested for their biological significance. There is clearly an urgent need for expanding research in this area of study, including in depth studies of those compounds which have shown promising results as well as a strong focus on identifying specific mechanisms of action and extensive clinical trials using the most promising lichen based drug therapies followed by large scale production of the best of those compounds.

Anticancer activities of selected species of North American lichen extracts.

Cancer is the second leading cause of human deaths in the USA. Despite continuous efforts to treat cancer over the past 50 years, human mortality rates have not decreased significantly. Natural products, such as lichens, have been good sources of anticancer drugs. This study reports the cytotoxic activity of crude extracts of 17 lichen species against Burkitts lymphoma (Raji) cells. Out of the 17 lichen species, extracts from 14 species showed cytotoxicity against Raji cells. On the basis of IC50 values, we selected Xanthoparmelia chlorochroa and Tuckermannopsis ciliaris to study the mechanism of cell death. Viability of normal lymphocytes was not affected by the extracts of X. chlorochroa and T. ciliaris. We found that extracts from both lichens decreased proliferation, accumulated cells at the G0/G1 stage, and caused apoptosis in a dose‐dependent manner. Both lichen extracts also caused upregulation of p53. The T. ciliaris extract upregulated the expression of TK1 but X. chlorochroa did not. We also found that usnic, salazinic, constictic, and norstictic acids were present in the extract of X. chlorochroa, whereas protolichesterinic acid in T. ciliaris extracts. Our data demonstrate that lichen extracts merit further research as a potential source of anticancer drugs. Copyright

The immunostimulating role of lichen polysaccharides: a review.

The immune system has capacity to suppress the development or progression of various malignancies including cancer. Research on the immunomodulating properties of polysaccharides obtained from plants, microorganisms, marine organisms, and fungi is growing rapidly. Among the various potential sources, lichens, symbiotic systems involving a fungus and an alga and/or a cyanobacterium, show promise as a potential source of immunomodulating compounds. It is well known that lichens produce an abundance of structurally diverse polysaccharides. However, only a limited number of studies have explored the immunostimulating properties of lichen polysaccharides. Published studies have shown that some lichen polysaccharides enhance production of nitrous oxide (NO) by macrophages and also alter the production levels of various proinflammatory and antiinflammatory cytokines (IL‐10, IL‐12, IL‐1β, TNF‐α, and IFN‐α/β) by macrophages and dendritic cells. Although there are only a limited number of studies examining the role of lichen polysaccharides, all results suggest that lichen polysaccharides can induce immunomodulatory responses in macrophages and dendritic cells. Thus, a detailed evaluation of immunomodulatory capacity of lichen polysaccharides could provide a unique opportunity for the discovery of novel therapeutic agents. Copyright

Predicting the distribution of the air pollution sensitive lichen species Usnea hirta

Usnea hirta, an important member of the lichen family Parmeliaceae, has long been used as a bio-monitor of air pollution, particularly of sulphur dioxide in North America. Although U. hirta has a wide geographical distribution, it is important to be able to identify accurately the optimal habitat conditions for air pollution-sensitive species, thus making it possible to more effectively and efficiently establish air quality bio-monitoring stations. We modelled the distribution of U. hirta as a function of nine variables, five macroclimatic variables: average monthly precipitation, average monthly minimum temperature, average monthly maximum temperature, solar radiation, and inte- grated moisture index, and four topographic variables: elevation, slope, aspect, and land forms and uses for the White River National Forest, Colorado. The response variable was developed based on the presence or absence of U. hirta at each of 72 bio-monitoring baseline sites established in selected portions of four intermountain area states. Our model was developed using Non-Parametric Multi- plicative Regression (NPMR) analysis, a modelling approach that analyzes environmental gradients, or predictor variables, against known locations for individuals of the model species. Finally, we evaluated our model on the basis of log b values and overall improvement over a naive model and the Monte Carlo Permutation Test with 1000 randomized runs. The best model for U. hirta included four variables - solar radiation, average monthly precipitation, and average monthly minimum and maximum temperatures (log b ¼ 3� 68). Among these four variables, average monthly maximum temperature was the most influential predictor (sensitivity ¼ 0� 71) for the distribution of U. hirta. The occurrence rate for U. hirta, based on field validation, was 45� 5%, 65� 4%, and 70� 4% for low, medium, and high probability areas, respectively. This study showed that our model was successful in predicting the distribution of U. hirta in the White River National Forest. Based on these results, the north-eastern and western portions of the forest appear to offer the most favourable conditions for the installation of future air quality bio-monitoring baseline sites.

In vitro evaluation of the antibacterial activity of extracts from 34 species of North American lichens

Abstract Context: The emergence of antibiotic resistant pathogens is a serious global health threat. Hence, the search for new antibiotic drugs from various natural sources should be given high priority. Lichens produce a variety of low molecular weight metabolic compounds and many cultures have utilized these compounds in traditional medicine for centuries. Objective: Report the antibiotic properties of extracts from 34 North American lichens screened against four pathogenic bacteria. Materials and methods: The micro-well dilution method was used to determine the minimum inhibitory concentration (MIC) of acetone and methanol extracts of 34 lichen species against four bacterial strains. Major chemical compounds in each species were identified using thin layer chromatography (TLC). Results: Most of the lichen extracts demonstrated inhibitory effects against Staphylococcus aureus, Pseudomonas aeruginosa, and methicillin-resistant S. aureus (MRSA) with MIC values ranging from 3.9 to 500 µg/ml. In addition, extracts from three species, Letharia columbiana (Nutt.) J. W. Thomson (Parmeliaceae), Letharia vulpina (L.) Hue (Parmeliaceae), and Vulpicida canadensis (Räsänen) J.-E. Mattsson & M. J. Lai (Parmeliaceae) (MIC = 125–500 µg/ml) were also effective against Escherichia coli. Generally, acetone extractions were found to be more effective than methanol extractions. Discussion and conclusion: Results of this study show that lichen extracts provide significant antimicrobial activity against both Gram-positive and Gram-negative bacteria. These results suggest that lichens may be an important potential source of antibacterial drugs.

Teuvoa , a new lichen genus in Megasporaceae (Ascomycota: Pertusariales ), including Teuvoa junipericola sp. nov.

The relationship of Aspicilia uxoris within Megasporaceae is assessed within a phylogenetic context. ‘ Aspicilia ’ uxoris and other related species are recovered as sister to the genus Lobothallia s. str. and described here as a new genus. Teuvoa (Ascomycota, Megasporaceae ) is erected based on nuclear ITS and LSU sequence data and morphological characters. In addition to Teuvoa uxoris , a second species, T. junipericola, is added to the new genus based on material collected from North America. Teuvoa junipericola, T. uxoris and T. tibetica form a group with 8-spored asci, absence of extrolites, rather short-sized conidia and ascospores, lack of a subhypothecial algal layer, and different substratum preferences (on organic substratum) with a sister relationship to genus Lobothallia s. lat. ( Aspicilia subgenus Pachyothallia Clauzade & C. Roux). Based on spore measurements of the holotypes, Lecanora ferganensis Tomin from central Asia (Kyrgyzstan, Tajikistan and Uzbekistan), Lecanora atrodiscata Gintovt, from Tajikistan and Lecanora takyroides Dzhur. from Turkmenistan are new synonyms to T. uxoris . A lectotype for Lecanora ferganensis is designated, expanding the known distribution of T. uxoris from Algeria, Morocco and Spain, into Central Asia.

Letharia vulpina, a vulpinic acid containing lichen, targets cell membrane and cell division processes in methicillin-resistant Staphylococcus aureus

Abstract Context: Antibiotic resistance in humans is a major concern. Drugs that target traditional sites and pathways are becoming obsolete; thus, compounds affecting novel targets are needed. Screening lichen metabolites for antimicrobials has yielded promising antimicrobial compounds, yet their mode of action is poorly understood. Letharia vulpina (L.) Hue (Parmeliaceae) has traditionally been used to poison predators, and treat stomach disorders; more recently L. vulpina extracts have demonstrated promising antimicrobial properties. Objective: This study investigates the mode of action of L. vulpina acetone extract against a methicillin-resistant Staphylococcus aureus (MRSA). Material and methods: We treated MRSA with L. vulpina extracts at 1×, 5×, and 10 × MIC values (MIC = 31.25 µg/ml) for 24 h and optical density (OD660) was measured over time to determine bacteriolytic activity; counted colony forming units (CFUs) to determine time kill dynamics; the propidium iodide (PI) assay and transmission electron microscopy were used to assess membrane-damage potential, and thin-layer chromatography was used to identify secondary compounds. Results: Bacteriolytic assays showed that L. vulpina extracts, containing only vulpinic acid, do not cause cell lysis, even at 10 × MIC values but there was 92% reduction in bacterial CFUs when treated with increased concentrations of lichen extracts over 24 h at 4 h intervals. Our data indicate that the L. vulpina extract compromises membrane integrity of the MRSA isolate and disrupts cell division processes. Discussion and conclusion: Based on this study, detailed examination of acetone extracts of L. vulpina as well as pure extracts of vulpinic acid as potential antibacterial compounds merit further study.

Lichen Phenolics: Environmental Effects

Abstract Lichens, obligate symbiotic associations between a fungus and an alga and/or cyanobacterium, produce an impressive diversity of secondary metabolites. These secondary compounds are derived from three fungal metabolic pathways—acetyl polymalonyl, mevalonic acid, and shikimic acid pathways. Lichen phenols are unique and are produced mainly by the acetyl-polymalonyl pathway and occasionally by the shikimic acid pathway. Lichen phenolic compounds have several potential pharmaceutical applications based on antibiotic, anticancer, antioxidant, and anti-viral properties. In addition, some lichen phenolics serve as “sun screen” effectively regulating the quality and quantity of light available to the algal partner (photobiont). The production and diversity of phenolics in lichens is influenced by various environmental factors such as light, UV radiation, temperature, elevation, geographic and seasonal variation, as well as culture-related conditions. Although phenolics in lichens are produced by a fungal pathway, the photobiont plays a significant role in terms of providing the “raw materials” for assembling the phenolic compounds. Research concerning the positive and negative effects of environmental parameters on the phenolic content of lichens has yielded contradictory results. These environmentally mediated differences may be attributed to the fact lichen phenolics are produced by different pathways regulated by variable gene groups which could potentially yield different responses under the same environmental conditions.

The Lichen Flora of Southwestern Colorado

Abstract. This paper reports on the lichen flora of the San Juan-Rio Grande National Forest located in southwestern Colorado. This survey is based on extensive collections made between 1993 and 1997, as part of an air quality bio-monitoring program and baseline. Southwest Colorado is a biologically diverse and habitat rich part of the state. Community types include arid, low-elevation shrublands, riparian communities, mid-elevation woodlands, upper elevation coniferous forests, and extensive alpine tundra. Lichen communities along the Colorado Front Range and adjacent portions of the Rocky Mountains have been studied for more than a half century; however, lichen communities in the western half of the state and particularly the southwest corner of the state are largely unknown. Based on our research we have identified a total of 316 species in 108 genera from 32 sites in southwest Colorado. Thirty-three species are reported as new records for the state; with a single new record for North America. The lichen flora for this region is diverse and well developed. Species representing all growth forms and substrates are reported. The flora is dominated by foliose species (44%), followed by crustose taxa (35%), squamulose species (14%), with fruticose species at 7%. Species were collected from all habitats and substrates including soil/moss, rock, bark/lignum, and other lichens (with 45% on rock, 25% on bark/lignum, 29% on soil/moss, and 1% on other lichens). The diversity of lichen species reported for this area reflects the complexity and variation in habitat and environmental parameters characteristic of southwest Colorado. Based on preliminary comparisons with adjacent lichen floras, in western New Mexico and southeastern Utah differences seem to be related to: 1) variation in collection intensity; 2) differences in habitat types; and 3) variation in the frequency of monsoonal summer rains.

The expansion of targetable biomarkers for CAR T cell therapy

Biomarkers are an integral part of cancer management due to their use in risk assessment, screening, differential diagnosis, prognosis, prediction of response to treatment, and monitoring progress of disease. Recently, with the advent of Chimeric Antigen Receptor (CAR) T cell therapy, a new category of targetable biomarkers has emerged. These biomarkers are associated with the surface of malignant cells and serve as targets for directing cytotoxic T cells. The first biomarker target used for CAR T cell therapy was CD19, a B cell marker expressed highly on malignant B cells. With the success of CD19, the last decade has shown an explosion of new targetable biomarkers on a range of human malignancies. These surface targets have made it possible to provide directed, specific therapy that reduces healthy tissue destruction and preserves the patient’s immune system during treatment. As of May 2018, there are over 100 clinical trials underway that target over 25 different surface biomarkers in almost every human tissue. This expansion has led to not only promising results in terms of patient outcome, but has also led to an exponential growth in the investigation of new biomarkers that could potentially be utilized in CAR T cell therapy for treating patients. In this review, we discuss the biomarkers currently under investigation and point out several promising biomarkers in the preclinical stage of development that may be useful as targets.