In vitro evaluation against Leishmania amazonensis and Leishmania chagasi of medicinal plant species of interest to the Unified Health System.

Leishmaniasis is a disease of public health relevance that demands new therapeutic alternatives due to the toxicity of conventional treatments. In this study, 27 plants of interest to the Unified Health System (SUS) were evaluated for cytotoxicity in macrophages, leishmanicidal activity and production of nitric oxide (NO). None of the species demonstrated cytotoxicity to macrophages (CC50 >100 µg/mL). Extracts from Chenopodium ambrosioides, Equisetum arvense, Maytenus ilicifolia showed greater efficacy in inducing the death of Leishmania amazonensis amastigotes with IC50 of 68.4, 82.3, 75.7 µg/mL, respectively. The species Cynara scolymus, Punica granatum and Passiflora alata were the most effective in inducing an increase in the indirect concentration of NO (41.31, 29.30 and 28.86 µM, respectively) in cultures of macrophages infected with L. amazonensis. Furthermore, Punica granatum was also the most effective species in inducing an increase in NO in macrophages infected by Leishmania chagasi (19.90 µM). The results obtained so far support the continuation of studies, with the possibility of developing safer and more effective treatments for leishmaniasis, using natural products. The identification of plants that stimulate the production of NO in macrophages infected by Leishmania opens doors for more detailed investigations of the mechanism of action of these natural products.

PI3K Signaling Pathways as a Molecular Target for Glioblastoma Multiforme.

Glioblastoma multiforme (GBM) is the most common type of cancer that affects the central nervous system (CNS). It currently accounts for about 2% of diagnosed malignant tumors worldwide, with 296,000 new cases reported per year. The first-choice treatment consists of surgical resection, radiotherapy, and adjuvant chemotherapy, which increases patients' survival by 15 months. New clinical and pre-clinical research aims to improve this prognosis by proposing the search for new drugs that effectively eliminate cancer cells, circumventing problems such as resistance to treatment. One of the promising therapeutic strategies in the treatment of GBM is the inhibition of the phosphatidylinositol 3-kinase (PI3K) pathway, which is closely related to the process of tumor carcinogenesis. This review sought to address the main scientific studies of synthetic or natural drug prototypes that target specific therapy co-directed via the PI3K pathway, against human glioblastoma.

New immunodiagnostic methods for human tegumentary leishmaniasis in the last 10 years: Technological prospecting.

Leishmaniasis is a neglected disease and more than 1 billion people live in endemic areas with the risk of infection worldwide. Although it is an important epidemiological issue, the gold standard method of diagnosis requires invasive sample collection and is accompanied by a high level of sensitivity variation in results. The present study aims to conduct a patent prospection of immunodiagnostic methods for human tegumentary leishmaniasis in the last 10 years, focused on those with high sensitivity and specificity, and simple usability. We searched seven patent databases: The LENS, WIPO, EPO, USPTO, Patent Inspiration, Google patents, and INPI. Eleven patents were found that satisfy our search criteria, with six of them being registered in 2017. Most patents were registered in Brazil. The information obtained here covers the main characteristics of the immunodiagnostic methods evaluated. Moreover, our prospective study reveals the latest biotechnological advancements achieved in the immunodiagnosis of tegumentary leishmaniasis, especially in Brazil, which holds the majority of patents in this subject. However, no patent for immunodiagnostic methods was found in the last three years, which raises concerns about the present and future trends of leishmaniasis diagnosis.

LQM10, a guanylhydrazone derivative, reduces nociceptive and inflammatory responses in mice.

In the present study, we examined the antinociceptive and anti-inflammatory activities of a guanylhydrazone derivative, (E)-(3,5-di-tert-butyl-4-hydroxybenzylidene)-2-guanylhydrazone hydrochloride (LQM10), in mice. The antinociceptive effect was determined by assessing behavioural responses in different pain models, while anti-inflammatory activity was examined in carrageenan-induced pleurisy. Intraperitoneal LQM10 administration reduced the acetic acid-induced nociceptive behaviour, a phenomenon that was unaltered by pretreatment with yohimbine, atropine, naloxone or glibenclamide. In the formalin assay, LQM10 reduced nociceptive behaviour only in the second phase, indicating an inhibitory effect on inflammatory pain. LQM10 did not alter the pain latency in the hot plate assay and did not impact the locomotor activity of mice in the rotarod assay. In the carrageenan-induced pleurisy assay, LQM10 treatment inhibited critical events involved in inflammatory responses, namely, leucocyte recruitment, plasma leakage and increased inflammatory mediators (tumour necrosis factor Like Properties of Chalchones and Flavonoid Derivatives [TNF]-α and interleukin [IL]-1ß) in the pleural exudate. Overall, these results indicate that LQM10 exhibits antinociceptive effects associated with peripheral mechanisms and anti-inflammatory activity mediated via a reduction in leucocyte migration and proinflammatory mediators, rendering this compound a promising candidate for treating pain and inflammatory process.

Pyroligneous extracts with therapeutic action: A technological prospect

Abstract The Pyroligneous extract is a product from the combustion of plant biomass with applications in the fields of health, industrial chemistry, and agriculture. The discovery of new molecules with therapeutic potential and of natural origin continues to be one of the great challenges for research centres around the world. The following work aims to analyze, through a technological prospection, the use of pyroligneous extracts for therapeutic purposes. To carry out the study, searches were carried out in documents deposited in Brazil, Europe, and the United States and searched on platforms specialized in patents. The number of inventions using pyroligneous extract with therapeutic applications is still quite small, however, innovations have been observed for the treatment of diseases of great clinical relevance such as cancer and hypertension. The systematic mapping of innovations is of great importance for the development of new technologies.

Obstacles to Glioblastoma Treatment Two Decades after Temozolomide.

Glioblastomas are considered the most common and aggressive primary brain tumor in adults, with an average of 15 months' survival rate. The treatment is surgery resection, followed by chemotherapy with temozolomide, and/or radiotherapy. Glioblastoma must have wild-type IDH gene and some characteristics, such as TERT promoter mutation, EGFR gene amplification, microvascular proliferation, among others. Glioblastomas have great heterogeneity at cellular and molecular levels, presenting distinct phenotypes and diversified molecular signatures in each tumor mass, making it difficult to define a specific therapeutic target. It is believed that the main responsibility for the emerge of these distinct patterns lies in subcellular populations of tumor stem cells, capable of tumor initiation and asymmetric division. Studies are now focused on understanding molecular mechanisms of chemoresistance, the tumor microenvironment, due to hypoxic and necrotic areas, cytoskeleton and extracellular matrix remodeling, and in controlling blood brain barrier permeabilization to improve drug delivery. Another promising therapeutic approach is the use of oncolytic viruses that are able to destroy specifically glioblastoma cells, preserving the neural tissue around the tumor. In this review, we summarize the main biological characteristics of glioblastoma and the cutting-edge therapeutic targets that are currently under study for promising new clinical trials.

Antarctic organisms as a source of antimicrobial compounds: a patent review.

Currently, antimicrobial resistance has become a global public health problem, which has made the need for new antimicrobial compounds to deal with resistant infections an emergency. However, environments that once offered so many innovative molecules, now already exhaustively exploited, do not meet this need. In this context, a geographically isolated, under-explored and extreme environment, such as Antarctica, which holds organisms with unique physiological and biochemical characteristics, assumes great importance as a potential source of new compounds with antimicrobial activity. In this patent review, we investigate the state of technological development in the field of antimicrobial compounds obtained from Antarctic organisms, highlighting the main countries and researchers active in the field, the species utilized, the compounds obtained, and their possible therapeutic applications. As results, few patent documents were found, however they encompass a wide diversity of compounds and species, indicating a great antimicrobial potential present in Antarctic biota, including compounds active against the most important human pathogenic microorganisms, such as including methicillin-resistant Staphylococcus aureus, vancomycin-resistant Enterococcus spp. and multi-resistant Mycobacterium tuberculosis. Furthermore, due to the increasing trend in patent applications, a significant rise in the number of patents in this area is expected in the coming years.

Can inflammasomes promote the pathophysiology of glioblastoma multiforme? A view about the potential of the anti-inflammasome therapy as pharmacological target.

Glioblastoma multiforme (GBM) is a malignant brain tumor with one of the worst general survivorship cases among the existing neoplasia. This aggressiveness is due to its complex molecular heterogeneity, immunohistochemistry and genetics. The current therapeutic approach brings little contribution to the improvement of the survival of the patients. Due to that, new forms of treatment have been explored, one of them being immunotherapy. In this aspect, the inflammasome pathway, which induces inflammation and immunosuppressive tumor response, contributing to the progression of the tumor, seems to be a new alternative to improve the treatment efficacy and the survival of the patients.

Antarctic environments as a source of bacterial and fungal therapeutic enzymes.

Microbial therapeutic enzymes are the protagonists in the pharmacological treatment of different human diseases. The intrinsic enzymatic characteristics, such as high affinity and specificity to the corresponding substrate, enable effective therapies, with minimal adverse effects and complete remission. However, immunogenicity, short half-life, low enzymatic yield, and low selectivity regarding available enzyme drugs are currently the main obstacles to their development and the broad adherence to therapeutic protocols. By harboring adapted and still unexplored microbial life, environments of extreme conditions, such as Antarctica, become especially important in the prospecting and development of new enzymatic compounds that present higher yields and the possibility of genetic improvement. Antarctic microorganisms have adaptation mechanisms, such as more fluid cell membranes, production of antifreeze proteins and enzymes with more malleable structures, more robust, stable, selective catalytic sites for their respective substrates, and high antioxidant capacity. In this context, this review aims to explore enzymes synthesized by bacteria and fungi from Antarctica as potential drug producers, capable of providing therapeutic efficacy, less adverse effects, and lower production costs with highlight to L-Asparaginase, collagenase, superoxide dismutase and ribonucleases. In addition, this review highlights the unique biotechnological profile of these Antarctic extremophile microorganisms.

Leishmanicidal activity of Morita-Baylis-Hillman adducts.

Leishmaniasis is a neglected disease that affects millions of people, mostly in developing countries. Although this disease has a high impact on public health, there are few drug options to treat the different leishmaniasis forms. Additionally, these current therapies have various adverse effects, including gastrointestinal disturbances, headache, pancreatitis, and hepatotoxicity. Thus, it is essential to develop new drug prototypes to treat leishmaniasis. Accordingly, the present study aimed to evaluate the leishmanicidal activity of Morita-Baylis-Hillman adducts and their O-acetylates, carboxylic acid derivatives, and acid and ester derivatives of 2-methyl-phenylpropanoids against Leishmania chagasi. Initially, we evaluated the cytotoxicity of 16 derivatives (1-16G) against J774A.1 macrophages. Eight derivatives (2G, 4G, 5G, 7G, 9G, 10G, 13G, and 15G) showed no cytotoxicity at up to the maximum concentration tested (100 µM). When evaluated for antileishmanial effect against promastigote forms, 1G, 6G, 8G, 10G, 11G, 13G, 14G, 15G, and 16G displayed significant toxicity compared to the control (0.1% DMSO). Additionally, the compounds 1G, 5G, 7G, 9G, 11G, 13G, 14G, and 16G reduced macrophage infection by amastigotes. Thus, we conclude that these derivatives have antileishmanial effects, particularly 1G, which showed activity against promastigotes and amastigotes, and low toxicity against macrophages.

LASSBio-596: a New Pre-clinical Candidate for Rheumatoid Arthritis?

Pain and inflammatory disorders are significant health problems because of prevalence and associated disabilities. In this context, LASSBio-596 is a hybrid compound able to modulate TNF-α and phosphodiesterases 4 and 5, exhibiting an anti-inflammatory effect in the pulmonary inflammatory model. Aiming at a better description of the activities of LASSBio-596, we initially conducted nociception tests (acetic acid-induced abdominal writhing, glutamate, and formalin-induced nociception and hot plate test) and later inflammatory tests (acute, peritonitis; and chronic, arthritis) that directed us to this last one. In the abdominal writhing test, there was a dose-dependent inhibition, whose response occurred at the maximum dose (50 mg/kg, p.o.), used in the subsequent tests. LASSBio-596 also inhibited nociception induced by chemical (glutamate by 31.9%; and formalin, in both phases, 1st phase: 25.7%; 2nd phase: 23.9%) and thermal agents (hotplate, by increased latency for pain at two different times). These effects were independent of the motor function, legitimated in rotarod. As there was a response in the inflammatory component of nociception, we performed the peritonitis test, in which migration was inhibited by LASSBio-596 by 39.9%. As the inflammatory process is present in autoimmune diseases, we also performed the arthritis test. LASSBio-596 reduced paw edema from the 15th day to the 21st day of treatment (no liver changes and with fewer paw injuries). In addition, LASSBio-596 decreased serum levels of TNF-α by 67.1%. These data demonstrated the antinociceptive effect of LASSBio-596 and reinforces its anti-inflammatory property (i.e., RA), amplifying the therapeutic potential of this molecule.

Synthesis, Antileishmanial Activity and in silico Studies of Aminoguanidine Hydrazones (AGH) and Thiosemicarbazones (TSC) Against Leishmania chagasi Amastigotes.

BACKGROUND: Leishmaniasis is a worldwide health problem, highly endemic in developing countries. Among the four main clinical forms of the disease, visceral leishmaniasis is the most severe, fatal in 95% of cases. The undesired side-effects from first-line chemotherapy and the reported drug resistance search for effective drugs that can replace or supplement those currently used in an urgent need. Aminoguanidine hydrazones (AGH's) have been explored for exhibiting a diverse spectrum of biological activities, in particular the antileishmanial activity of MGBG. The bioisosteres thiosemicarbazones (TSC's) offer a similar biological activity diversity, including antiprotozoal effects against Leishmania species and Trypanosoma cruzi. OBJECTIVES: Considering the impact of leishmaniasis worldwide, this work aimed to design, synthesize, and perform a screening upon L. chagasi amastigotes and for the cytotoxicity of the small "inhouse" library of both AGH and TSC derivatives and their structurally-related compounds. METHODS: A set of AGH's (3-7), TSC's (9, 10), and semicarbazones (11) were initially synthesized. Subsequently, different semi-constrained analogs were designed and also prepared, including thiazolidines (12), dihydrothiazines (13), imidazolines (15), pyrimidines (16, 18) azines (19, 20), and benzotriazepinones (23-25). All intermediates and target compounds were obtained with satisfactory yields and exhibited spectral data consistent with their structures. All final compounds were evaluated against L. chagasi amastigotes and J774.A1 cell line. Molecular docking was performed towards trypanothione reductase using GOLD® software. RESULTS: The AGH's 3i, 4a, and 5d, and the TSC's 9i, 9k, and 9o were selected as valuable hits. These compounds presented antileishmanial activity compared with pentamidine, showing IC50 values ranged from 0.6 to 7.27 µM, maximal effects up to 55.3%, and satisfactory SI values (ranged from 11 to 87). On the other hand, most of the resulting semi-constrained analogs were found cytotoxic or presented reduced antileishmanial activity. In general, TSC class is more promising than its isosteric AGH analogs, and the beneficial aromatic substituent effects are not similar in both series. In silico studies have suggested that these hits are capable of inhibiting the trypanothione reductase from the amastigote forms. CONCLUSION: The promising antileishmanial activity of three AGH's and three TSC's was characterized. These compounds presented antileishmanial activity compared with PTD, showing IC50 values ranged from 0.6 to 7.27 µM, and satisfactory SI values. Further pharmacological assays involving other Leishmania strains are in progress, which will help choose the best hits for in vivo experiments.

Fibrinolytic Enzymes From Extremophilic Microorganisms in the Development of New Thrombolytic Therapies: Technological Prospecting.

Extremophilic microorganisms from a wide variety of extreme natural environments have been researched, and many biotechnological applications have been carried out, due to their capacity to produce biomolecules resistant to extreme conditions, such as fibrinolytic proteases. The search for new fibrinolytic enzymes is important in the development of new therapies against cardiovascular diseases. This article aimed to evaluate the patents filed about protease with fibrinolytic activity produced by extremophilic microorganisms whose use is aimed at the development of new drugs for the treatment of cardiovascular diseases. The prospecting was carried out using data on deposits and patent concessions made available on the technological bases: European Patent Office (EPO), United States Patent and Trademark Office (USPTO), World Intellectual Property Organization (WIPO), Instituto Nacional de Propriedade Industrial - Brazil (INPI), The LENS and Patent Inspiration. The International Patent Classification and subclasses and groups for each document were also evaluated. Although 382 patents were selected using terms related to extreme environments, such as "thermophile" and "acidophiles", few were related to clinical use and were mainly performed using Bacillus subtilis and Streptomyces megasporus strains. A highlight of nattokinase was produced by Bacillus subtilis GDN and actinokinase by Streptomyces megasporus SD5. The low number of patents on enzymes with this profile (extreme environments) revealed a little-explored field, promising in the development of new microbial thrombolytic drugs, such as fibrinolytic enzymes with less adverse effects.

Technological Prospecting: Mapping Patents on L-asparaginases from Extremophilic Microorganisms.

BACKGROUND: L-asparaginase (L-ASNase, L-asparagine amidohydrolase, E.C.3.5.1.1) is an enzyme with wide therapeutic applicability. Currently, the commercialized L-ASNase comes from mesophilic organisms, presenting low specificity to the substrate and limitations regarding thermostability and active pH range. Such factors prevent the maximum performance of the enzyme in different applications. Therefore, extremophilic organisms may represent important candidates for obtaining amidohydrolases with particular characteristics desired by the biotechnological market. OBJECTIVES: The present study aims to carry out a technological prospecting of patents related to the L-asparaginases derived from extremophilic organisms, contributing to pave the way for further rational investigation and application of such enzymes. METHODS: This patent literature review used six patents databases: The LENS, WIPO, EPO, USPTO, Patent Inspiration, and INPI. RESULTS: It was analyzed 2860 patents, and 14 were selected according to combinations of descriptors and study criteria. Approximately 57.14% of the patents refer to enzymes obtained from archaea, especially from the speciesPyrococcus yayanosii (35.71% of the totality). CONCLUSION: The present prospective study has singular relevance since there are no recent patent reviews for L-asparaginases, especially produced by extremophilic microorganisms. Although such enzymes have well-defined applications, corroborated by the patents compiled in this review, the most recent studies allude to new uses, such as the treatment of infections. The characterization of the catalytic profiles allows us to infer that there are potential sources still unexplored. Hence, the search for new L-ASNases with different characteristics will continue to grow in the coming years and, possibly, ramifications of the technological routes will be witnessed.

Design and Synthesis In Silico Drug-like Prediction and Pharmacological Evaluation of Cyclopolymethylenic Homologous of LASSBio-1514.

Acylhydrazones are still an important framework to the design of new bioactive compounds. As treatment of chronic pain represents a clinical challenge, we decided to modify the structure of LASSBio-1514 (1), previously described as anti-inflammatory and analgesic prototype. Applying the homologation as a strategy for molecular modification, we designed a series of cyclopentyl- (2a-e), cyclobutyl- (3a-e), and cyclopropylacylhydrazones (4a-e) that were synthetized and evaluated in murine models of inflammation and pain. A comparison of their in silico physicochemical and drug-like profile was conducted, as well as their anti-inflammatory and analgesic effect. Compounds 4a (LASSBio-1755) and 4e (LASSBio-1757) displayed excellent in silico drug-like profiles and were identified as new analgesic lead-candidates in acute and chronic model of pain, through oral administration.

Leismanicidal Activity of Propolis Collected in the Semiarid Region of Brazil.

Objective: The aim of the current study is to investigate the chemical composition, cytotoxic effect, and leishmanicidal activity of propolis collected in the semi-arid region of Bahia, Brazil. Methods: EtOH extract, hexane, EtOAc and MeOH fractions from propolis were analyzed by ultra-performance liquid chromatography coupled with diode array detector and quadrupole time-of-flight mass spectrometry. The identification was based on the exact mass, general fragmentation behaviors and UV absorption of the flavonoids. The in vitro cytotoxic effect and leishmanicidal activity of ethanolic extract, hexane, ethyl acetate, and methanolic fractions of propolis were evaluated. Results: Five triterpenes and twenty-four flavonoids were identified. The propolis did not present toxicity to the host cell up to the maximum concentration tested. In addition, all tested samples showed statistically significant activity against promastigotes of Leishmania chagasi and Leishmania amazonensis. Regarding the activity against amastigote forms of L. amazonensis, the hexane fraction, presented statistically significant activity with IC50 of 1.3 ± 0.1 µg/ml. Conclusion: The results support the idea that propolis can be used for future antileishmania studies.

Pigments from Antarctic bacteria and their biotechnological applications.

Pigments from microorganisms have triggered great interest in the market, mostly by their "natural" appeal, their favorable production conditions, in addition to the potential new chemical structures or naturally overproducing strains. They have been used in: food, feed, dairy, textile, pharmaceutical, and cosmetic industries. The high rate of pigment production in microorganisms recovered from Antarctica in response to selective pressures such as: high UV radiation, low temperatures, and freezing and thawing cycles makes this a unique biome which means that much of its biological heritage cannot be found elsewhere on the planet. This vast arsenal of pigmented molecules has different functions in bacteria and may exhibit different biotechnological activities, such as: extracellular sunscreens, photoprotective function, antimicrobial activity, biodegradability, etc. However, many challenges for the commercial use of these compounds have yet to be overcome, such as: the low stability of natural pigments in cosmetic formulations, the change in color when subjected to pH variations, the low yield and the high costs in their production. This review surveys the different types of natural pigments found in Antarctic bacteria, classifying them according to their chemical structure. Finally, we give an overview of the main pigments that are used commercially today.

Costus spiralis (Jacq.) Roscoe leaves fractions have potential to reduce effects of inflammatory diseases.

ETHNOPHARMACOLOGICAL RELEVANCE: Since drugs currently used to manage pain and inflammatory conditions present several side effects, the investigation of new anti-inflammatory and antinociceptive agents from folk-medicine plants is an important approach. Costus spiralis (Costaceae) has been used in Brazilian medicinal teas to treat urinary infection, cough, inflammation, arthritis, among others. AIM OF THE STUDY: The current study focused on investigating anti-inflammatory and antinociceptive effects of fractions from C. spiralis leaves using animal models. MATERIALS AND METHODS: Adults Swiss mice were used in the following experimental models: acetic acid-induced abdominal writhing, formalin-induced nociception, hot plate, zymosan-induced peritonitis, and arthritis induced by complete Freund's adjuvant. RESULTS: The presence of steroids was confirmed in all fractions. Flavonoids, condensed tannins and saponins were observed in EFL. In methanolic fraction leaves (MFL), the presence of flavonoids and pentacyclic triterpenoids was confirmed. Orally administered leaf fractions significantly reduced abdominal writhing. Fractions were ineffective in the neurogenic stage of the formalin test, but in the inflammatory stage, ethyl acetate fraction levaes (AcFL), ethanolic fraction leaves (EFL), and MFL significantly reduced paw licking time by 69.6 ± 11.9%, 58.2 ± 9.4%, and 79.6 ± 8.3%, respectively. In the hot plate test, the reaction latency was similar for treated animals and controls. However, in the peritonitis test, cell migration was significantly reduced in animals treated with chloroform fractions leaves ClFL (61.8 ± 11.4%), AcFL (58.7 ± 8.3%), EFL (39.2 ± 5.0%), and MFL (64.8 ± 4.4%). This was similar to the result observed in the chronic inflammation model, this time only the chloroform fraction was able to reduce paw edema. CONCLUSION: Our results show that leaf fractions of Costus spiralis are capable of modulating peripheral nociceptive and inflammatory responses without effects on central nervous system being potential substrates for phytochemical purification, structural and mechanistic studies.

Citrus limon (L.) Burm f. Essential Oil Has Anxiolytic and Sedative Properties by Modulating GABAA-Receptors

Abstract The high prevalence of anxiety disorders associated with pharmacotherapy side effects have motivated the search for new pharmacological agents. Species from Citrus genus, such as Citrus limon (sicilian lemon), have been used in folk medicine as a potential therapy to minimize emotional disorders. In order to searching for new effective treatments with fewer side effects, the present study evaluated the anxiolytic mechanism of action and the hypnotic-sedative activity from the Citrus limon fruit's peels essential oil (CLEO). Adults male Swiss mice were submitted to barbiturate-induced sleep test; elevated plus-maze (EPM) and light-dark box (LDB) (evaluation of the mechanism of action); rotarod; and catalepsy tests. CLEO oral treatment decreased latency and increased the sleep total time; moreover it induced in animals an increased the number of entries and percentage of time spent into open arms of the EPM; an increased the number of transitions and the percentage of time into light compartment in the LDB; which were only antagonized by flumazenil pretreatment, with no injury at motor function. Thus, results suggest that CLEO treatment induced an anxiolytic behavior suggestively modulated by the benzodiazepine binding site of the GABAA receptor or by an increase of GABAergic neurotransmission, without cause impairment in the motor coordination.