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Biotechnology Advances Nov 2020Limonene and its derivatives have great market potential with diverse applications in food, pharmaceuticals, cosmetics, etc. Commercial production of limonene and its... (Review)
Review
Limonene and its derivatives have great market potential with diverse applications in food, pharmaceuticals, cosmetics, etc. Commercial production of limonene and its derivatives through extraction from plants suffers from the unstable market supply, while chemical synthesis of these compounds is hindered by high energy consumption and pollutant emission. Microbial biosynthesis provides a promising alternative approach for the sustainable supply of limonene and its derivatives. However, low efficiency and specificity of the biosynthetic enzymes and pathways in heterologous hosts make it still challenging for the commercialization of microbial limonene production. On the other hand, the limonene toxicity heavily reduces cellular fitness, which poses a serious challenge for improving limonene titer. Here, we critically review the recent progresses in engineering microbes for limonene biosynthesis and derivation with the emphasis on enzyme characterization and pathway optimization. In particular, we introduce the current trends in microbial limonene decoration for the biosynthesis of bio-active molecules such as α-terpineol and perillyl alcohol. We also discuss the feasible strategies for relieving limonene toxicity and enhancing the robustness of microbial cell factories.
Topics: Cyclohexenes; Limonene; Metabolic Engineering
PubMed: 32882371
DOI: 10.1016/j.biotechadv.2020.107628 -
PloS One 2020The prognosis for patients with glioblastoma (GB) remains grim. Concurrent temozolomide (TMZ) radiation-the cornerstone of glioma control-extends the overall median...
The prognosis for patients with glioblastoma (GB) remains grim. Concurrent temozolomide (TMZ) radiation-the cornerstone of glioma control-extends the overall median survival of GB patients by only a few months over radiotherapy alone. While these survival gains could be partly attributed to radiosensitization, this benefit is greatly minimized in tumors expressing O6-methylguanine DNA methyltransferase (MGMT), which specifically reverses O6-methylguanine lesions. Theoretically, non-O6-methylguanine lesions (i.e., the N-methylpurine adducts), which represent up to 90% of TMZ-generated DNA adducts, could also contribute to radiosensitization. Unfortunately, at concentrations attainable in clinical practice, the alkylation capacity of TMZ cannot overwhelm the repair of N-methylpurine adducts to efficiently exploit these lesions. The current therapeutic application of TMZ therefore faces two main obstacles: (i) the stochastic presence of MGMT and (ii) a blunted radiosensitization potential at physiologic concentrations. To circumvent these limitations, we are developing a novel molecule called NEO212-a derivatization of TMZ generated by coupling TMZ to perillyl alcohol. Based on gas chromatography/mass spectrometry and high-performance liquid chromatography analyses, we determined that NEO212 had greater tumor cell uptake than TMZ. In mouse models, NEO212 was more efficient than TMZ at crossing the blood-brain barrier, preferentially accumulating in tumoral over normal brain tissue. Moreover, in vitro analyses with GB cell lines, including TMZ-resistant isogenic variants, revealed more potent cytotoxic and radiosensitizing activities for NEO212 at physiologic concentrations. Mechanistically, these advantages of NEO212 over TMZ could be attributed to its enhanced tumor uptake presumably leading to more extensive DNA alkylation at equivalent dosages which, ultimately, allows for N-methylpurine lesions to be better exploited for radiosensitization. This effect cannot be achieved with TMZ at clinically relevant concentrations and is independent of MGMT. Our findings establish NEO212 as a superior radiosensitizer and a potentially better alternative to TMZ for newly diagnosed GB patients, irrespective of their MGMT status.
Topics: Animals; Blood-Brain Barrier; Brain; Brain Neoplasms; Cell Line, Tumor; Cell Survival; DNA Damage; Dacarbazine; Drug Resistance, Neoplasm; Gas Chromatography-Mass Spectrometry; Glioma; Humans; Mice; Mice, Inbred C57BL; O(6)-Methylguanine-DNA Methyltransferase; Radiation-Sensitizing Agents; Temozolomide; Xenograft Model Antitumor Assays
PubMed: 32881880
DOI: 10.1371/journal.pone.0238238 -
Neuro-oncology Jan 2021Intracarotid injection of mannitol has been applied for decades to support brain entry of therapeutics that otherwise do not effectively cross the blood-brain barrier...
BACKGROUND
Intracarotid injection of mannitol has been applied for decades to support brain entry of therapeutics that otherwise do not effectively cross the blood-brain barrier (BBB). However, the elaborate and high-risk nature of this procedure has kept its use restricted to well-equipped medical centers. We are developing a more straightforward approach to safely open the BBB, based on the intra-arterial (IA) injection of NEO100, a highly purified version of the natural monoterpene perillyl alcohol.
METHODS
In vitro barrier permeability with NEO100 was evaluated by transepithelial/transendothelial electrical resistance and antibody diffusion assays. Its mechanism of action was studied by western blot, microarray analysis, and electron microscopy. In mouse models, we performed ultrasound-guided intracardiac administration of NEO100, followed by intravenous application of Evan's blue, methotrexate, checkpoint-inhibitory antibodies, or chimeric antigen receptor (CAR) T cells.
RESULTS
NEO100 opened the BBB in a reversible and nontoxic fashion in vitro and in vivo. It enabled greatly increased brain entry of all tested therapeutics and was well tolerated by animals. Mechanistic studies revealed effects of NEO100 on different BBB transport pathways, along with translocation of tight junction proteins from the membrane to the cytoplasm in brain endothelial cells.
CONCLUSION
We envision that this procedure can be translated to patients in the form of transfemoral arterial catheterization and cannulation to the cerebral arteries, which represents a low-risk procedure commonly used in a variety of clinical settings. Combined with NEO100, it is expected to provide a safe, widely available approach to enhance brain entry of any therapeutic.
Topics: Animals; Blood-Brain Barrier; Brain; Endothelial Cells; Humans; Mice; Monoterpenes; Tight Junctions
PubMed: 32877532
DOI: 10.1093/neuonc/noaa206 -
Circulation Research Oct 2020Brain arteriovenous malformations (AVMs) are abnormal tangles of vessels where arteries and veins directly connect without intervening capillary nets, increasing the...
RATIONALE
Brain arteriovenous malformations (AVMs) are abnormal tangles of vessels where arteries and veins directly connect without intervening capillary nets, increasing the risk of intracerebral hemorrhage and stroke. Current treatments are highly invasive and often not feasible. Thus, effective noninvasive treatments are needed. We previously showed that AVM-brain endothelial cells (BECs) secreted higher VEGF (vascular endothelial growth factor) and lower TSP-1 (thrombospondin-1) levels than control BEC; and that microRNA-18a (miR-18a) normalized AVM-BEC function and phenotype, although its mechanism remained unclear.
OBJECTIVE
To elucidate the mechanism of action and potential clinical application of miR-18a as an effective noninvasive treatment to selectively restore the phenotype and functionality of AVM vasculature.
METHODS AND RESULTS
The molecular pathways affected by miR-18a in patient-derived BECs and AVM-BECs were determined by Western blot, RT-qPCR (quantitative reverse transcription polymerase chain reaction), ELISA, co-IP, immunostaining, knockdown and overexpression studies, flow cytometry, and luciferase reporter assays. miR-18a was shown to increase TSP-1 and decrease VEGF by reducing PAI-1 (plasminogen activator inhibitor-1/SERPINE1) levels. Furthermore, miR-18a decreased the expression of BMP4 (bone morphogenetic protein 4) and HIF-1α (hypoxia-inducible factor 1α), blocking the BMP4/ALK (activin-like kinase) 2/ALK1/ALK5 and Notch signaling pathways. As determined by Boyden chamber assays, miR-18a also reduced the abnormal AVM-BEC invasiveness, which correlated with a decrease in MMP2 (matrix metalloproteinase 2), MMP9, and ADAM10 (ADAM metallopeptidase domain 10) levels. In vivo pharmacokinetic studies showed that miR-18a reaches the brain following intravenous and intranasal administration. Intranasal co-delivery of miR-18a and NEO100, a good manufacturing practices-quality form of perillyl alcohol, improved the pharmacokinetic profile of miR-18a in the brain without affecting its pharmacological properties. Ultra-high-resolution computed tomography angiography and immunostaining studies in an Mgp AVM mouse model showed that miR-18a decreased abnormal cerebral vasculature and restored the functionality of the bone marrow, lungs, spleen, and liver.
CONCLUSIONS
miR-18a may have significant clinical value in preventing, reducing, and potentially reversing AVM.
Topics: ADAM10 Protein; Activin Receptors, Type I; Activin Receptors, Type II; Amyloid Precursor Protein Secretases; Animals; Bone Morphogenetic Protein 4; Brain; Endothelial Cells; Humans; Hypoxia-Inducible Factor 1, alpha Subunit; Intracranial Arteriovenous Malformations; Matrix Metalloproteinase 2; Matrix Metalloproteinase 9; Membrane Proteins; Mice; MicroRNAs; Monoterpenes; Plasminogen Activator Inhibitor 1; Receptor, Transforming Growth Factor-beta Type I; Thrombospondin 1; Vascular Endothelial Growth Factors
PubMed: 32755283
DOI: 10.1161/CIRCRESAHA.119.316317 -
BMC Complementary Medicine and Therapies Jul 2020Pulmonary artery hypertension (PAH) is a vascular disease in the lung characterized by elevated pulmonary arterial pressure (PAP). Many miRNAs play a role in the...
Perillyle alcohol and Quercetin ameliorate monocrotaline-induced pulmonary artery hypertension in rats through PARP1-mediated miR-204 down-regulation and its downstream pathway.
BACKGROUND
Pulmonary artery hypertension (PAH) is a vascular disease in the lung characterized by elevated pulmonary arterial pressure (PAP). Many miRNAs play a role in the pathophysiology of PAH. Perillyle alcohol (PA) and Quercetin (QS) are plant derivatives with antioxidant and anti-proliferative properties. We investigated the effect of PA and QS on PAP, expression of PARP1, miR-204, and their targets, HIF1α and NFATc2, in experimental PAH.
METHODS
Thirty rats were divided into control, MCT, MCT + Veh, MCT + PA and MCT + QS groups. MCT (60 mg/kg) was injected subcutaneously to induce PAH. PA (50 mg/kg daily) and QS (30 mg/kg daily) were administered for 3 weeks after inducing PAH. PAP, lung pathology, expression of miRNA and mRNA, and target proteins were evaluated through right ventricle cannulation, H&E staining, real-time qPCR, and western blotting, respectively.
RESULTS
Inflammation and lung arteriole thickness in the MCT group increased compared to control group. PA and QS ameliorated inflammation and reduced arteriole thickness significantly. miR-204 expression decreased in PAH rats (p < 0.001). PA (p < 0.001) and QS (p < 0.01) significantly increased miR-204 expression. Expression of PARP1, HIF1α, NFATc2, and α-SMA mRNA increased significantly in MCT + veh rats (all p < 0.001), and these were reduced after treatment with PA and QS (both p < 0.01). PA and QS also decreased the expression of PARP1, HIF1α, and NFATc2 proteins that had increased in MCT + Veh group.
CONCLUSION
PA and QS improved PAH possibly by affecting the expression of PARP1 and miR-204 and their downstream targets, HIF1a and NFATc2. PA and QS may be therapeutic goals in the treatment of PAH.
Topics: Animals; Disease Models, Animal; Down-Regulation; Hypertension, Pulmonary; Hypertrophy, Right Ventricular; Hypoxia-Inducible Factor 1, alpha Subunit; Male; MicroRNAs; Monocrotaline; Monoterpenes; NFATC Transcription Factors; Poly (ADP-Ribose) Polymerase-1; Pulmonary Artery; Quercetin; Rats; Rats, Wistar
PubMed: 32660602
DOI: 10.1186/s12906-020-03015-1 -
International Immunopharmacology Sep 2020Limonene (LIM) and its main metabolite perillyl alcohol (POH) are ingredients found in food with promising chemical entities due to their pharmacological profile. In...
Limonene, a food additive, and its active metabolite perillyl alcohol improve regeneration and attenuate neuropathic pain after peripheral nerve injury: Evidence for IL-1β, TNF-α, GAP, NGF and ERK involvement.
BACKGROUND
Limonene (LIM) and its main metabolite perillyl alcohol (POH) are ingredients found in food with promising chemical entities due to their pharmacological profile. In this study, we hypothesized that LIM and POH are two molecules capable of accelerating the regenerative process and alleviating neuropathic pain.
METHODS
Animals were treated daily (LIM, POH and saline) for 28 days and during this period evaluated for mechanical hyperalgesia, astrocyte participation by immunofluorescence for GFAP, and ELISA was used to quantify IL-1β and TNF-α in the spinal cord. Western blot analysis of the following proteins was also performed: GFAP, GAP-43, NGF and ERK. For motor deficit analysis, tests were performed to assess hind paw muscle strength and footprints through gait (SFI).
RESULTS
Both POH and LIM accelerated the regenerative process and improved motor deficits comparing to positive control; however, POH was more effective, particularly between the 2nd and 3rd week after the nerve injury, increasing GAP-43, NGF and the phosphorylated ERK immunocontent. Moreover, POH and LIM were able to reduce hyperalgesia and astrocytosis.
CONCLUSIONS
Both substances, LIM and POH, improved the regeneration process and sensory and motor function recovery in the PNI model in mice by mitigating the inflammatory reactions and up-regulating the neurotrophic process.
Topics: Animals; Anti-Inflammatory Agents; Disease Models, Animal; Extracellular Signal-Regulated MAP Kinases; Food Additives; GTPase-Activating Proteins; Humans; Interleukin-1beta; Limonene; Male; Mice; Monoterpenes; Motor Neurons; Nerve Growth Factor; Neuralgia; Peripheral Nerve Injuries; Regeneration; Tumor Necrosis Factor-alpha
PubMed: 32652504
DOI: 10.1016/j.intimp.2020.106766 -
Molecules (Basel, Switzerland) Jun 2020The interaction between a drug molecule and its carrier's components is an important factor which influences the drug release profile. For this purpose, molecular...
The interaction between a drug molecule and its carrier's components is an important factor which influences the drug release profile. For this purpose, molecular dynamics (MD) may be the in silico tool which can help to understand the mechanism of drug loading/release. The aim of this work is to explain the effect of interactions between different types of terpenes, namely perillyl alcohol, forskolin, ursolic acid, and the nanoemulsion droplet core, on the release by means of experimental and theoretical studies. The basic nanoemulsion was composed of caprylic/capric triglyceride as the oil phase, polysorbate 80 as the emulsifier, and water. The in vitro release tests from a terpene-loaded nanoemulsion were carried out to determine the release profiles. The behavior of terpenoids in the nanoemulsion was also theoretically investigated using the molecular dynamics method. The forskolin-loaded nanoemulsion showed the highest percentage of drug release (almost 80% /) in contrast to ursolic acid and perillyl alcohol-loaded nanoemulsions (about 53% / and 19% /, respectively). The results confirmed that the kinetic model of release was terpene-type dependent. The zero-order model was the best to describe the ursolic acid release profile, while the forskolin and the perillyl alcohol followed a first-order and Higuchi model, respectively. Molecular dynamics simulations, especially energetical analysis, confirmed that the driving force of terpenes diffusion from nanoemulsion interior was their interaction energy with a surfactant.
Topics: Emulsifying Agents; Emulsions; Kinetics; Models, Chemical; Nanostructures; Polysorbates; Terpenes
PubMed: 32545817
DOI: 10.3390/molecules25122747 -
Neurotoxicity Research Aug 2020In this study, we aim to assess the phytomedicinal potential of perillyl alcohol (PA), a dietary monoterpenoid, in a unilateral 6-hydroxydopamine (6-OHDA) lesion rat...
Perillyl Alcohol Mitigates Behavioural Changes and Limits Cell Death and Mitochondrial Changes in Unilateral 6-OHDA Lesion Model of Parkinson's Disease Through Alleviation of Oxidative Stress.
In this study, we aim to assess the phytomedicinal potential of perillyl alcohol (PA), a dietary monoterpenoid, in a unilateral 6-hydroxydopamine (6-OHDA) lesion rat model of Parkinson's disease (PD). We observed that PA supplementation alleviated behavioural abnormalities such as loss of coordination, reduced rearing and motor asymmetry in lesioned animals. We also observed that PA-treated animals exhibited reduced oxidative stress, DNA fragmentation and caspase 3 activity indicating alleviation of apoptotic cell death. We found reduced mRNA levels of pro-apoptotic regulator BAX and pro-inflammatory mediators IL18 and TNFα in PA-treated animals. Further, PA treatment successfully increased mRNA and protein levels of Bcl2, mitochondrial biogenesis regulator PGC1α and tyrosine hydroxylase (TH) in lesioned animals. We observed that PA treatment blocked BAX and Drp1 translocation to mitochondria, an event often associated with the inception of apoptosis. Further, 6-OHDA exposure reduced expression of electron transport chain complexes I and IV, thereby disturbing energy metabolism. Conversely, expression levels of both complexes were upregulated with PA treatment in lesioned rats. Finally, we found that protein levels of Nrf2, the transcription factor responsible for antioxidant gene expression, were markedly reduced in cytosolic and nuclear fraction on 6-OHDA exposure, and PA increased expression of Nrf2 in both fractions. We believe that our data hints towards PA having the ability to provide cytoprotection in a hemiparkinsonian rat model through alleviation of motor deficits, oxidative stress, mitochondrial dysfunction and apoptosis.
Topics: Animals; Behavior, Animal; Caspase 3; DNA Fragmentation; Dynamins; Electron Transport Complex I; Electron Transport Complex IV; Enzyme Inhibitors; Mitochondria; Monoterpenes; Movement; NF-E2-Related Factor 2; Oxidative Stress; Oxidopamine; Parkinsonian Disorders; Peroxisome Proliferator-Activated Receptor Gamma Coactivator 1-alpha; Proto-Oncogene Proteins c-bcl-2; Rats; Sympatholytics; Tyrosine 3-Monooxygenase; bcl-2-Associated X Protein
PubMed: 32394056
DOI: 10.1007/s12640-020-00213-0 -
Journal of Applied Oral Science :... 2020Natural products have emerged as a rich source of bioactive compounds for adjunctive treatments of many infectious and inflammatory conditions, including periodontitis....
Natural products have emerged as a rich source of bioactive compounds for adjunctive treatments of many infectious and inflammatory conditions, including periodontitis. Among the monoterpenes with significant biological properties, there is the perillyl alcohol (POH), which can be found in several essential oils and has shown immunomodulatory properties in recent studies, which may be interesting in the treatment of non-neoplastic inflammatory disorders. Objective To determine the antibacterial and immune modulatory activities of the POH. Methodology The minimum inhibitory concentration (MIC) and the minimum bactericidal concentration (MBC) of the POH for two significant Gram-negative periodontal pathogens were determined by macrodilution and subculture, respectively. Cell proliferation and cytotoxicity in RAW 264.7 macrophages were determined by Trypan Blue and mitochondrial enzymatic activity assay. The modulation of reactive oxygen species (ROS) was analyzed by flow cytometry and expression of TNF and arginase-1 by real-time PCR. Results The POH was effective against P. gingivalis (ATCC 33277) and F. nucleatum (ATCC 25586) with MIC= MBC=1600 μM. No cytotoxicity up to 100 µM was observed on macrophages. The cell proliferation was inhibited from 48 hours at 100 μM (p<0.05) and 250 μM (p<0.01). The POH increased ROS production at both 10 μM and 100 μM (p<0.05) in unstimulated cells. The PMA-induced ROS production was not affected by POH, whereas 100 μM significantly reduced lipopolysaccharide-induced (LPS-induced) ROS. The expression of TNF was not affected by POH in unstimulated cells or in cells polarized to M1 phenotype, whereas both concentrations of POH reduced (p<0.05) the expression of arginase-1 in M2-polarized macrophages. Conclusion The POH has antibacterial activity against periodontal pathogens and reduced proliferation of murine macrophages without significant cytotoxicity at concentrations up to 100 μM. In addition, the POH reduced the LPS-induced ROS and the expression of arginase-1 in M2-polarized macrophages.
Topics: Animals; Anti-Bacterial Agents; Arginase; Biological Products; Cell Proliferation; Flow Cytometry; Fusobacterium nucleatum; Gene Expression; Lipopolysaccharides; Macrophages; Mice; Microbial Sensitivity Tests; Monoterpenes; Porphyromonas; RAW 264.7 Cells; Reactive Oxygen Species; Real-Time Polymerase Chain Reaction; Reproducibility of Results; Time Factors; Tumor Necrosis Factor-alpha
PubMed: 32348444
DOI: 10.1590/1678-7757-2019-0519 -
ACS Omega Apr 2020In this work, a drug delivery system for perillyl alcohol based on the peptide self-assembly containing 3-(2-benzothiazolyl)-7-(diethylamino)coumarin (C6) as a...
In this work, a drug delivery system for perillyl alcohol based on the peptide self-assembly containing 3-(2-benzothiazolyl)-7-(diethylamino)coumarin (C6) as a fluorescent additive is obtained, and its photophysical characteristics as well as its release dynamics were studied by steady-state and time-resolved fluorescence spectroscopy. Results proved the dynamics of drug release from the peptide nanostructures and showed that the system formed by the self-assembled peptide and C6, along with perillyl alcohol, presents unique photophysical properties that can be exploited to generate singlet oxygen (O) upon irradiation, which is not achieved by the sole components. Through epifluorescence microscopy combined with time-correlated single photon counting fluorescence spectroscopy, the release mechanism was proven to occur upon peptide structure interconversion, which is controlled by environmental changes.
PubMed: 32337442
DOI: 10.1021/acsomega.0c00381