Research Title: A NOVEL BRAZILIAN SPIDER TOXIN ANALOGUE IS ANTINOCICEPTIVE ACTING VIA THE CANNABINOID SYSTEM

Author: Wafa Moh'd Khair Hourani, Published Year: 2018

The 28th Annual International Cannabinoid Research Society Symposium on the Cannabinoids, Leiden university, the Netherlands

Faculty: Pharmacy

Abstract: Peptide toxins produced by a variety of organisms have evolved with different targets, including GPCR, enzymes and ion channels, where there are many examples of ligands with high potency and selectivity. The Brazilian scientific team developed peptide analogues of an active component of the venom of the Brazilian wandering spider (Phoneutria nigriventer). The synthetic nontoxic peptide PnPP-19 (BR 10 2012 020800-8 A2) induced antinociception involving the inhibition of neutral endopeptidase and activation of CB1 cannabinoid receptors as well as µ and δ opioid receptors (Freitas, Br J Pharmacol PMID: 26947933). In this work, a novel synthetic nontoxic peptide, Pn1 (11 amino acid residues, MW= 1.35 kDa, in patent application) modeled from δCtenitoxin-Pn1a toxin was examined. Nociceptive thresholds were measured by the in vivo paw pressure test on male Wistar rats (180- 200 g). First, to investigate the role of Pn1 in nociception, the peptide was injected (1.5, 2.5, 5 and 10 µg per paw) or vehicle (50 µl) into rat paws-that were hyperalgesic following the administration of carrageenan (250 µg). Pn1 induced a dose-dependent antinociceptive response, with the maximal effects at 10 µg per paw, which peaked at 30 min post-administration. The role of the CB1 cannabinoid receptor in these effects was investigated with intraplantar co-administration of AM251 (40, 80 and 160 µg per paw). The highest dose totally inhibited the antinociceptive effect of Pn1 (10 µg per paw). Moreover, we investigated whether enzymatic degradation by FAAH, MAGL and anandamide endocannabinoid transporter pathways could be contributing to the antinociceptive activity using intraplantar administration of JZL184 (14 µg per paw); MAFP, (4 µg per paw); or VDM11 (40 µg per paw). The results showed that inhibition of MAGL or the transporter potentiated the antinociceptive activity of the Pn-1. In in vitro studies, we evaluated the selectivity of Pn1 using CHO-CB1 and CHO-CB2 human receptors and [35S]-GTPγS binding assays. The preliminary results indicated that Pn1 (10 µM) appears to exhibit selectivity and affinity for the CB1 receptor. Further studies of the downstream signalling pathways for the CB1 receptor (calcium elevation and ERK1/2 phosphorylation), however, failed to show significant effects of Pn1 (10 µM). Antinociception induced by Pn1 appears to involve the inhibition of MAGL and anandamide endocannabinoid transporter and the selective activation of CB1 receptors. This novel peptide could be useful as a new antinociceptive drug candidate

Keywords: cannabinoid receptors,Peptide toxins, Antinociception

Research Title: COUPLING OF THE HUMAN RECOMBINANT CB2 CANNABINOID RECEPTOR TO AKT/PROTEIN KINASE B ACTIVATION/PHOSPHORYLATION IN CHINESE HAMSTER OVARY CELLS

Author: Wafa Moh'd Khair Hourani, Published Year: 2018

The 28th Annual International Cannabinoid Research Society Symposium on the Cannabinoids, Leiden university, the Netherlands

Faculty: Pharmacy

Abstract: Background: Cannabinoids affect the proliferation and differentiation of neural precursor cells in rats and mice. In oligodendrocytes, cannabinoid receptors influence stem cell survival and differentiation via the phosphatidylinositol 3-kinase:protein kinase B (PI3K-Akt, http://www.guidetopharmacology.org/GRAC/FamilyDisplayForward?familyId=285) signalling pathway(Gomez, Sanchez-Rodriguez et al. 2011). Recruitment to the plasma membrane and binding to phosphatidylinositol 3,4,5-trisphosphate allows phosphorylation and activation of Akt. The main objective of this investigation was to quantify the coupling of CB2 receptors to activation of the Akt signalling pathway. Methods: Chinese hamster ovary cells were stably transfected with human recombinant CB2 cannabinoid receptors. Activation of the Akt signalling pathway was assessed using a phosphopan-Akt antibody and in cell immunocytochemistry in 96-well microtitre plates quantified with a Licor Odyssey using fetal bovine serum as a positive control. Results: Multiple CB2 cannabinoid receptor agonists were investigated including the endogenous ligands; 2-AG and anandamide, as well as the synthetic ligands; CP55940, fenofibrate, HU210 ,WIN55,212-2, JWH015 and JWH133. A rank order of potency was CP55490 (pEC50 7.9 ± 0.04) > WIN55,212-2 (7.6 ± 0.1), HU210 (7.4 ± 0.1) > fenofibrate (7.1 ± 0.1), JWH015 (7.0 ± 0.1) > 2- AG (6.7 ± 0.2), JWH133 (6.3 ± 0.1) > anandamide (5.8 ± 0.1). The maximal effect induced by the different agonists was indicated as the Emax relative to the FBS response with 2-AG producing a maximum response at 149 ± 4 %, HU210 (142 ± 10), CP55940 (139 ± 12), fenofibrate (139 ± 11), WIN55,212-2 (116 ± 10), JWH015 (100 ± 10), JWH133 (98 ± 7), anandamide (71 ± 13). The phosphorylation/activation of the Akt signalling pathway was significantly attenuated by pretreatment with the CB2 selective antagonist AM630 (1 µM) or pertussis toxin (100 ng/mL). Conclusion This is the first study to quantify coupling of the CB2 receptor to Akt phosphorylation using cell immunocytochemistry. In common with other signalling pathways, 2-AG appears to act as a full agonist, while anandamide is a partial agonist, in the phosphorylation of Akt.

Keywords: Cannabinoids, cannabinoid receptors,anandamide, protein kinase B

Research Title: Coupling of the human recombinant CB2 cannabinoid receptor to extracellular signal-regulated kinase and calcium elevation in Chinese hamster ovary cells

Author: Wafa Moh'd Khair Hourani, Published Year: 2017

8th European Workshop on Cannabinoid Research, University of Roehampton, London, UK

Faculty: Pharmacy

Abstract: Background: CB1 cannabinoid receptors are characterised as coupling via the Gi/o family of G proteins to the inhibition of adenylyl cyclase activity and the activation of extracellular signal-regulated kinase (ERK)1. CB1 receptors in neurones also inhibit voltage-gated calcium channels, activate potassium channels and stimulate calcium mobilisation1. CB2 cannabinoid receptors are described to inhibit adenylyl cyclase and activate ERK, but not to regulate ion channel function1. In this study, we have investigated the coupling of human recombinant CB2 cannabinoid receptors expressed in Chinese hamster ovary cells. Methods: Human recombinant CB2 cannabinoid receptors were expressed in Chinese hamster ovary cells. ERK activation was assessed using in cell immunocytochemistry quantified with a Li-Cor Odyssey, while intracellular calcium levels were assessed using fluo-4 with a FlexStation with ATP as a positive control. Results:A range of cannabinoid agonists both endogenous (anandamide and 2-arachidonoylglycerol), natural (THC, β-caryophyllene and caryophyllene N-oxide) and synthetic (CP55940, WIN55,212-2, fenofibrate,HU210, JWH133, and JWH015) were tested. The following agents evoked maximal responses similar to the ATP (78-90 %) with the rank order: WIN55212-2 (pEC50 7.7 ± 0.1)> HU210 (7.4 ± 0.2)> fenofibrate (7.2 ± 0.1), JWH015 (7.2 ± 0.1)>2-AG (6.8 ± 0.1)>JWH133 (6.5 ± 0.1). AEA evoked a reduced maximal response (51%, 5.7 ± 0.2), while β-caryophyllene and caryophyllene N-oxide were inactive. 1 µM AM630, a CB2-selective antagonist, or pretreatment with 100 ng/mL pertussis toxin blocked responses to all these agonists. Calcium ion levels were stimulated by cannabinoid agonists, albeit in a slower manner than ATP. These responses were quantified as area under the curve: WIN55212-2 (21 ± 5 % ATP response, 7.3 ± 0.1) , HU210 (24 ± 3, 6.8 ± 0.3), CP55940 (37 ± 5, 6.7 ± 0.3), JWH015 (24 ± 3, 6.4 ± 0.1), fenofibrate (26 ± 3, 6.3 ± 0.2), 2-AG (34 ± 6, 6.2 ± 0.2), JWH133 (29 ± 7, 6.0 ± 0.1) , AEA (11 ± 1, 5.7 ± 0.3). Responses to THC failed to reach statistical significance, while those to β-caryophyllene and caryophyllene N-oxide were not distinguishable from background. Calcium responses to cannabinoid ligands were blocked by pretreatment with 1 µM AM630 or 100 ng/mL pertussis toxin. Cannabinoid ligand-evoked calcium responses were unaltered in the presence of 10 µM U0126, a selective inhibitor of ERK activation. Conclusions: This is the first study to determine coupling of the CB2 cannabinoid receptor to calcium elevation in recombinant expression.

Keywords: cannabinoid receptors,extracellular signal-regulated kinase,calcium

Research Title: A Voltammetric Sensor Based on Iodine-Coated Platinum Electrode for Determination of Iron in Blood Serum

Author: Wafa Moh'd Khair Hourani, Published Year: 2018

ANALYTICAL & BIOANALYTICAL ELECTROCHEMISTRY , Volume 10 , Number 1

Faculty: Pharmacy

Abstract: A novel, simple, convenient, inexpensive and green voltammetric sensing for iron in human blood serum was developed. The method is based on linear sweep voltammetry at an iodine-coated polycrystalline platinum electrode. A miniaturized home-made small scale 0. 5-mL cell was used for the analysis. Oxidation of iron at the iodine-coated platinum electrode was manifested by a peak centered at 0. 5 V. The established calibration curve for the relationship between iron (II) concentration and current extracted from the linear sweep voltammograms revealed an excellent linearity(R2=0. 9923). The calculated lowest detection limit, LOD, is 0. 26 ppm and the limit of quantification is 0. 87 ppm. Recovery studies for a concentration of 1. 0 ppm yielded a value of 1. 02 ppm (102% percent recovery). No interference with other common essential elements in blood serum was observed which attests to the suitability and accuracy of the method. The developed method was applied to analysis of real blood samples. The results of analysis were compared with the results from conventional standard methods used in medical laboratories where an excellent agreement between the two analytical methods was observed.

Keywords: Voltammetric sensors,Iodine-coated platinum electrode,Serium analysis,Iron determination in blood

Research Title: n−3 polyunsaturated N -acylethanolamines are CB 2 cannabinoid receptor-preferring endocannabinoids

Author: Wafa Moh'd Khair Hourani, Published Year: 2018

Molecular and Cell Biology of Lipids , Volume 1863, Issue 1

Faculty: Pharmacy

Abstract: Anandamide, the first identified endogenous cannabinoid and TRPV1 agonist, is one of a series of endogenous N-acylethanolamines, NAEs. We have generated novel assays to quantify the levels of multiple NAEs in biological tissues and their rates of hydrolysis through fatty acid amide hydrolase. This range of NAEs was also tested in rapid response assays of CB1, CB2 cannabinoid and TRPV1 receptors. The data indicate that PEA, SEA and OEA are not endocannabinoids or endovanilloids, and that the higher endogenous levels of these metabolites compared to polyunsaturated analogues are a correlate of their slow rates of hydrolysis. The n−6 NAEs (AEA, docosatetraenoyl and docosapentaenoyl derivatives) activated both CB1 and CB2 receptors, as well as TRPV1 channels, suggesting them to be ‘genuine’ endocannabinoids and ‘endovanilloids’. The n−3 NAEs (eicosapentaenoyl, docosapentaenoyl and docosahexaenoyl derivatives) activated CB2 receptors and some n−3 NAEs (docosapentaenoyl and docosahexaenoyl derivatives) also activated TRPV1 channels, but failed to activate the CB1 receptor. We hypothesise that the preferential activation of CB2 receptors by n−3 PUFA NAEs contributes, at least in some part, to their broad anti-inflammatory profile.

Keywords: AnandamideEndocannabinoidsCannabinoid receptorsVanilloid receptorsFatty acid amide hydrolasePolyunsaturated fatty acids

Research Title: Cannabinoid ligands, receptors and enzymes: Pharmacological tools and therapeutic potential

Author: Wafa Moh'd Khair Hourani, Published Year: 2018

Brain and Neuroscience Advances, Volume 2: 1–8

Faculty: Pharmacy

Abstract: Endocannabinoids have been identified to have roles in numerous physiological and pathological processes. Largely due to the association of the effects of Cannabis administration on mental states, the CNS impact of the endocannabinoid system has been the most intensively studied. Here, we provide a brief summary of the endocannabinoid system, comprising the receptors and the multiple endogenous lipid derivatives which activate them, as well as the enzymes which control the levels of these lipid derivatives. We identify pharmacological tools which may be used to interrogate the endocannabinoid system, as well as current and future options to exploit the system in the clinic.

Keywords: Cannabis, cannabinoids, the endocannabinoid system, anandamide, 2-arachidonoylglycerol

Research Title: A comparative study of Lane Change Assistance Protocols at Road Network

Author: Maram Bani Younes, Published Year: 2020

Faculty: Information Technology

Abstract: Driving over road networks is an everyday challenge that several research studies have been proposed to make it more safe and convenient. One of the main issues need to be efficiently and accurately handled is lane change scenarios. Drivers need to change their driving lane to take an exit on a road or to open the way for faster or emergency vehicle on that lane. Changing lane over highway road scenarios is a more serious challenge than changing it on downtown areas. This is due to the relatively high speed moving vehicles there. Vehicles in blind spots and wrong estimations of the distances between vehicles are the main two challenges that en-face drivers while changing their driving lanes. These cases may cause fatal accidents and lead to several damage consequences. This paper aims to investigate and compare the different previous proposed protocols that have been used to control safe lane changing for autonomous vehicles. Moreover, it reviews the intelligent driving assistance protocols that help drivers to safely change their lanes in several scenarios.

Keywords: Lang Change, Highway Road Scenario, Driving Assistance Protocol, Autonomous vehicle.

Research Title: The value of caring behavior and its impact on students' self‐efficacy: Perceptions of undergraduate nursing students

Author: Maha Mohammed Wahbi Atout, Published Year: 2020

Nursing Forum, 55

Faculty: Nursing

Abstract: Background Clinical instructors are the key mediators in helping students conquer clinical experiences by preparing them for clinical workplaces. The caring behavior of instructors plays an important role in the instructor‐student relationship.Objective: This study aimed to (a) assess the perception of nursing students of clinical instructors' caring behavior and (b) explore the correlation between their perception of instructors' caring behavior and their self‐efficacy.Design: A quantitative descriptive correlation design was adopted.Setting: The sample was collected from the faculty of nursing at a major governmental university for females in the Kingdom of Saudi Arabia.Participants: A sample of nursing students (N = 200) was recruited.Methods: The data were collected using the inventory for nursing students' perceptions of instructor caring (NSPIC) and general self‐efficacy (GSE) scale. Descriptive and inferential statistics, such as Pearson r correlation and regression analysis, were used.Results: The total mean of NSPIC was moderate to high (mean = 3.06, SD = 1.04). The level of GSE among students was high (mean = 3.45, SD = 0.849). NSPIC is significantly correlated with its GSE (r = .282). In addition, GSE turned out to be a major indicator of caring behavior, with P = .021 and R2 = 0.642.Conclusion: Caring relationships between clinical instructors and nursing students enable students to grow as caring professionals. Therefore, nursing programs must have highly qualified clinical instructors to teach and train students and be a good role model in the workplace.

Keywords: caring behavior, clinical instructor, nursing, self‐efficacy, students' perception

Research Title: Development and validation of RP-HPLC for related substance of pemetrexed disodium

Author: Balakumar Chandrasekarn, Published Year: 2012

International Journal of Pharmaceutical Sciences Review and Research , 15

Faculty: Pharmacy

Abstract: Simple and sensitive method for the quantification of related substance of pemetrexed disodium injection by RP-HPLC has been developed. Isocratic separation of pemetrexed disodium is carried out using a reversed-phase x-bridge column C18 (150 mm × 4.6 mm, 5 μm) with mobile phase consisting of acetonitrile and buffer (pH adjusted to 5 with orthophosphoric acid) in the ratio 15:85 (v/v) and quantified by UV detection at 230 nm. Analytical run time was 15 min. The assay exhibited good linear relationship with an accuracy and precision were over the concentration range of 0.1-10 μg/ml. This method can be used for routine analysis of related substances of pemetrexed disodium in injectables.

Keywords: HPLC, validation, related substances, pemetrexed disodium

Research Title: Pharmaceutical compounds

Author: Balakumar Chandrasekarn, Published Year: 2018

Faculty: Pharmacy

Abstract: A compound having the general formula (I) or (II), in which R, Rl, R2 and R3 are as described herein, or a pharmaceutically acceptable salt thereof, is provided. An associated method for treating a mycobacterial infection is also provided.

Keywords: Pharmaceutical compounds