HISTORIC LIST OF MAPLETHORPE FELLOWS FROM OCTOBER 1987
Derek Anthony John Balon (KCL) October 1987 - September 1990
“To develop a computer based method to assist diagnosis of minor conditions and to assess its use in undergraduate pharmacists.”
Melwyn Rex Euerby (Sch Pharm) October 1987- October1988
“War on Worms.”
Kenneth King Lam Ho (KCL) October 1987 - February 1989
“An evaluation of current techniques employed in the analysis of aerosols in Medicine.”
Annette Louis Leeman (Sch Pharm) October1987 - September 1989
“Towards optimising the administration of high dose methotrexate in cancer patients.”
Lam Siu Ping (KCL) October 1988 - September 1989
“Amine-imine tautomerism as a factor in the metabolism of amino azaheterocycles to toxic or non-toxic N-oxidation products.”
Marguerita Sui Lin Lim (Sch Pharm) October 1988 - September 1990
“The structural characterisation of the AIDS retroviral protease and of AIDS viral peptides for vaccine production.”
Amala Raman (KCL) October 1988 - August 1989
“The metabolic N-Hydroxylation of cyclic amides in plants.”
Felicity Julia Smith (Sch Pharm) October 1988 - September 1989
“Health care in the residential homes of the elderly.”
Subsequent Post: Professor of Pharmacy Practice, Department of Practice and Policy, SoP
Majed Sharif Abu Shamat (KCL) October 1989 - September 1992
“Metabolic and absorption studies involving the large intestine.”
Colin William Wright (Sch Pharm) October 1989 - September 1992
“Natural products and the development of novel antiamoebic and antimalarial agents.”
Geoffrey Harding (Sch Pharm) January 1990 - August 1991
“Developing courses in the Social and Behavioural Sciences in Pharmacy and research in the area of Pharmacy practice.”
Praful Udaykant Jani (Sch Pharm) October 1990 - March 1993
“Oral absorption of peptides and proteins in microspherical and vesicular carriers.”
Caron Wood (KCL) October 1990 - August 1992
“bsorption of collogen peptides and their role in oral tolerance and autoimmunity.”
Mark Helliwell (KCL) October 1990 - March 1993 + 3 months
“The lymphatic absorption of microencapsulated drugs.”
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Klara Valko (Sch Pharm) September 1991 - August 1993+ 3 months
“Parasite specific drug development based upon high technology metabolite screening of parasites and hosts.”
Alison Heather Batts Lansley (KCL) October 1991 - October 1993
“Development and use of a respiratory cell culture system to study drug transport and metabolism.”
Philip Charles Gordge (Sch Pharm) April 1992 - October 1993
“Isolation and characterisation of three phorbol ester kinase receptors from rat brain which are not known isozymes of protein Kinase C.”
Imogen Ann Temple Savage (KCL) April 1992 - June 1994
“Factors affecting the frequency and quality of counselling provided by pharmacists with prescribed medicines.”
Janine Lesley Bragger (KCL) October 1994 - September 1997
“Colonic oxidation-reduction potentials: Pharmaceutical and clinical applications.”
Reports: February 1996, September 1997, April 1998.
Publications: Bragger, J., Lloyd, A.W., Soozandehfar, S.H., Bloomfield, S.F.B., Marriott, C. and Martin, G.P. Investigations into the azo reducing activity of a colonic microorganism,
Int. J. Pharm. (1997) 157 61-71.
Bragger, J., Yazaki, E.T. and Evans, D.F. Colonic redox potentials in man, Reproducibility of measurements, Proc. 5th UKaps Annual Conference. Ed. I.W. Kellaway, G.Parr p.47
(1997).
Bragger, J., Yazaki, E.T. and Evans, D.F. Measurement of Colonic redox potentials in man. Pharm. Res. 14 S-656.
Soozandehfar, S.H., Lloyd, A.W., Bragger, J., and Martin, G.P. Azopolymer synthesis and degradation. Submitted to J. Contr. Rel.
Stirrup, V., Ledingham, S.J., Thomas, W.M., Pye, G., lftikhar, S.Y. , Bragger J. and Evans, D.F. Redox potential measurement in the gastrointestinal tract in man. In preparation.
Summary of Work: It has been proposed that the low redox potential prevailing in the colon may have a role in the degradation of azo- and disulphide-linked polymers designed for
colon-specific drug delivery. Due to the inherent difficulties in studying the colonic environment, there are limited data on redox potentials in man. The objectives of this project were
to determine inter- and intra-subject variability of colonic redox potentials in healthy subjects, using redox-sensitive radiotelemetry capsules and to compare pre- and post-study
calibrations. A Pt/Ag/AgCl radiotelemetry capsule, calibrated in saturated quinhydrone buffers pH 7 and 1.6, was swallowed and allowed to pass freely through the gastrointestinal
tract. Signals were recorded up to 48 h using a portable solid state receiver and recording system, allowing the volunteer to maintain normal ambulatory activities. Inter-subject
variability in colonic redox potentials was -491mV±43mV (n=8), and intra-subjectvariation -470mV±14mV (n=4). Post-study capsule calibration showed a maximum 'drift' in
recordings of +80 mV, which was reduced to +10 mV when the connection between body of the capsule and reference cap was sealed with silicone adhesive. These studies provide
further evidence of a uniquely low colonic redox potential in man and the post-study calibrations confirm that these low potentials are not an artefact of drift in capsule recordings
during transit through the gastrointestinal tract.
Subsequent employment: Employed by the Medicines Control Agency MCA (now the MHRA)
Anya Marie Hillery (Sch Pharm) October 1994 - April 1995 Relinquished
“The development and optimization of a novel particulate delivery system for the oral absorption of peptides.”
Reports: Nil
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Gary Stuart Tilbrook (KCL) October 1994 - September 1996
“Novel aluminium chelators for neurobiological application.”
Reports: July 1995, February 1996, April 1998
Summary of Work: Two new families of tribasic tetracoordinate chelating agents have been designed on the basis of the coordination requirements of aluminitim at physiological pH
(pH 7.4). Molecular modelling studies indicated the optimal linker dimensions for metal complexation. A multi-step synthetic route has been developed for the preparation of each
class of ligalid on a multi-gram scale. Two test compounds where selected for biological assessment using a rabbit model of aluminium-overload on the basis of physicochemical
analysis. Both ligands tested failed to induce aluminium excretion above control levels. In contrast, desferrioxamine, a ligand licensed for aluminium poisoning in man, was able to
induce a 50% increase in aluminium excretion. Further in-vivo experiments are therefore necessary to delineate the reasons for the observed ii-lactivity of these compounds. A
manuscript is in preparation detailing aspects of this work and will be submitted to "Inorganic Chemistry" for publication shortly.
Subsequent Post: Director of Chemistry, CeNeS (Responsibility for all chemistry activities relating to drug discovery and development for the company. Also, responsible for project
which is the lead optimisation for new treatments of Parkinson's disease)
Jacqueline Kay Miller (Sch Pharm) October 1995 - May 1996 Relinquished
“A study of the distribution of 5-HT and GABA receptor sites in healthy and diseased human whole brain sections using quantitative receptor autoradiography and in situ
hybridization.”
Reports: May 1996.
Susan Helene Kenyon (Sch Pharm) October 1996 - September 1998
“The involvement of vitamin B12 dependent methionine synthase and its dependent pathways in cancer formation and treatment.”
Reports: March 1997, September 1997, September 1998
Publications: Kenyon, S.H., Nicolaou, A. and Gibbons, W.A. (1998) 'The effect of ethanol and its metabolites upon methionine synthase in vitro.'Alcohol 15 305-309.
Kerai, M.D.J., Waterfield, C.J., Kenyon, S.H., Asker, D.S. and Timbrell, J.A. (1998) 'Taurine: Protective properties against ethanol-induced hepatic steatosis and lipid peroxidation
during chronic ethanol consumption in rats.' Amino Acids 15 53-76
Kenyon, S.H., Waterfield, C.J., Asker, D.S., Kudo, M., Moss, D.W., Bates, T.E., Nicolaou, A., Gibbons, W.A. and Timbrell, J.A. (1998) 'The of hydrazine upon vitamin B,2dependent
methionine synthase activity and the sulphur amino acid pathway in isolated rat hepatocytes.'Biochem. Pharmacol. submitted.
Kenyon, S.H., Nicolaou, A. and Gibbons, W.A. (1998) 'Acetaldehyde, but not alcohol or acetate, inhibits vitamin B12-dependent methionine synthase in vitro.' Ned. J. Med.52 s9.
Kenyon, S.H., Waterfield, C.J., Asker, D.S., Nicolaou, A., Timbrell, J.A. and
Gibbons, W.A. (1998) 'The effect of hydrazine on vitamin B12-dependent methionine synthase activity in isolated rat hepatocytes.' Ned J Med. 52 s 10.
Nicolaou, A., Kenyon, S.H., Waterfield, C.J., Gibbons, W.A. (1998) 'Control and regulation of methionine synthase.' Ned. J. Med. 52 sl2
Kerai, M.D.J., Waterfield, C.J., Kenyon, S.H. Asker, D., and Timbrell, J. (1998) 'Taurine: protective properties against ethanol-induced hepatic steatosis and lipid peroxidation during
ethanol consumption in rats'Human & Expt. Tox. in press
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Summary of Work: The enzyme methionine synthase (Bl2-MS) produces methionine from homocysteine and requires vitamin B12 and folate for activity. Some tumour cells have
been identified as methionine dependent; the aim of this project was to investigate whether B12-MS activity was involved in tumour formation. Inhibition of methionine synthase by
cancer causing compounds was also investigated. Alcohol, a 'known' inhibitor of B12-MS was not found to inhibit enzyme activity in vitro, but acetaldehyde (a metabolite of alcohol)
did. In vivo methionine synthase was inhibited in the liver, but not the brain, of rats given daily alcohol for one month. Taurine a sulphur amino acid and antioxidant, given
concurrently with the alcohol, did not prevent inhibition of methionine synthase activity. Hydrazine, a laboratory cancer inducer, was also found, in isolated rat hepatocytes, to inhibit
methionine synthase activity, increase homocysteine excretion and increase total taurine levels. The rat hepatoma cell line, Phi 1, was able to upregulate methionine synthase activity
by 10 fold and could be described as methionine independent. Another rat hepatoma cell line, HTC, was unable to increase enzyme activity more than 5 fold and the maximal
methionine synthase activity in these cells was only a fifth that of Phi 1 cells, HTC cells could be considered to be methionine dependent. Ability to increase methionine synthase
activity was postulated to be partly responsible for methionine dependency.
Belinda Janine Krishek (Sch Pharm) October 1996 – April 1999
“Modulation of GABAA receptors by H+: physiological and pharmacological implications.”
Reports: February 1997, November 1997, March 1998, November 1998, June 2000.
Publications: Krishek, Moss & Smart: Interaction of H+ and Zn2+ on recombinant & native rat neuronal GABAA receptors (Journal of Physiology1998, 507,3, pp. 639-652)
Summary of Work: Utilising both Xenopus oocytes and hypothalamic neurones, the pH dependence of zinc-induced inhibition of GABA-induced responses was determined. The
results demonstrate that the inhibition of the GABA response by zinc was modulated by changing the external pH around mammalian GABAA receptors. Interestingly, for the first
time, the inclusion of the δ subunit within the GABAA receptor complex was shown to influence the zinc-induced antagonism. The completion of this study culminated in a paper
entitled "interaction of H+ and Zn2+ on recombinant and native rat neuronal GABAA receptors" and was published in The Journal of Physiology.
Using rat cultured cerebellar granule cell neurones the modulation of GABAA receptors by H+ was studied over a three week period, when the composition of in situ GABAA receptors
is postulated to change. The results show a clear differential modulation of GABA responses by H+ that is dependent on the stage of neuronal development, and presumably subunit
composition. This modulation is most clearly marked between day 11 and 12 in vitro, with low pH inhibiting GABA responses before day 11 and enhancing GABA responses after
day 12. The completion of this study has culminated in a paper entitled "Proton sensitivity of rat cerebellar granule cell GABAA receptors: Dependence on neuronal development"
and has been submitted to The Journal of Physiology.
Fiona Cathleen Mortimer (née Sheppard) (KCL) October 1996 - September 1998
Topic: “An evaluation of fluorescent probes and their application to antimicrobial susceptibility testing”
Reports: February 1997, October 1997, March 1999.
Summary of Work: The traditional methods for determining the antimicrobial susceptibility of infecting organisms require at least an 18-24 hour delay before obtaining results. Thus
a more rapid method is desired. The aim of this study was to evaluate a variety of fluorescent probes to establish which could be utilised in detecting changes to the functionality of
bacterial cells as a result of antimicrobial action. Three membrane integrity probes, Propidium Iodide (PI), TO-PRO-1 and SYTOX Green, two membrane potential probes, bis-(1,3dibutylbarbituric acid)trimethine oxonol [IDiBAC4(3)] and 5,5',6,6'-tetrachloro-1,1',3,3'-tetraethylbenzimidazolyl-carbocyanine iodide [JC-1] and two metabolic dyes, MITOTRACKER
ORANGE and MITOTRACKER RED, were assessed. Cells were analysed flow cytometrically and by fluorescence microscopy.
All the membrane integrity probes were able to discriminate between log phase and permeabilised cells for both Gram positive (Staphylococcus aureus) and Gram negative
(Escherichia coli) populations. DiBAC4(3) could distinguish between cells with a high and low membrane potential in both population types but JC-1 only worked with the Staph.
aureus cells. Similarly the metabolic dyes were only useful in the Gram Positive population.
The membrane integrity and membrane potential dyes detected changes in cell populations exposed to various antimicrobial agents. Generally the cell-associated fluorescence
however, did not reflect the loss in viability of the population determined by plate counts. It was noted that the mechanism of action of the antimicrobial frequently influenced the
quantity of cell-associated fluorescence. Large fluorescence changes with all dyes were observed with the B-lactam antibiotics but smaller changes, or no change, occurred when
cells were exposed to antimicrobials acting against the nucleic acids.
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The study highlighted a number of problems associated with using fluorescent probes to monitor antimicrobial activity. Variation in dye response between bacterial species, and the
differing responses by the dyes to the antimicrobials, could impede the further development of a rapid antimicrobial susceptibility tests using these fluorescent probes.
Publications: Mortimer, F C, Mason, D J & Grant, V A (1998). The use of cell impermeable fluorescent probes to assess antibiotic-induced injury. Cytometry Suppl. 9 127.
Subsequent employment: Employed by the Medicines Control Agency MCA (now the MHRA)
Christopher James Richardson (KCL) January 1997 - December 1999
“An expert system for sequence-based protein 3D structure prediction.”
Reports: March 1997, November 1997, November 1998
Employment: Bioinformatics Computing Officer, Centre for Structural Biology, Institute of Cancer Research, Chester Beatty Laboratories, Fulham Road, South Kensington, London.
Summary of Work: A portfolio of predictive techniques have been developed to aid in sequence-based protein 3-D structure prediction, combining database searches of known
protein structures, to identify likely main chain conformations for short (5 residue) sequence motifs; a battery of artificial neural networks and a Voronoi tessellation-based procedure
for predicting protein structural class, a simple statistical procedure for predicting a protein’s set of buried residues; and a genetic algorithm for predicting protein secondary structure.
The utility of these various protein structure prediction methods was established by comparison with equivalent methods developed by other workers, and also through their combined
application in the sequence-based prediction of the 3-dimensional structure of the human duodenal ferric reductase implicated in the intestinal absorption of iron.
Xian Ming Zeng (KCL) October 1997 – January 1999
“Engineering of Dry Powder for Inhalation Aerosols.”
Reports: March 1998, November 1998, April 1999.
Summary of Work: To investigate the effects of molecular weight (MW) of polyvinylpyrrolidone (PVP) on glass transition and crystallization of sucrose. Methods: Sucrose was colyophilized with 2.5% and 5.0% w/w PVP of different molecular weights, which were characterized using a gel permeation chromatography. Freeze drying was carried out for 48
hours at a shelf temperature of –40oC and a pressure of about 270 mTorr. The samples were then dried in a vacuum oven at 24oC for 12 hours before drying for a further 12 hours
at 40oC. Differential scanning calorimetry (DSC) was employed to measure the glass transition temperature (Tg), dynamic crystallization temperature (Tc) and isothermal
crystallization induction time (tc) at 85oC of sucrose. Isothermal water vapour sorption of each sample was also measured at different relative humidities. Results: Tg values of
sucrose varied from 48.3 ± 0.8oC for freeze dried (FD) sucrose alone to 58.8 ±0.8oC for the mixture containing 5.0% PVP of a nominal MW 300K. PVP increased sucrose Tg
signficantly (ANOVA p<0.05). Although there was no significant difference (p>0.05) in Tgof the mixtures containing 2.5% w/w PVP of different MW, samples with 5.0% PVP of MW
300K produced a significantly higher (p<0.05) Tg than the other mixtures. All mixtures were shown to possess higher (p<0.01) Tg than sucrose alone, which exhibited a Tc of
approximately 85oC. PVP of MW 300K consistently induced a significantly (p<0.05) higher Tc of sucrose than PVP of smaller MW. Increasing PVP concentration from 2.5% to 5.0%
also resulted in a substantial increase in sucrose Tc. Sucrose tc was increased over 10 times when it was co-lyophilized with 2.5% PVP, different MW of which also led to differing
values of sucrose Tc For example, PVP of MW 300K resulted in a sucrose Tc at 85oC (89.1-95.6 min), which was approximately 7 times higher than that of 2.5% PVP of MW 24K or
40K. A longer Tc of sucrose was also observed for mixtures containing PVP of MW 300K than those with PVP of smaller MW when measured with isothermal water vapour
sporption. Conclusions: The effect of PVP on sucrose Tg, Tc and it is dependent upon MW. PVP of higher MW may be more efficient in inhibiting sucrose crystallization and by
stabilizing glassy structures of the sugar, these polymers may improve the stability of co-lyophilized proteins and peptides.
Publications: Xian Ming Zeng, Gary Peter Martin, Seah-Kee Tee and Christopher Marriott: The role of fine particle lactose on the dispersion and deaggregation of salbutamol
sulphate in an air stream in vitro. International Journal of Pharmaceutics. 176(1998) 99-110.
Xian Ming Zeng, Gary Peter Martin, Seah-Kee Tee, Abeer Abu Ghoush and Christopher Marriott: Effects of particle size and adding sequence of fine lactose on the deposition of
salbutamol sulphate from a dry powder formulation. International Journal of Pharmaceutics (in press).
Xian Ming Zeng, Gary Peter Martin and Christopher Marriott: Effects of molecular weight of polyvinylpyrrolidone on the glass transition and crystallization of co-lyophilized sucrose.
(Submitted for publication)
Xian Ming Zeng, Kiranpal H Pandhal and Gary P Martin: The role of fine particle lactose on the content homogeneity and dispersibility of beclomethasone dipropionate from powder
aerosols. (Submitted for publication)
Xian Ming Zeng, Gary Peter Martin, Christopher Marriott and John Pritchard: The influence of crystallization conditions on the morphology of lactose intended for use as a carrier for
dry powder aerosols. (Submitted for publication).
Xian Ming Zeng, Gary Peter Martin and Christopher Marriott: The influence of the carrier morphology on the drug delivery by dry powder inhalers (Submitted for publication)
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Xian Ming Zeng, Gary Peter Martin and Christopher Marriott: The effects of carrier size and morphology on the dispersion of salbutamol sulphate after aerosolisation at different flow
rates. (Submitted for publication)
Xian Ming Zeng, Gary Peter Martin and Christopher Marriott: Crystallisation of lactose from carbopol gels. (Submitted for publication)
Xian Ming Zeng, Gary Peter Martin and Christopher Marriott: Influence of particle engineering of the carrier on drug delivery by dry powder aerosols (Submitted for publication)
Xian Ming Zeng, Gary Peter Martin and Christopher Marriott: Lactose as a carrier in dry powder formulations: The influence of particle characteristics on drug delivery (Submitted for
publication)
Hassan Larhrib, Xian Ming Zeng, Gary Peter Martin, Christopher Marriott and John Pritchard: Using different characterised batches of lactose as the carrier for salbutamol sulphate
formulation for inhalation (Submitted for publication).
Xian Ming Zeng, Gary Peter Martin and Christopher Marriott: Particulate interactions in dry powder formulations (Accepted for publication by Taylor and Francis).
Subsequent employment: Initially held Chair at Medway School at Kent but now major industrialist.
Christopher Frederick van der Walle (KCL) October 1997 - February 1999
Topic: “Cloning, expression and structure determination of the major extracellular domain of the PepT1 oligopeptide transporter.”
Reports: February 1998, November 1998 & February 1999.
Summary of Work: While the project has not been given the chance to run its full course we have been successful in synthesising a relatively large gene, which was subsequently
expressed in E. coli. The over expression of a mammalian protein in E. coli is suggested as being a reason for the poor folding of the protein in the cell and its subsequent
accumulation as inclusion bodies. Refolding of the protein purified from inclusion bodies does not look optimistic, however, because of the large loss of protein during the refolding
step.
Expression of the MBP or GST fusion proteins in E. coli would provide good yields of soluble protein, easily purified in one step using affinity chromatography. This route does,
however, require the purchase of enzyme to cleave the fusion product. Alternative eukaryotic expression systems, including yeast and insect expression systems could also be
tested, facilities permitting.
Moreover, the cloned human, rat or rabbit PepTl and PepT2 genes have been obtained as a kind gift from Prof. Leibach (Fei et al., 1994) and it would now be possible to design
several protein constructs using standard PCR techniques encompassing the PepTl extracellular loop domain alone, or as a larger construct with flanking protein sequences.
Publications: C F van der Walle, C Prodromou & D J Barlow, Cloning, expression and structure determination of the major extracellular domain of PepT1 oligopeptide transporter, J.
Pharmacy & Pharmacol., 1998, 50, S234.
Subsequent Post: Academic post at the University of Strathclyde
Cheryl Joy Hemingway (Sch Pharm) November 1997 – June 2000
“Regulation of acetyl-CoA carboxylase in skeletal muscle by insulin and by the AMP-PK cascade - a potential target for gemfibrozil with implications for glucose intolerance in
diabetes.”
Reports: February 1998, November 1998, November 1999, June 2000
Summary of Work: Gemfibrozil is currently used in the treatment of hyperlipidaemia and inhibits the hepatic output of very low-density lipoproteins (VLDL) and increases lipoprotein
clearance. This is achieved through increased oxidation, decreased endogenous synthesis of the lipid components of VLDL and inhibition of esterification of fatty acids. The
inhibition of cholesterol and triglyceride synthesis is achieved via the activation of AMP-activated protein kinase (AMP-PK) possibly via phosphorylation) which in turn phosphorylates
and inactivates acetyl-CoA carboxylase (ACC) and 3-hydroxy 3-methylglutaryl-CoA reductase (HMG-CoAR), the enzymes controlling the first committed steps in fatty acid and
cholesterol synthesis, respectively. Gemfibrozil inhibits the microsomal isoform of the rate-limiting enzyme for esterification glycerol-phosphate acyltransferase (GPAT) possibly via
phosphorylation, with no effect on the mitochondrial isoform suggesting different functions isoform regulation. Antagonism of insulin action resulting in increased plasma glucose and
hepatic glycogen storage may have serious implications for the gemfibrozil-treatment of hyperlipidaemic patients with NIDDM. Purified brain AMP-PK has similar characteristics to
liver AMP-PK and it may be found that brain AMP-PK, like liver has an important role in cellular stress response. Inhibitory peptides developed to AMP-PK are effective in vitro and
delivery of these peptides into cells will serve as a useful laboratory tool in opening a new era in determining the function of AMP-PK.
Publications:
Hemingway C J and Munday M R 'Gemfibrozil activation of AMP-activated protein kinase'. Biochem.Soc.Trans. (1997) 25, 148.
Hemingway, C J & Munday M R: The lipid-lowering agent gemfibrozil inhibits fatty acid esterification. J.Pharmacy and Pharmacology (1998) 50 (supplement) 208.
Munday M R and Hemingway C J: The effect of the lipid-lowering agent gemfibrozil on glucose handling. J.Pharmacy and Pharmacology (1998) 50 (supplement) 209.
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Munday M R and Hemingway C J: The regulation of Acetyl-CoA carboxylase – a potential target for the action of hypolipidaemic agents. Advances in Enzyme Regulation (1998) 39,
205-234.
Hemingway C J, Tey K K &Munday M R: Short-term inhibition of fatty acid and cholesterol biosynthesis by the hypolipidaemic drug gemfibrozil mediated by stimulation of AMPactivated protein kinase. Biochem.J. (1999 (in press)
Hemingway C J and Munday M R: The fibrate, gemfibrozil inhibits fatty acid esterification. Biochim. Biophys. Acta (1999) submitted.
Hemingway C J and Munday M R: The effects of fibrates on carbohydrate metabolism in the rat. Biochim. Biophys. Acta (1999) submitted.
Thiagarajan Sakthivel (Sch Pharm) October 1998 – September 2000
“Bacterial protein mediated dendrimer carrier system to promote cellular entry.”
Reports: March 1999, November 1999
Publications: Florence A T, Sakthivel T, Toth I: Oral uptake and transport of a lipidic dendrimer. J.Control.Rel. (1999 in press).
D Shah, T Sakthivel, I Toth, A T Florence and A F Wilderspin: Investigation of DNA transfection and cell viability using high concentration of lipidic dendrimeric lysine. J.Pharm.
Pharmcol. 1999, 51 (supplement) 138.
Reem Kayyali (KCL) 1 October 1998 – 30 December 2000
“Investigation of the mechanism of agranulocytosis induction.”
Reports: March 1999, November 1999, February 2000, December 2000.
Publications: Kayyali R S, Porte J B, Lui Z D, Nugent J and Hider R C: Structure function investigation of the interaction of 1- and 2 – substituted 3-hydroxpyridin-4-one with 5lipozygenase and ribonucleotide reductase. J.Biol.Chem. (In press 1999)
Kayyali R S, Pizzy A, Porter J B and Hider R C: Apoptosis induction by iron chelators in thymocytes and proliferating leukaemia cells. J.Biochem.Pharmacol. (In preparation 1999)
Hider R C, Kayyali R and Evans P: The production of hydroxyl radicals by deferiprone-iron compounds under physiological conditions. Blood (Abstract) In press 1999.
Kayyali R S, Waterfield C J, Asker D, Evans P, Timbrell J A and Hider R C: Investigation of the mechanism of agranulocytosis induction by 3-hydroxypyridin-4-one iron chelators.
J.Biol. Pharmcol. (In preparation 1999)
Kayyali PS, Porter JB, Lui ZD, Davies NA, Nugent J, Cooper CE and Hider PC (2001): Structure function investigation of the interaction of 1 - and 2- substituted 3-hydroxypyridin-4one with 5-lipoxygenase and ribonucleotide reductase. J. BioA Chem., Jnprparation.
Kayyali RS, Lynagh GR, Fizzy A, Porter JB and Hider PC (2001): Apoptosis induction by iron chelators in thymocytes and proliferating leukaemia cells. J Biochem. I)hcirrnacol.,
Jnprepara(ion.
Lui ZD, Kayyali PS, Theobald AE and Hider PC (2001): The design and biological evaluation of novel 2-substituted 3-hydroxypyridin-4-one iron chelators. Pharm. Res., Jn
preparation.
Summary of Work: The orally active bidentate hydroxypyridinone (HPO) chelator currently in clinical trials, deferiprone, was found to induce bone marrow hypoplasia in laboratory
animals and agranulocytosis in a small proportion of humans. The mechanism of agranulocytosis induction by this class of chelators is not fully understood, In order to contribute to
the understanding of the mechanism of bone marrow toxicity induced by these chelators, we studied metabolic activation and hydroxyl radical generation as possible triggers for the
event, In this study, the amount of hydroxyl radical produced by two HPO chelalors, deferiprone and CP502 has been compared under physiologically relevant conditions using the
deoxyribose assay. These two chelators have different affinities for iron at pH 7.4, deferiprone (pM=19.5) and CP502 (pM=21.7). Metabolic activation as a mechanism of toxicity was
investigated by monitoring the depletion of intracellular glutathione (GSH) with varying concentrations of deferiprone and CP502 and their iron complexes in both neutrophils; a
primary site of toxicity by HPO chelators and hepatocytes; an important site of iron storage. The viability of neutrophils was monitored using the trypan blue exclusion dye and that of
hepatocytes by using lactate dehydrogenase leakage. Furthermore, lipid peroxidation in these cells was measured as an estimate of 'OH production. Hydroxyl radicals were
generated in the presence of both the 2:1 and 3:1 deferiprone and CP502 complexes with iron. However, the amount of hydroxyl radical generated in the presence of the 3:1
deferiprone and CP502 complexes was significantly lower than that produced in the presence of the equivalent 2:1 complexes, with the amount of hydroxyl radical generated in the
presence of the 2:1 and 3:1 CP502 complexes with iron being significantly lower than that generated by the equivalent deferiprone complexes. Jn the simultaneous presence of
physiological levels of citrate and GSH, the amount of hydroxyl radical generated in the presence of the 2:1 and 3:1 deferiprone:Fe and CP502:Fe complexes became negligible.
When the direct toxicity of the 2:1 deferiprone:Fe complex on neutrophils was investigated using GSH depletion as an indicator, we found that the 2:1 deferiprone:Fe caused
significant depletion of GSH in neutrophils. A direct correlation between the concentration of the 2:1 deferiprone:Fe complex and GSH levels was observed; GSH levels decrease with
increasing 2:1 complex concentration. A less significant GSH depletion was observed with the 2:1 CP502 complex. When similar experiments were performed using hepatocytes, a
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less marked GSH depletion was observed. The difference possibly resulting from the higher availability of H202 in neutrophils. It must be pointed out that GSH depletion occurred in
these cells without affecting their viability and without any evidence of lipid peroxidation occurring. The results of this study may offer an explanation for the cause of agranulocytosis
that is observed in some patients receiving treatment with deferiprone. In principle, this toxicity could be overcome by the use of an FIPO chelator with a higher affinity for iron, such
as CP502, as its OH radical generation potential is outbalanced by its higher affinity for iron.
Atiya Rao (now Shah) (KCL) April 1999 – July 2000
“Probing the cellular mechanisms of drug absorption from the lungs.”
Reports: November 1999, July 2000.
Publications: Atiya Shah, Benjamin Forbes, Gary Peter Martin and Alison Lansley: Characterisation of the 16HBE14o- cell culture model of the airways. Pharm. Res. (Submitted).
Atiya Shah, Gary Peter Martin and Alison Lansley: Transport of a series of model peptides across the 16HBE14o- cell model of the airways and alveolar type II cells of the deep lung.
Pharm. Res. (Submitted)
Summary of Work: Three peptides attached with a fluorescent label (anthranilic acid) were synthesised for the purpose of analysing their transport rates and pathways across a
human bronchial epithelial cell culture model of the airways (16HBE14o- cell line). The peptides and the label ranged approximately 10-rold in molecular weights from 143 to 1142.
All peptides and label were analysed using HPLC and matrix-assisted laser desorption ionisation (MALDI) instruments, indicating that the peptides were 99% pure, relatively
hydrophilic and of the correct mass. 16HBE14o- cells were cultured on monofibrillar Vitrogen-coated polycarbonate filters at a seeding density of 2.5 x 105 cells/cm2.. The cells were
cultured for a period of 6 days, when the transepithelial electrical resistance (TER) across the cells was measured. TER across the 16HBE14o- cells were 278 ± 58 Ω cm2 (n=30).
Initially, radiolabelled mannitol (an inert paracellular marker, MW 182) was transported across the cells. By applying the apparent permeability equation (Papp=dQ/dt x 1/A.C0, where
dQ/dt is permeability rate of mannitol, A= area of filter and Co is the initial concentration of mannitol) the apparent permeability rate of mannitol (mW 182) was calculated to be 1.96 ±
0.34 x 10-6 (n=12). The transport of anthranilic acid and the peptides was also investigated at a range of concentrations (1 mg/mL, 0.5 mg/mL, 0.25 mg/mL, 0.1 mg/mL, 0.01mg/mL,
0.001 mg/mL and 0.0001 mg/mL) across the 16HBE14o- cell line in the apical-basolateral direction. Papp values were 22.9 ± 0.45 x 10-7 (n=18), 9.62 ± 0.74 x 10-7 (n=27), 6.79 ±
0.39 x 10-7 (n=30) cm s-1 for anthranilic acid, S-Q-S-D-AA, S-Q-S-S-Q-S-D-AA and S-Q-S-S-Q-S-S-Q-S-D-AA, respectively. The transport rates of the label and each of the peptides
were similar, irrespective of the initial concentration, indicating passive transport. The various molecular weights of the peptides indicate that they were transported via the
paracellular route and the only constraints on transport were the size of the tight junctions between cells. The lower the molecular weight of the peptide, the greater the transport rate.
HLPC and MALDI analysis on peptides both pre- and post-transport indicated the peptides were still intact post-transport, with the label attached and the masses (from MALDI data)
were equivalent to the peptides pre-transport.
Subsequent employment: Promotional Affairs Adviser, AstraZeneca.
Ana Miljkovic-Brake (KCL) 18 October 1999 – 31 January 2001
"Exploration of novel bioactive secondary metabolites in previously untapped filamentous fungi employing a newly developed method based on the use of novel radiolabelled
precursors."
Reports: March 2000, November 2000
Publications:
Does Apoptosis cause renal atrophy in Balkan endemic nephropathy? Lancet, 352: 1118-119 (Mantle, P.G., Miljkovic, A., Udupa, V., Dobrota, M. (1998)
Debrisoquine 4-hydroxylation and the Balkan Endemic nephropathy, Nephron, 82 (3): 347-348 (Mantle, P G, Amirtharajah, M, Slippel, S, Miljkovic, A, Naik, J, Nesler, S. (1999)
Influence of halogen salts on the production of the ochratoxins by Aspergillus ochraceus, Journal of Agricultural and Food Chemistry, 48, 5, 1865-1871 (Stander, M A, Steyn, P S,
Foodtek, A L, Miljkovic, A, Mantle, P G, Foodtek, G M (2000)
Mantle, P.G., Miljkovic, A., Udupa, V., Dobrota, M. (1998): Does Apoptosis Cause Renal Atrophy in Balkan Endemic Nephropathy? Lancet, 352: 1118-1119.
Mantle, P. G., Amirtharajah, M., Klippel, S., Miljkovic, A., Naik, J., Nesler, S. (1999): Debrisoquine 4-hydroxylation and the Balkan Endemic Nephropathy, Nephron, 82 (3): 347-348.
Stander, M. A., Steyn, P. S., Foodtek, A. L., Miljkovic, A., Mantle, P. G., Foodtek, G. M. (2000): Influence of halogen salts on the production of the ochratoxins by Aspergillus
ochraceus, Journal of Agricultural and Food Chemistry, 48, 5, 1865-1871.
Mantle, P. G., Miljkovic, A., Dobrota, M. (2000): Dose and route responses of nephrotoxic mycotoxins, in “Mycotoxins and Phycotoxins”, refereed proceedings of the X International
IUPAC Symposium on Mycotoxins and Phycotoxins, (in press).
Miljkovic, A., Mantle, P. G., Williams, D., Rassing, B. (2001): A new natural azaanthraquinone produced by a Bispora-like tropical fungus, Journal of Natural Products, 64: 1251-1253.
8
Miljkovic, A., Pfohl-Leszkowicz, A., Dobrota, M., Mantle, P., (in press): Comparative responses to mode of oral administration and dose of ochratoxin A or nephrotoxic extract of
Penicillium polonicum in rats, Experimental and Toxicologic Pathology.
Summary of Work:
The continuous need for drug discovery has induced novel approaches to the study of biologically active compounds, one of which is based on the diversity and chemical creativity of
microorganisms. Of the conservatively-estimated 1.5 million fungal species, only approximately 5% have been described and characterised, while only 0.3% have been screened for
biological activity.1 However, fungal secondary metabolites represent a large proportion of the known biologically-active compounds. Exploration of ecologically “unusual” habitats,
which was particularly intensified in the last couple of decades, has led to the discovery of a considerable number of novel and more diverse microbial species. These untapped
microbial “factories” possess immense potential for discovery of novel compounds and biotransformations as a source of valuable drugs. Despite the wide structural diversity of
fungal secondary metabolites, they are biosynthesised from only a few key metabolic precursors and intermediates such as acetate, mevalonate, shikimate and amino acids.2
Administration of such simple “building blocks” to a fungus and following their incorporation into the end-products can allow for predictions regarding the structure of the secondary
metabolites and their biosynthetic origin, which consequently may indicate likely biological activity. A novel miniaturized method exploiting radiolabelled biosynthetic probes has been
developed.3
Subsequent Appointment: Lecturer in the Department of Pharmacy at KCL.
Weng Li Yoon (Sch Pharm) 18 February 2000 – 17 March 2002
"The rapid identification of counterfeit pharmaceutical products using near infrared spectroscopy."
Reports: November 2000, November 2001,
Publications:
The Optimisation of sample presentation in the analysis of pharmaceutical excipients using near infrared spectroscopy, Journal of Pharmacy and Pharmacology, Vol 49, Supplement
(W L Yoon, R D Jee and A C Moffat) (1998)
The optimisation of sample presentation for the near-infrared spectra of pharmaceutical excipients, The Analyst, 1029-1034 (W L Yoon, R D Jee and A C Moffat) (1998)
The Transfer of near infrared spectra of solvents between different instruments, Journal of Pharmacy and Pharmacology, Vol 50, Supplement (W L Yoon, R D Jee, A C Moffat, D C
Lee, P D Blackler and K Yeung) (1998)
Water sorption and near infrared spectroscopy to study the differences between microcrystalline cellulose before and after wet granulation, International Journal of Pharmaceutics,
181, 41-47 (G Buckton, E Yonemochi, W L Yoon and A C Moffat) (1999)
Construction and transferability of a spectral library for the identification of common solvents by near infrared transflectance spectroscopy. The Analyst 124, 1197-1203 (W L Yoon, R
D Jee, A C Moffat, P D Blackler, Ken Yeung and D C Lee) (1999)
An interlaboratory trial to study the transferability of a spectral library for the identification of solvents using near infrared spectroscopy, Journal of Pharmacy and Pharmacology, Vol
51, Supplement (W L Yoon, R D Jee and A C Moffat) (1999)
W.L Yoon, G. Lee, A. Charvill, R. D. Jee and A. C. Moffat, The Use of Near-infrared spectroscopy to determine the authenticity of parallel imports, British Pharmaceutical Conference
2001, Abstract Book:12, Pharmaceutical Press, London, ISBN 0 85369 5121.
Weng Li Yoon, Roger D. Jee, Gerard Lee, Andrew Charvill, Anthony C. Moffat, A Non-destructive Method To Detect Counterfeit Medicines Using Near-infrared Spectroscopy, 2001,
Vol.3, ISSN 1522-0893, M2396
Weng Li Yoon, Roger D. Jee, Gerard Lee, Andrew Charvill, Anthony C. Moffat, Application of near-infrared spectroscopy to the determination of the sites of manufacture for
proprietary products, Analyst (pending submission)*.
Weng Li Yoon, Roger D. Jee, Gerard Lee, Andrew Charvill, Anthony C. Moffat, The use of near-infrared spectroscopy to the detect counterfeit drug products, Analyst (pending
submission)*.
* Publication awaiting clearance within the Medicines Control Agency.
Summary of Work: Given that the definition of a counterfeit product is a deliberate mislabelling of the content and/or origin of manufacture, this project attempts to use this technique
to: (1) determine the authenticity of using real-life suspect tablets (2) trace the origin of tablet (manufacturing site) using parallel-imported tablets. Different batches of four proprietary
tablets originating from up to 4 different sites of manufacture were scanned. For one of the products which was unnamed, 4 different real-life suspects were available. Counterfeit
tablets gave evident differences in spectral patterns which were detectable by eye. However, genuine tablets originating from different sites, which are manufactured using the same
formulation, gave spectra which were identical. However, upon the application of multivariate method of analysis called principal component analysis, tablets of different origin formed
separate clusters. Further analysis using algorithms called SIMCA (Soft Independent Modelling Class Analogy) revealed that each of these clusters was a statistically separate
9
population (P < 0.05). This library was incorporated into the Vision software of the FOSS NIRSystems instrument. All known counterfeits were identified as FAIL and a classification
rate of 99.7% were achieved when identifying tablets according to their sites of manufacture. Recent findings also revealed that this technique offers the potential to help us
understand that differences between the tablet and even the potential to identify and quantify the aspects of the formulation which have been altered.
Emma Louise Dunne (Sch Pharm) 1 July 2000 – 31 July 2001
"Molecular definition of a modulatory site on the GABAa receptor complex."
Reports: July 2001.
Publications:
MOLECULAR IDENTIFICATION OF RESIDUES AND INTERSUBUNIT INTERACTIONS COORDINATING ZINC INHIBITION ON GABAA RECEPTORS
Dunne EL, Hosie AM, Harvey RJ, Smart TG
Manuscript submitted to Neuron
N-TERMINAL HISTIDINE REGULATES Zn2+ INHIBITION ON THE MURINE GABAA RECEPTOR B3 SUBUNIT
Dunne EL, Hosie AM, Wooltorton JRA, Duguid IC, Moss SJ, Harvey RJ, Smart TG
Manuscript submitted to British Journal of Pharmacology
Summary of work:
The activity of γ-aminobutyric acid type A (GABAA) receptors, the predominant mediators of inhibitory neurotransmission in the mammalian central nervous system, is regulated by a
number of endogenous and pharmacologically important ligands. A prominent endogenous modulator is the transition metal ion, zinc. Zinc antagonises GABAA receptors
allosterically, by binding to discrete sites on the protein and preventing the channel opening in response to GABA. To date little is known about the structure of these sites and how
they regulate receptor function. The aim of the present study was to identify residues that mediate zinc inhbition of GABAA receptors. Through systematic site directed mutagenesis of
potential zinc binding residues in murine GABAA receptor α1 and β3 subunits, two clusters of residues on the α1 subunit were identified that profoundly influenced zinc potency. One
cluster is likely to contribute directly to the zinc binding site whilst the other helps mediate the allosteric actions of zinc. In addition, marked changes in zinc and GABA potency were
observed when specific α and β subunit residues were mutated in concert but less marked effects were seen when residues in one or other subunit were mutated alone. Although
expected for allosteric proteins, such non-linear interactions have not been reported previously. The present study provides the first complete description of multiple zinc binding sites
in a receptor protein and has important implications in the molecular understanding of receptor function.
Subsequent Appointment: Trainee Patent Attorney.
Bhavesh K Patel (KCL) 1 September 2000 – 30 September 2002
"Molecular cloning and functional characterization of two human 2-arylpropionyl-CoA epimerases / α-methylacyl-CoA racemases."
Reports: September 2001, April 2003, December 2003
Publications:
• B.K. Patel, J. Valentova and A.J. Hutt, Stereospecific HPLC analysis of flurbiprofen and its two major metabolites in serum and urine (in preparation).
• S.C. Tan, B.K. Patel, S.H.D. Jackson, C.G. Swift and A.J. Hutt, Stereoselectivity of ibuprofen metabolism and pharmacokinetics following the administration of the racemate to
healthy volunteers (in preparation).
• B.K. Patel, J. Valentova and A.J. Hutt, Chromatographic separation and enantiomeric resolution of flurbiprofen and its major metabolites, Chromatographia (submitted)
• J.R.S. Hoult, B.R. Jackson, E. Benicka, B.K. Patel and A.J. Hutt, Chromatographic resolution, chiroptical characterization and preliminary pharmacological evaluation of the
enantiomers of butibufen, a comparison with ibuprofen, J. Pharm. Pharmacol. 51 (1999) 1201-1205.
• B.K. Patel and A.J. Hutt, Enantiospecific analysis of flurbiprofen and its major metabolite in plasma by chiral-phase chromatography. J. Pharm. Pharmacol. 51 (1999) 75.
• S.C. Tan, B.K. Patel, S.H.D. Jackson, C.G. Swift and A.J. Hutt, Ibuprofen Stereochemistry : double-the-trouble? Enantiomer, 4 (1999) 195-203.
• A. Slovakova, X.F. von Maltzan, B.K. Patel, A.F. Drake and A.J. Hutt, Chromatographic resolution, chiroptical characterization and urinary excretion of the enantiomers of
sulindac. Chromatographia, 48 (1998) 369-376.
• A.J. Hutt and B.K. Patel, Enantiospecific Bioanalysis. In: N.J. Gooderham (ed.), Drug Metabolism: Towards the Next Millenium. IOS Press, Amsterdam, 1998, pp196-212.
•
B.K. Patel, S.H.D. Jackson, C.G. Swift and A.J. Hutt (2003). Disposition of flurbiprofen in humans – effect of stereochemistry and aging. Xenobiotica., submitted.
10
• S.C. Tan, B.K. Patel, S.H.D. Jackson, C.G. Swift and A.J. Hutt (2003). Influence of age on the enantiomeric disposition of ibuprofen in healthy volunteers. Br. J. Clin. Pharmacol.,
in press.
• B.K. Patel, J. Valentova and A.J. Hutt (2003). Stereospecific analysis of flurbiprofen and its major metabolites in plasma and urine by chiral-phase liquid chromatography.
Chromatographia,57, 7-18.
• S.C. Tan, B.K. Patel, S.H.D. Jackson, C.G. Swift and A.J. Hutt (2002). Stereoselectivity of ibuprofen metabolism and pharmacokinetics following the administration of the
racemate to healthy volunteers. Xenobiotica, 32, 683-697.
•
B.K. Patel, J. Valentova and A.J. Hutt. (2002) Chromatographic separation and enantiomeric resolution of flurbiprofen and its major metabolites. Chromatographia, 55, 135-142.
• J.R.S. Hoult, B.R. Jackson, E. Benicka, B.K. Patel and A.J. Hutt (1999). Chromatographic resolution, chiroptical characterization and preliminary pharmacological evaluation of
the enantiomers of butibufen, a comparison with ibuprofen, J. Pharm. Pharmacol., 51, 1201-1205.
•
S.C. Tan, B.K. Patel, S.H.D. Jackson, C.G. Swift and A.J. Hutt (1999). Ibuprofen Stereochemistry : double-the-trouble? Enantiomer, 4, 195-203.
• A. Slovakova, X.F. von Maltzan, B.K. Patel, A.F. Drake and A.J. Hutt (1998). Chromatographic resolution, chiroptical characterization and urinary excretion of the enantiomers of
sulindac. Chromatographia, 48, 369-376.
Summary of Work:
The 2-arylpropionic acids, e.g. ibuprofen and ketoprofen, are an important group of chiral NSAIDs. The majority of these agents are used as racemates even though their major
pharmacological activity resides in the enantiomers of the S-configuration. However, in vivo the R-enantiomers undergo metabolic chiral inversion to their active S-enantiomers, a key
enzyme in this process is 2-arylpropionyl-CoA epimerase (2-APCAE) and the extent of inversion is structure-dependent. 2-APCAE, equivalent to the enzyme 
-methyacyl-CoA
racemase, also catalyzes a requisite inversion during bile acid synthesis and is involved in the metabolism of dietary-derived phytanic acid. Inactivating mutations in the 2-APCAE
gene underlie a late-onset neuropathy. In addition to these rare mutations a number of common non-synonymous polymorphisms also exist at this locus. The aim of the work was to
express both the native and polymorphic 2-APCAE variants in Sf-9 insect cells via recombinant baculoviruses for functional and biochemical analyses. A viral shuttle vector
(pFastBac-HTa) containing a cDNA encoding native 2-APCAE was constructed and used as template to generate variants via site-directed mutagenesis. Recombinant baculoviruses
were subsequently generated by transfection of Sf-9 insect cells with recombinant bacmid DNA obtained via site-specific transposition (Bac-to-Bac). This expression system can be
used to obtain affinity-purified (His)10-tagged 2-APCAE to enable detailed functional investigations using a range of racemic profen drugs. Altered activity with one or more variant
forms of the enzyme may be of potential pharmacogenetic significance for profen therapy.
Subsequent Appointment: Hoping to continue career in academia in the USA.
John Paul Malkinson (Sch Pharm) 1 October 2000 – October 2002
"Solid phase synthesis of C- and N-terminal carbohydrate modified peptides and lipopeptides."
Reports: April 2001, February 2002.
Publications:
Toth, I; Malkinson, J P: Flinn, N S; Drouillat, B; Horvath, A; Erchegyi, J: Idei, M; Venetianer, A; artursson, P; Lazorova, L; Szende, B; Keri, G J Med. Chem. 1999, 42, 4010-4013
Malkinson, J P; Falconer, R A; Toth, I J Org. Chem. 2000, 65, 5249-5252.
Malkinson, J. P.; Falconer, R. A. Solid-phase synthesis of C-terminal thio-linked glycopeptides. Tetrahedron Lett. 2002, 43(52), 9549-9552.
Summary of Work:
Chemical modification represents an attractive means by which to improve the delivery and bioavailability of a variety of potential peptide drugs, normally very poorly absorbed and
subject to rapid degradation, metabolism and clearance after oral administration. Conjugation of a glycolipid moiety to either the C-terminus or the N-terminus of a peptide results in
improved absorption across biological membranes and inhibition of recognition by proteases. A series of glycolipid-modified peptides were assembled using standard solid-phase
peptide synthesis. The stereochemistry and alkyl chain length of the lipid moiety were varied. The glycosyl component was varied to include simple monosaccharides, aminosugars,
uronic acids and disaccharides. Conjugation of the glycosyl component to the lipid component was achieved via O-glycoside, thioglycoside and glycosyl amide linkages. Varying the
nature of the glycosyl and lipid moieties, and the linkage between them allows optimisation of the absorption profiles of the conjugates and their stability towards enzymatic
degradation. C-terminal N-linked glycopeptides can be prepared from resin-bound glycosyl azides via a modified Staudinger reaction. C-terminal S-linked glycopeptides were
prepared from resin-bound 1-thiosugars using a Mitsunobu condensation.
Subsequent Appointment: Lecturer at the School of Pharmacy
11
Seah Kee Tee (KCL) 1 January 2001 – 31 March 2002
"The use of biodegradable microspheres for the pulmonary delivery of hydrophobic drugs."
Reports: December 2001, April 2002.
Publications:
Tee, S.K., Forbes, B., Larhrib, H., Marriott, C., Martin, G.P. Development of an in vitro dissolution-absorption model to evaluate the delivery of aerosolised drug from dry
powder inhaler formulations. Proceedings of drug delivery to the lung XII, London. (2001) 115-118.
Tee, S.K., Martin, G.P., Leeds, A.R., Walker, C., Kicman, A., Cowan, D.A., Marriott, C. The influence of a tertiary component on the in vivo disposition of salbutamol isomers
aerosolised from a dry powder inhaler formulation. Thorax, (2001) p51.
Tee, S.K., Marriott, C., Leeds, A.R., Walker, C., Kicman, Cowan, D.A., Martin, G.P. In vivo determination of salbutamol sulphate delivered from a dry powder inhaler
formulation in human plasma and urine by solid phase extraction/gas chromatography-mass spectrometry (SPE/GC-MS). British Pharmaceutical Conference 2001 Abstract
Book, Glasgow. (2001) 15.
Tee, S.K., Martin, G.P., Leeds, A.L., Walker, C., Kicman, A., Cowan, D.A., Marriott, C. The effects of fine lactose on the in vitro and in vivo delivery of salbutamol from a dry
powder inhaler formulation. Am. J. Respir. Crit. Care Med. 163 (2001) p. A163.
Tee, S.K., Martin, G.P., Leeds, A.L., Walker, C., Kicman, A., Cowan, D.A., Fallon, J., Marriott, C. In vitro/in vivo evaluation of the performance of two lactose carrier-based dry
powder aerosol formulation for the delivery of salbutamol to the lung. J. Aerosol Med. 14 (2001) p130.
Tee, S.K., Marriott, C., Zeng, X.M. and Martin, G.P. The use of different sugars as fine and coarse carriers for aerosolised salbutamol sulphate. Int. J. Pharm. 208 (2000) 111123.
Zeng, X.M., Martin, G.P., Tee, S.K., Ghoush, A. and Marriott, C. Effects of particle size and adding sequence of fine lactose on the deposition of salbutamol sulphate from a
dry powder formulation. Int. J. Pharm. 182 (1999) 133-144.
Tee, S.K., Zeng, X.M., Martin, G.P. and Marriott, C. The role of carrier type and fine particle concentration on the deposition of salbutamol sulphate from dry powder aerosols
in vitro. J. Pharm. Pharmacol. 51 (1999) S325.
Tee, S. K., Martin, G.P., Marriott, C. and Zeng, X. M. The effect of type and concentration of various fine sugar particles on the deposition of salbutamol sulphate from dry
powder aerosols in vitro. Proceedings of drug delivery to the lung X, London. (1999) 33-37.
Zeng, X.M., Martin, G.P., Tee, S.K. and Marriott, C. The role of fine particle lactose on the dispersion and deaggregation of salbutamol sulphate in an air stream in vitro. Int. J.
Pharm. 176 (1998) 99-110.
Tee, S. K., Zeng, X. M., Martin, G. P., and Marriott, C. The influence of carriers and mixing time on the mixing homogeneity of salbutamol sulphate in dry powder aerosols.
Proceedings of drug delivery to the lung IX, London. (1998) 188-191.
Zeng, X. M., Tee, S. K., Martin, G. P., and Marriott, C. Improving the delivery efficiency of dry powder inhalers (DPIs) by adding fine carrier particlers to powder formulation.
Thorax. 51 (1996).
Zeng, X.M., Tee, S.K., Martin, G.P. and Marriott, C. Effects of mixing procedure and particle size distribution of carrier particles on the deposition of salbutamol sulphate from
dry powder inhaler formulations. Proceedings of drug delivery to the lung VII, London. (1996) 40-43.
Summary of Work:Following the deposition dry powder aerosols on the surface of the respiratory epithelium, drug bioavailability will depend on dissolution and absorption. To
study these processes in vitro requires the reproducible delivery of formulations to cultured respiratory epithelial monolayer. In this study respirable formulations were
manufactured using discretely-sized monodisperse fractions (1.0, 3.0 and 4.5 µm) of salbutamol sulphate (SS), prepared using an Andersen Cascade Impactor, blended with
lactose carrier (63-90 µm) in the drug to carrier ratio of 1:67.5. The twin stage impinger of the British Pharmacopoeia was modified to deliver SS precisely and evenly to
Transwell cell culture supports which contained monolayered respiratory epithelial (16HBE14o-) cells (Tranepithelial resistance ~ 150-200 Ω.cm2, across the Transwell filters
(6.5 mm i.d., 0.4 µm pore size)). Formulations containing 3.0 and 4.5 µm SS resulted in better deposition than that containing 1.5 µm. The amount of formulation aerosolised
was then adjusted to deliver equivalent mass (3.0-3.8 µg) of SS from each particle-sized formulation evenly across the surface of the cell cultured supports. The dissolution-
12
absorption of SS across the monolayered cell membrane was evaluated using the solid phase extraction/gas chromatography-mass spectrometry method. In conclusion, an in
vitro dissolution-absorption model using a cultured respiratory epithelial cells on a Transwell supports in a modified twin stage impinger has been developed, and it has the
value in evaluating the effect of formulation factors on the delivery of aerosolised drug from dry powder inhaler formulations.
Subsequent Employment: Position:
Product Development and Formulation Scientist
Team:
Intranasal Product Development
Department:
Company:
New Chemical Entities, Pharmaceutical Development
GlaxoSmithKline
Nicola Potz (KCL) 8 January 2001 – 4 June 2002
"Elucidation of the genes involved in bacterial resistance acquired due to prolonged exposure to biocide."
Reports: November 2001, May 2002
Publications:
Potz, N. A. C. & Forbes, B. (2002) A colorimetric assay for detecting catalase activity in Staphylococcus aureus. J. Microbiol. Meths. In Preparation.
Potz, N. A. C. & Forbes, B. (2001) The effect of adaptive tolerance to chlorhexidine on susceptibility to other biocides. J. Pharm. Pharmacol. 53 (Supplement), 114
Potz, N. A. C. & Forbes, B. (2001) Characterisation of adaptive tolerance to chlorhexidine: Laboratory experiments. Abstracts of the Society for Applied Microbiology Summer
Conference 2001.
Summary of Work: Pseudomonas aeruginosa has been shown to have the ability to acquire tolerance to in-use levels of the biocide Hibitane® (chlorhexidine gluconate) in response
to prolonged exposure to increasing concentrations. Upon removal of the biocide, the tolerance level decreases, although to a level greater than that of the wild-type organism. This
intermediate level of tolerance has been shown to be stable. Pseudomonads tolerant to an in-use concentration of Hibitane® showed increased susceptibility to the biocides Hycolin®
and Milton®, but slightly decreased susceptibility to Tego2000®. This decrease in Tego2000® susceptibility, however, was not to a level approaching the in-use concentration of this
biocide. The acquisition of tolerance to Hibitane® by Ps. aeruginosa has also been shown to increase the organisms susceptibility to the anti-pseudomonad antibiotics ciprofloxacin,
gentamicin and ceftazidime. The long feared link between bacterial biocide and antibiotic resistance was not present under the circumstances tested, and continual exposure to a
biocide, whilst possibly causing an increase in tolerance to that biocide, may not ultimately cause failure of treatment with other biocides or antibiotics.
Subsequent Employment: Clinical Scientist (Grade 11) at the Antibiotic Resistance Monitoring and Reference Laboratory (ARMRL), Central Public Health Laboratory, 61 Colindale
Avenue, London, NW9 5HT. I will be responsible for the sentinel surveillance of resistance i.e. the management of national antibiotic resistance surveys.
Zu Dong Liu (KCL) – 1 October 2001 – 30 June 2003
"Design and application of novel iron-specific fluorescent probes for monitoring intracellular labile iron pools"
Reports: April 2002, August 2004
Publications:
Liu, Z. D.; Kayyali, R.; Hider, R. C. Porter, J. B. and Theobald, A. E. (2002) Design, synthesis, and evaluation of novel 2-substituted 3-hydroxypyridin-4-ones: Structure-activity
investigation of metalloenzyme inhibition by iron chelators. J. Med. Chem. 45: 631-639.
Liu, Z. D. and Hider, R. C. (2002) Design of clinically useful iron(III)-selective chelators. Med. Res. Rev. 22: 26-64.
Liu, D.Y.; Liu, Z. D.; Piyamongkol, S.; Lu, S. L. and Hider, R. C. (2002) Characterization of two isomeric beta-D-glucosiduronic acids derived from 1,2-diethyl-3-hydroxypyridin-4-one
(CP94) in rat liver homogenate incubates. J. Pharm. Pharmacol 54: 951-957.
Liu, D.Y.; Liu, Z. D. and Hider, R. C. (2002) Oral iron chelators - development and application. Best Practice & Research Clinical Haematology 15 (2): 369-384.
Yoshimura, E.; Liu, Z. D.; Khodr, H and Hider R. C. (2002) Purity of commercial morin products and their spectrophotometric characterization. Bunseki Kagaku 51 (10): 979-982.
Liu, Z. D. and Hider, R. C. (2002) Design of iron chelators with therapeutic application. Coordin. Chem. Rev. 232 (1-2): 151-171.
Liu, Z. D.; Liu, D. Y. and Hider, R. C. (2002) Iron chelator chemistry. In: Iron Chelation Therapy. Hershko C., Eds. Adv. Exp. Med. Biol. 509: 141-166.
Hider, R.C. and Liu, Z. D. (2003) Emerging understanding of the advantage of small molecules such as hydroxypyridinones in the treatment of iron overload. Curr. Med. Chem. 10
(12): 1051-1064.
Zhou, T.; Hider, R. C.; Liu, Z. D. and Neubert, H (2004) Iron(III)-selective dendritic chelators. Tetrahedron Lett 45 (51): 9393-9396
13
Ma, Y. M.; Luo, W.; Quinn, P. J.; Liu, Z. D. and Hider, R. C. (2004) Design, synthesis, physicochemical properties, and evaluation of novel iron chelators with fluorescent sensors. J.
Med. Chem. 47 (25): 6349-6362.
Luo, W.; Ma, Y. M.; Quinn, P. J.; Hider, R. C. and Liu, Z. D. (2004) Design, synthesis and properties of novel iron(III)-specific fluorescent probes. J. Pharm. Pharmacol 56 (4): 529536
Hider, R. C. and Liu, Z. D. (2004) Siderophores. In: Encyclopedia of Supramolecular Chemistry. Atwood J. L. and Steed J. W. Eds, DOI: 10.1081/E-ESMC-120012714, Marcel
Dekker.
Piyamongkol, S.; Zhou, T.; Liu, Z. D.; Khodr, H. H. and Hider, R. C. (2005) Design and characterisation of novel hexadentate 3-hydroxypyridin-4-one ligands. Tetrahedron Lett 46 (8):
1333-1336.
Ma, Y. M.; Luo, W.; Camplo, M.; Liu, Z. D. and Hider, R. C. (2005) Novel iron-specific fluorescent probes. Bioorgan. Med. Chem. Lett. 15 (14): 3450-3452.
Z D Liu, D Y Liu and R C Hider (2001) Design and Application of Novel Iron-specific Fluorescent Probes for Monitoring Intracellular Labile Iron Pools. Bioiron 2001 – World Congress
on Iron Metabolism. Cairns, Australia (August 18-22) 092 (Oral Presentation)
Z D Liu, Y M Ma and R C Hider (2002) Design of Iron-specific Fluorescent Probes for Monitoring Intracellular Labile Iron Pools. Biometals 2002: 3rd International Biometals
Symposium. London, UK (April 11-13) C39 (Poster Presentation)
Summary of Work: Dr Liu continued his work on the design of therapeutically useful iron chelators which led to a series of original peer reviewed publications and four review
articles. During this period he also began work on the design of iron-sensitive fluorescent probes. This line of research has also been productive leading to three peer reviewed
publications. Dr Liu’s work continues to be a main theme of Professor Hider’s research effort.
Subsequent Appointment: In 2003 Dr Liu was appointed Managing Director of Great Chinese Herbal Medicine Ltd which trades as ‘Dr China’. This is an extremely successful
company coordinating over 80 clinical centres in Southern England. The clinical centres practise, Traditional Chinese Herbal Medicine, Massage and Acupuncture. Dr Liu is keen to
improve the quality control of these traditional treatments and towards this end has recently formed a UK recognised charity “Natural Medicines Research (UK)”. This charity will fund
staff in the newly formed “Centre for Natural Medicines Research” which is based at King's College London. Dr Liu plans to spend 40% of his time in this Centre in order to facilitate
the development of state of the art methods for the quality control and formulation of herbal medicines.
Vikas Jaitely (Sch Pharm) 25 October 2001 – 31 October 2004
"Ionic liquid mediated pulsatile delivery of proteins and peptides"
Reports: April 2002, May 2004, April 2005
Publications in process for submission
V. Jaitely and A.T. Florence, Water Immiscible Room Temperature Ionic Liquids: Some Properties Relevant to their Pharmaceutical Use, Pharmaceutical Research.
V. Jaitely, P. Song, A. Karatos and A.T. Florence, Water-immiscible room temperature ionic liquids as new matrix for electrically controlled drug delivery vehicles, Pharmaceutical
Research. Submitted
Summary of Work:
Water immiscible room temperature ionic liquids (RTILs) also referred to as ‘green solvents’ were investigated during the tenure of the Maplethorpe Fellowship for their potential for
use in pharmaceutical applications especially with regard to the formulation of controlled release delivery systems. Their use as reservoirs releasing solubilised drugs under the
influence of an electrical current was investigated, and promised the basis of an interesting pulsed release system. Butyl, hexyl and octyl derivatives of 1-methyl imidazole were
synthesized combined with hexafluorophosphate cation. Highly viscous RTILs had high densities and surface tensions the rate of 35-45 mN/m depending on alkyl chain length.
Although largely immiscible with water, hydrophilic RTILs have a tendency to absorb water to the saturation levels of around 10%, 5% and 3% w/w respectively in the butyl, hexyl and
octyl analogues. The presence of water markedly decreases their viscosities and density. The nature of RTILs allow solubilisation of water-soluble solutes such as sucrose.
Partition coefficients of some water soluble and some water insoluble drugs were determined using the shake flask method. The log P values of sucrose, penicillin V potassium salt,
dexamethasone, progesterone and dehydroxyepiandosterone between BMIM, HMIM, OMIM and water were found to be linearly correlated with their octanol-water log P values.
Release profiles monitored showed a slow release patterns for both water soluble sucrose and water insoluble dexamethasone recorded over a period of 48 h. The application of
low intensity electric pulses to the RTIL significantly enhanced the release of these molecules. The rate of release for sucrose was 3 times faster on application of current XMA and
the rate for dexamethasone was doubled. An increase in release rates were generally observed with an increase in current from 1mA to 5mA. The change in release patterns on
passage of current is due to the change in molecular packing of the ionic liquid array. The changes in surface tension and viscosity recorded for RTILs partially explained the release
data finding. These results could be instrumental in the future in design of pulsatile systems. Further combinations of RTIL with a polymer would be an ideal situation for making
electrically stimulated polymers. Toxicity studies undertaken using the MTT assay protocol using Ca Co-2 cells with saturated aqueous solutions of RTILs. 90% of cells were
14
predominantly in viable state, therefore indicating towards non-toxic behaviour of RTILs. An increase in alkyl chain length increases toxicity levels.
CONCLUSIONS
During the study we were able to demonstrate the potential of RTILs in pharmaceutical formulation design. Hydrophobic RTILs could serve as alterations to polymer matrix in
designing of controlled release systems and in design of electrically modulated pulsatile systems. Further studies by use of potential drug molecules would be desirable to exploit the
growth potential of technology.
WORK PROGRESS
With support from the Heptagon Fund, Bloomsbury Consortium funding for commercial assessment of the work was undertaken.
Patent application was filed on 25th March Application number: 0505779.9.
Current Post: Postdoctoral Fellow, SoP
Nilesh Patel (KCL) 2 January 2002 – 1 January 2004
"Delivery of DNA by pressurised metered dose inhalers; formulations and testing"
Reports: November 2002, January 2004
Publications:
Fluorinated Ionic Surfactant Microemulsions in Hydrofluorocarbon 134a (HFC 134a). J. Coll. Inter. Sci. Received 11/10/02. In press
Formation of Fluorinated Non-Ionic Surfactant Microemulsions in Hydrofluorocarbon 134a (HFC 134a). J. Coll. Inter. Sci. Received 11/10/02. In press.
Dodia V, Patel N, Hutt AJ, Forbes B. (-)-R-Enantiomers of Profen NSAIDS exhibit antiproliferative activity in vitro. AAPS: submitted (2002).
Dodia V, Patel N, Hutt AJ, Forbes B. Antiproliferative effects of sixteen non-steroidal anti-inflammatory drugs on Caco-2 cells. J. Pharm. Pharmacol. 54: S101 (2002).
Dodia V, Patel N, Hutt AJ, Forbes B. Stereoselective and metabolite-mediated antiproliferative effects of non-steroidal anti-inflammatory drugs. J. Pharm. Pharmacol. 54: S100-101
(2002).
Patel N, Murray JG, Forbes B. Development of physiological in vitro drug transport methods. 8th United Kingdom-Ireland Controlled Release Society Symposium. Loughborough, UK.
17th January (2002).
Patel N, Murray JG, Forbes B. Simulated intestinal fluids for use with the Caco-2 drug absorption model. British Pharmaceutical Conference Abstracts 55 (2001).
Arnot L.F, Minet A, Patel N, Royall P, Forbes B (submitted)
Solution calorimetry as a tool for the investigation of drug dissolution in complex media. Thermochimica acta
Patel N, Murray J, Forbes B (submitted)
Design and biocompatibility of Simulated Intestinal Fluids for use with the Caco-2 oral drug absorption model
Patel N, Marlow M, Lawrence MJ (2003)
Fluorinated ionic surfactant microemulsions in hydrofluorocarbon 134a (HFC 134a): J. Coll. Inter. Sci., 258, Issue 2, 354-362
Patel N, Marlow M, Lawrence MJ (2003)
Formation of fluorinated nonionic surfactant microemulsions in hydrofluorocarbon 134a (HFC 134a): J. Coll. Inter. Sci., 258, Issue 2, 345-353
Summary of Work:
Complexation of pDNA encoding for luciferase with the chosen lipids (DOTAP:DOPE 1:1 w/w) in aqueous solution was shown using gel electrophoresis. However in the presence of
surfactant (Tween 80), complexation efficiency was diminished or removed. In the absence of surfactant these complexes were shown to be able to transfect A549 cells. Zeta
potential measurements showed that at low surfactant concentrations no potential could be detected compared to the dispersions without surfactant. Visual assessment of the
aerosol formulations showed that without surfactant the lipoplex could not be formulated. In the presence of varying concentrations of surfactant, a clear solution was obtained up
until the solubility limit of surfactant in propellant was reached. The addition of ethanol gave milky uniform dispersions, which did not phase separate on standing. However. an
optimum formulation still requires elucidation. Spraying of the aerosol formulations however, gave a demonstrable release of DNA, of which the ability to transfect cells still remains
unknown.
Subsequent Employment: Lecturer in Pharmaceutics within the Pharmacy Department of King's College London
15
Sara Garfield (Sch Pharm) – 16 April 2002 – 12 August 2005
"Building a theoretical model to measure patients' desire for involvement in the medicines management of chronic conditions"
Reports: March 2003, March 2004, February 2005, April 2006.
Publications:
in
progess
Summary of Work: A concordant approach to medicine taking where patients and healthcare professionals make decisions about the use of medicines in partnership has been
advocated in the UK (RPSGB & MSD, 1997). However, it has also been recognised that patients may wish to hand over control to the healthcare professional (RPSGB & MSD
1997). Previous studies, mainly conducted in the US, have investigated preferences for general involvement in medical decision-making generally (Faden, Becker, Lewis, Freeman,
& Faden, 1981, Lidz, Meisel, Osterweis, Holden, Marx, & Menetz ,1983, Strull, Lo, & Charles, 1984, Sutherland, Llewellyn-Thomas, Lockwood, & Trichler, 1989, Ende, Kazis , Ash, &
Moskowitz, 1989, Beisecker & Beisecker, 1990, Nease & Brooks, 1995, Bradley, Zia, & Hamilton, 1996, Blanchard, Labrecque, Ruckdeschel, & Blanchard, 1998, Mckinstry, 2000,
Arora & Mcorney, 2000, Degner, & Sloan, 1992, Hack, Degner, & Dyck ,1994, Degner et al. 1997, Blackhall , Murphy, Frank, Michel, &, Azen, 1995, Young, & Klingle, 1996, Deber,
Kraetschmer & Irvine, 1996, Entwistle, Skea, & ODonnell, 2001, Krantz, Baum & Wideman, 1980) but have not focussed on decisions regarding medicine use. In addition, many
studies have used small, convenience samples, have been limited to bivariate analysis and have asked about hypothetical rather than real situations. The current study developed a
theoretical model of patients’ desire for involvement in decision-making concerning the use of medicines for chronic conditions in the UK. The model was tested in a large
representative sample of patients with one of two clinical conditions. Multivariate analysis revealed that age, social class and clinical condition were associated with preferences for
involvement in decision-making concerning the use of medicines for chronic illness but gender, ethnic group, concerns about medicines, beliefs about necessity of medicines, health
status, quality of life and time since diagnosis were not. In total, the model explained 14% of the variation in preference for involvement in decision-making. Variation in preference
may be very difficult to predict within a population. To work concordantly, prescribers may therefore need to ask patients about their decision-making preference during consultations.
Hendrik Neubert (KCL) 1 August 2002 – 31 July 2004
"The Development of an Affinity Capture MALDI TOF Mass Spectrometric Method for the Detection and Quantification of the Glycoprotein Hormones Human Chorionic Gonadotropin
(hCG) and Human Erythropoietin (hEPO) from Biological Fluids".
Reports: March 2003, October 2004
Publications:
Published Articles and those in press
Neubert H, Knights KA, de Miguel YR, Cowan DA. MALDI TOF Post-Source Decay Investigation of Alkali Metal Adducts of Apolar Polypentylresorcinol Dendrimers, Macromolecules,
2003;36(22):8297-8303.
Fernandez Ocaña M, Neubert H, Przyborowska AM, Parker R, Bramley PM, Halket JM, Patel RKP. BSE Control: Detection of Gelatine-Derived Peptides in Animal Feed by Mass
Spectrometry, Analyst 2004;129(2):111-115.
Neubert H, Fernandez Ocaña M, Patel RKP, Halket JM. α-Cyano-4-Hydroxycinnamic Acid Cluster Interferences in MALDI Spectra of a Hydrochloric Acid Digest, J. Am. Soc. Mass
Spectrom. 2004;15(3):336-343.
Fernandez Ocaña M, Jarvis J, Parker R, Bramley PM, Halket JM, Patel RKP, Neubert H. C-Terminal Sequencing by Mass Spectrometry: Application to Gelatine-Derived Proline-Rich
Peptides, Proteomics 2005;5(3): in press.
Zhou T, Hider RC, Liu ZD, Neubert H, Iron(III)-Selective Dendritic Chelators, Tetrahedron Lett. 2005 in press.
Saurin AT, Neubert H, Brennan JP, Eaton P, Widespread sulfenic acid formation in tissues in response to hydrogen peroxide, Proc. Natl. Acad. Sci. U. S. A., minor revision.
Submitted Publications
Kayyali R, Evans PJ, Waterfield CJ, Khodr H, Neubert H, Devanur LD, Hider RC, The Fenton-like Activity of Iron(III) in the Presence of 3-Hydroxy-pyridin-4-ones and its Effect on
Intracellular Glutathione Levels, Biochem. Pharmacol., submitted.
Malatos S, Neubert H, Kicman AT, Iles RK, Molecular identification of hCG associated fragments (HAF), implicated as the causative agents of HIV Kaposi’s sarcoma apoptosis, J.
Natl. Cancer Inst., submitted.
Aldred S, Sozzi T, Mudway I, Grant MM, Neubert H, Kelly FJ, Griffiths HR, Alpha tocopherol supplementation elevates pro-apolipoprotein AI but decreases HDL cholesterol
concentration; an explanation for the vitamin E paradox?, Circulation, submitted.
16
Manuscripts in Preparation
Halket JM, Waterman D, Przyborowska AM, Gröger T, Bernhard M, Zwitkovits M, Tüchler M, Bramley PM, Fraser PD, Fernandez Ocaña M, Neubert H, Patel RKP, Smith NW.
Charge Derivatization and Ion Trap LC/MSn with Mass Spectral Library Matching for Organic Acid Analysis and Metabolic Profiling, Rapid Commun. Mass Spectrom., in preparation.
Berndt UEC, Zhou T, Hider RC, Liu ZD, Neubert H, Mass Spectrometry of Chelator-Terminated Dendrimers and Their Synthetic Intermediates, Eur. J. Mass Spectrom., in
preparation.
Some other publications are expected resulting from collaborative proteomics studies. Some key examples are the mapping of cell surface receptor ligands in a diabetes study with
the Immunobiology group at Guy’s Hospital, characterisation of posttranslational modifications of a cardiac membrane protein with the Cardiovascular Division at St. Thomas’
Hospital and identification of differentially expressed bacterial proteins in collaboration with the Microbiology Group at the School of Life Sciences.
Summary of Work:
The research accomplished during the term of the Maplethorpe Fellowship is categorised into three main areas; in fundamental advances in peptide mass spectrometry, in
proteomics studies utilising mass spectrometry and in the characterisation of synthetic macromolecules. Firstly, a novel sequencing method was developed for proline-rich peptides
that are often unusually difficult to characterise by standard collision-induced dissociation mass spectrometry. In addition, a fundamental investigation into the chemical nature of
MALDI-matrix interferences enabled discerning their mass spectral signals from those of peptides and allowed using them for internal mass calibration. Secondly, a proteomics study
identified human chorionic gonadotropin associated fragments as proteins with possible activity against HIV-related Kaposi’s sarcoma and another investigation established how
proteolytic fragments of gelatine can reveal contamination of cattle feed with meat and bone meal. Other studies established how the widespread oxidative post-translational
modification of protein cysteinyl thiols by hydrogen peroxide could be detected and how alpha-tocopherol (AT) exerts an up-regulation of serum apolipoprotein A1 that suggests a
novel mechanism of AT action. Thirdly, the research presented herein provided innovative mass spectrometric tools advancing the analysis of dendrons and dendrimers. In particular,
structures of polypentylresorcinol dendrimers and iron(III)-selective hexadentate chelator-terminated dendrimers were verified and their properties established.
Subsequent Post: Manager, Clinical Proteomics & Biomarker Discovery Clinical Assay Group, Pfizer Global Research & Development, Kent.
Miss Nicola Wilson (Sch Pharm) 7 October 2002 – 6 October 2004
"The Identification of Counterfeit Medicines by Near Infrared Spectroscopy and Near Infrared Microsocopy".
Reports: April 2003, October 2004.
Publications:
British Pharmaceutical Conference (BPC 2003, Harrogate)
"NIR Methods of Detecting Counterfeit Products"
Invited speaker; Combating the Counterfeiting of Medicines Session.
BPC 2004 (Manchester)
"The Use of Near Infrared Microscopy for the Detection of Counterfeit Viagra Tablets"
Speaker, Short Papers in Pharmaceutical Analysis Session, organised by the Joint Pharmaceutical Analysis Group (JPAG). Awarded a JPAG prize for the oral presentation.
Also: Press Release issued by the Royal Pharmaceutical Society of Great Britain, which attracted national media interest.
Hospital Pharmacy Europe, October 2004, invited article (In Press)
"New Techniques to Identify Counterfeit Medicines: Near Infrared Spectroscopy and Near Infrared Microscopy"
American Association of Pharmaceutical Scientists (AAPS 2004, Baltimore)
"The Use of Near Infrared Microscopy for the Identification of Counterfeit Viagra Tablets"
Poster
Summary of Work:
A Near Infrared (NIR) microscopy method was developed that allows the identification of counterfeit Viagra and shows the distribution within the tablet of sildenafil, the active
pharmaceutical ingredient (API) and tablet excipients. The method is also able to suggest the identity of components found in counterfeit Viagra tablets and shows their distribution
2
), individual
within the tablet. An image map is obtained which contains NIR spectra obtained from successive small areas of the tablet. Because these areas are so small (30
m
particles can be identified. These spectra are compared to a spectral library containing sildenafil citrate, Viagra excipients and compounds likely to be present in counterfeit Viagra.
Samples of authentic Viagra and counterfeit Viagra were studied, in addition to several other proprietary versions of sildenafil citrate tablets to help validate the method. Samples of
Viagra were also obtained over the internet. It was found the sildenafil in counterfeit tablets showed greater clumping and that counterfeit tablets appeared to contain excipients such
as lactose and starch which are not present in genuine Viagra. Half of the internet Viagra samples obtained were identified as possible counterfeits using this NIR microscopy
17
method. This was confirmed by Pfizer using conventional techniques. The distribution maps of the sildenafil content of the identified counterfeit Viagra tablets were stored in a
database. This will allow comparison with other counterfeit samples, which will help in relating samples to a common source and subsequently help with the fight against the
trafficking of counterfeit medicines.
Subsequent Post: From 1 December 2004 - Postdoctoral Fellow at the Centre for Pharmacognosy and Phytotherapy, SoP.
Dr Olivia Corcoran (KCL) 2 January 2003 – 1 January 2005
"NMR studies on the protein binding of reactive metabolites from nonsteroidal anti-inflammatory drugs."
Reports: December 2003, May 2005, December 2005.
Publications:
Corcoran, O. NMR spectroscopy as a versatile analytical platform for toxicology research. Handbook of Toxicogenomics: Strategies and Applications. (2005) Wiley-VCH. Edited by J.
Borlak. Chapter 8, p163-183.
Griffin, J. L. and Corcoran, O. High-resolution magic-angle spinning 13C NMR spectroscopy of cerebral tissue. Magnetic resonance materials in physics, biology and medicine, (2005)
18, 1, 51-56.
O. Corcoran. Identification of reactive metabolites and potential reactive intermediates using NMR spectroscopy: NSAID case histories. Invited speaker, UK Drug Metabolism Group
1-day meeting, 18 Feb (2004), Northants, UK.
O. Corcoran. “Hyphenated NMR in clinical chemistry.” Invited speaker, San Diego NMR Meeting, December (2003), California.
O. Corcoran and M. Spraul. “LC-NMR-MS in drug discovery.” Drug Discovery Today, (2003), 8, 14, 624-631.
M. Spraul, A. S. Freund, R. E. Nast, R. S. Withers, W. E. Maas and O. Corcoran. “Advancing NMR sensitivity for LC-NMR-MS using a cryoflow probe: Application to the analysis of
acetaminophen metabolites in urine.” Analytical Chemistry, (2003), 75, 6, 1536-1541.
O. Corcoran, A. S. Freund, W. E. Maas and M. Spraul. “A novel cryoflow probe for LC-NMR-MS.” HPLC2003 Abstracts Nice, France (2003).
O. Corcoran, K. Mountfort and S. Jickells. “Chemical characterisation of latent fingerprint residue using ultra-sensitive NMR spectroscopy. Analytical Chemistry, in preparation.
K. Chow, P. J. Houghton and O. Corcoran. Identification of oligomeric proanthocyanidin complexes (OPCs) from the bark of Funtumia elastica. Planta medica, in preparation.
Summary of Work: Reactive acyl glucuronide metabolites of several non-steroidal anti-inflammatory drugs (NSAIDs) are known to bind physiologically important proteins. However,
there remains controversy as to whether the formation of chemically reactive acyl glucuronide metabolites is linked to a higher incidence of serious adverse reactions of NSAIDs.
Toxicity may be caused by formation of immuno-reactive glucuronide protein adducts but mechanisms involving modification of active sites of enzymes and interactions with
structural proteins have also been suggested. As yet, no common toxicity mechanism for -1-O-acyl glucuronides has been established because -1-O-acyl glucuronides of different
drugs bind covalently to different proteins and on different sites on specific proteins. Specifically, recent evidence indicates that the isomeric glucuronide metabolites of diclofenac
bind important bile transporter proteins including MRP2 causing cholestasis. However, no methods were previously available that can distinguish between the positional isomers of
diclofenac -1-O-acyl glucuronides, and thus the reports on protein binding to MRP2 cannot distinguish which isomers are involved in the binding process. The knowledge of which
structural isomer is involved would be critical in constructing a quantitiative structure-binding model to explain the binding mechanism and, ultimately, to determine whether the
differences in protein binding reactivity between NSAID glucuronides is due to differential affinity of glucuronide isomers for physiologically important proteins.
The chemical reactivity of selected isomeric glucuronide metabolites was investigated in the course of this project. High performance liquid chromatography (HPLC) methods have
been developed for the measurement of the intrinsic chemical reaction kinetics of naproxen, ibuprofen and diclofenac glucuronides. Isolation of the glucuronide metabolites from
urine samples was achieved for naproxen and ibuprofen by analytical HPLC, however the isolation of multiple diclofenac glucuronides was not a viable approach. The isomeric
glucuronide products were found to have measured half lives of less than 5 mins under in vitro physiological conditions (37o C and pH 7.4). Structural identification of isomer NSAID
glucuronides was confirmed by NMR spectroscopy, but synthetic approaches will be required in order to generate sufficient diclofenac-beta-1-O-acyl glucuronide for protein-binding
studies.
For the first time, definitive structural identification of the isomeric glucuronides of tolmetin was achieved. HPLC assays were also investigated for diflunisal and tolmetin glucuronides.
These chemical reactivity studies of the isolated and purified NSAID acyl glucuronides under identical conditions of physiological pH and temperature will allow comparison of intrinsic
chemical reactivity that may affect protein-binding.
18
Other research undertaken during the course of this fellowship was the investigation of the chemistry of fingerprints by NMR with Dr Jickells, Department of Forensic Analysis at KCL
and the investigation of extracts from the tree Funtumia elastica with Professor Houghton.
Subsequent Post: In November 2004, Dr Corcoran was offered a permanent position as NMR Research Officer in the 5* Department of Chemistry at the University of Bristol. She
subsequently was offered and accepted a Senior Lectureship in Forensics in the School of Health and Bioscience at the University of East London, Stratford. She started at UEL on 1
January 2005 and is also a Visiting Research Fellow in the Department of Pharmacy until December 2007.
Dr Alessandra Gaeta (KCL) 1 June 2003 – 31 May 2005
"Design of iron chelators with the potential of treating neurodegenerative disorders."
Reports: March 2004, April 2005.
Publications:
“The crucial role of metal ions in neurodegeneration. The basis for a promising therapeutic strategy”
Review article submitted to the British Journal of Pharmacology.
Summary of Work:
Over the last two years, as a Maplethorpe fellowship, I have been working on a project whose title is “Design and Synthesis of Novel Iron Chelators with the Potential of Treating
Neurodegenerative Diseases”. The variety of factors and events involved in neurodegeneration renders the subject a major challenge. Neurodegenerative disorders, such as
Alzheimer's disease and Parkinson disease, can be considered as different pathological conditions sharing similar critical metabolic processes such as protein aggregation and
oxidative stress. Interestingly, both these metabolic alterations seem to be associated with the involvement of metal ions, particularly iron and copper. Therefore, chelation therapy
could be a valuable therapeutic approach, since metals are considered to be a pharmacological target for the rationale design of new therapeutic agents directed towards the
treatment of neurodegeneration. During the first year, a set of new iron chelators, with a 3-hydroxypyirid-4-one based molecular structure, has been successfully synthesised. Once
the synthesis was completed, I focused on the physical chemical characterisation, with both lipophicility and iron affinity being investigated as important requirements to evaluate and
improve the pharmacological activity. Furthermore, studies on two iron containing enzymes, lipoxygenase and tyrosine hydroxylase, have been carried out in order to verify the
potential toxicity, due to any inhibitory effect, of the new iron chelators. The physical chemical characterisation together with the biological work have been of considerable relevance
to the pharmacological in vitro studies, which at the same time, were undertaken on the synthesised compounds. All the new compounds have shown a certain degree of
neuroprotection in cortical neurones culture and three analogues have been chosen as lead compounds for further pharmacological investigation and synthetic work.
Subsequent Post:
Dr Naseem Mushtaq (SoP) 1 August 2003 – 31 July 2005
!Attenuation of bacterial virulence through enzyme-mediated capsule hydrolysis."
Reports: March 2004, July 2005
Publications:
Mushtaq, N., Redpath, M.B., Luzio, J.P., and Taylor, P.W. (2004) Prevention and cure of systematic Escherichia coli K1 infection by modification of the bacterial phenotype.
Anitmicrobial Agents and Chemotherapy. 48:5:0
Mushtaq, N., Redpath, M.B., Luzio, J.P., and Taylor, P.W. (2005) Treatment of experimental E. coli K1 infection with recombinant bacteriophage derived capsule depolymerase.
Journal of Antimicrobial Chemotherapy published online 24th May 2005.
Mushtaq, N., Redpath, M.B., Luzio, J.P., and Taylor, P.W. (2004) Prevention and cure of systematic Escherichia coli K1 infection by modification of the bacterial phenotype.
Antimicrobial Agents and Chemotherapy. 48:5:1503-1508.
Summary of Work:
The K1 capsular polysaccharide of E. coli is a homopolymer of α2-8 linked sialic acid (PSA) and is the virulence determinant of E. coli strains isolated from cases of neonatal sepsis
and meningitis. The strain is found to be profoundly resistant to attack by the humoral immune factors, and very difficult to treat through conventional therapy, therefore modifying the
phenotype of the strain through the removal of the K1 capsule with a depolymerase known as endoE will allow the immune system to recognise and destroy the antigen. The
therapeutic efficacy of endoE has revealed that an intraperitoneal injection between 20-0.25ug/100ul of PBS is able to clear systemic bacteraemia with almost 0% morbidity, whereas
in the controls there is almost a 100% morbidity. Therefore, an intraperitoneal injection of the enzyme is able to depolymerise the K1 capsular polysaccharide, thus stimulating
phagocytosis of the pathogen as observed with macrophages in vitro. The dissemination of bacteria appears to be the most concentrated during symptomatic illness in the lungs,
and the least in the brain, however during asymptomactic infection, the spread is fairly uniform amongst the organs, and the bacterial load far less extreme. The spread of
19
bacteraemia has also been found to be markedly age-dependent since two day-old rats have been found to be more susceptible to infection than five-day-old pups. Through these
studies it can be concluded that recombinant bacteriophage endoE is capable of not only curbing neonatal E. coli K1 infection, but can also be used prophylactically.
Subsequent Post: Working for a biotechnology company in Cambridge.
Dr Behrooz Nasseri (SoP) 1 October 2003 – 30 September 2005
"Development of a vesicular shuttle system within tether nanotube networks."
Reports: March 2004, October 2004, May 2006
Publications:
1. Nasseri, Behrooz, Fasciolo, Alessandro and Florence, Alexander T. (2005a) Exploring soft matter: liposomal vesicles and their tethers. J. Drug Target. 13(8-9): 471-7.
2. Nasseri, Behrooz and Florence, Alexander T. (2005b) The relative flow of the walls of phospholipid tether bilayers. Int. J. Pharm. 25; 298(2): 372-7.
3. Nasseri, Behrooz and Florence, Alexander T. (2005c) Effect of cholesterol and temperature on the elastic properties of niosomal membranes. Int. J. Pharm. 300; 95–101.
Nasseri, B. and Florence, A T, A vesicular shuttle: Transport of a vesicle within a flexible microtube, J.Control. Rel. 92 (2003) 233-240.
Summary of Work: An investigation into the formation of flexible tether nanotubes produced from surfactant vesicles. The study looked into the factors governing the
flexibility of such nanotube tethers which included the investigation of their composition, in particular the effect of cholesterol on the surface modulus of elasticity of the
vesicular membranes. The tether nanotubes once formed where subsequently used to transport particles and vesicles inside their channels. These channels were walled by
number of surfactant bilayers (multilamellar) whose relative movement was also looked into where it was found that the inner bilayers moved faster and further in relation to the
outer bilayers. Entrapment of polystyrene microspheres inside vesicles was achieved where the movement dynamics were studied inside such microchambers by the
utilization of particle tracking and matrix laboratory (MATLAB) software. It was possible to differentiate two types of movement inside a disturbed vesicle where the particles
positioned in the centre of a vesicle moved slower than the particles nearing the vesicle chamber wall. It was also found that under sink conditions particles followed a
Brownian motion until a point where their population effected the neighbouring nearby particles i.e. a caging effect.
Subsequent Post: Teaching position at King’s College London.
Dr Omid Masoud (KCL) – 1 April 2004 – 30 September 2005
"The in vitro assessment of aerosolised drug deposition using novel oropharyngeal models and the incorporation of inhalation profiles using an artificial lung."
Reports: March 2005, September 2005
Publications:
Summary of Work: Recent environmental concerns of global warming and the thinning of the ozone layer has led to the phase out of chlorofluorocarbons (CFCs) which have been
employed as propellants in the formulation of metered dose inhalers (MDIs). Replacement hydrofluoroalkanes (HFAs) appear to be a safe alternative, however, recent reports
suggest that HFA-containing salbutamol MDIs can block during continued use and therefore the required dose may not reach the patient’s airways. The compendial in vitro apparatus
for assessing drug deposition from inhaled formulations involves the use of an impinger or impactor. The systems mimic the respiratory tract in that they fractionate the emitted dose
according to aerodynamic particle size, larger particles are deposited higher up, and smaller ones further down the apparatus. The design of the inlet which is intended to mimic the
oropharynx has always been considered to be crucial but this has not been associated with concomitant sophistication. The biggest challenge encountered in pulmonary drug delivery
is to increase the bioavailability of the drug and achieve a more reproducible peripheral lung deposition. Dry Powder Inhaler (DPI) devices differ in their design and formulation
components and as a result exhibit different internal resistances. At a given pressure drop (4P), the flow (F) through the inhaler is governed by the internal resistance (R) of that
device where F = 4P/R. The internal resistance of a device has a great influence on the emptying of blister, redispersion of the powder mixture and respirable fraction. It is believed
that DPIs of the same design will exhibit consistent internal resistances. The Performance comparison of CFC and HFA-based MDIs (n=3) in terms of the particle size emitted upon
actuation and whether cleaning as per manufacturer instructions affected the size distribution during the lifetime use of the inhalers was investigated using the Twin Stage Impinger
and Spray-Tech techniques. Statistical Analysis of in vitro Deposition Studies of Dry Powder Inhalers (DPI) at 4 kPa, Low, Medium and High Flow Rates using different oropharyngeal
inlets was carried out using the minitab statistics package. The resistance of all UK marketed Accuhaler® devices was compared and inter-batch variability investigated. The HFA
based formulation generated particles with the most appropriate size range to enable efficient deposition to the target site of the lower airways. This was true whether the actuator
was washed or not. Analysis of variance (ANOVA) at 95% confidence interval, demonstrated that %inlet deposition and %FPF data differences for inlet, device and flow rate, were
significant. ANOVA also confirmed that there is only a 5% probability that interactions between inlets, devices and flow rate could be attributed to chance. The study indicates that
inlets constructed from modern imaging techniques are potentially more representative of the oropharyngeal region, and the respirable fraction of drugs predicted by in vitro
deposition of systems correlates better with the fraction attained in vivo from aerosolized formulations. Significant differences in resistance between different Accuhalers® were
20
observed. ANCOVA (α = 0.5) demonstrated that the means of Seretide® 250 µg Accuhalers® and Seretide® 500 µg Accuhalers® significantly differed from the other Accuhalers®
tested. The extended study on different batches of Seretide® Accuhalers® goes on to show that different batches of Accuhalers® of the same composition also show variations in
resistance. This suggests that drug delivery from the same inhaler device containing the same formulation and strength is also different.
Subsequent Post: Lecturer in the Department of Pharmacy, KCL
Dr Laila Kudsiova (KCL) – 1 October 2004 to 30 September 2006
"Cellular fate of non-viral gene delivery vectors"
Reports: July 2005, September 2006
Publications:
Laila Kudsiova and M. Jayne Lawrence, The effect of chitosan and chitosan-coated liposomes on the monolayer integrity and the permeability across Caco-2 and 16HBE14o-cells.
(currently being revised for submission).
Laila Kudsiova, M. Jayne Lawrence, and Anthony PR Brain. Potential gene delivery vectors composed of chitosan-coated liposomes encapsulating DNA. (currently being revised for
submission).
Richard D. Harvey, David J. Barlow, Anthony PR Brain, Philip Callow, Laila Kudsiova and M. Jayne Lawrence. The effect of electrolyte on the lamellarity of nonionic surfactant
vesicles. (submitted to Pharmaceutical Research).
Kudsiova L, Ho J, Hart S, Tabor AB, Lawrence MJ and Hailes HC. Lipoplex Ternary Delivery Systems Incorporating C14 Glycerol-Based Lipids. (To be submitted to Bioconjugate
Chemistry)
Ho J, Kudsiova L, Harvey R, Hart S, Tabor AB, Lawrence MJ and Hailes HC. Synthesis and Characterisation of Lipoplex Non-Viral Gene Delivery Systems Containing C14 GlycerolBased Lipids. (To be submitted to Bioconjugate Chemistry)
Elbs M, Mustapa MFM, Kudsiova L, Grosse S, Cook HEJ, Warley A, Lawrence MJ, Hart SL, Hailes HC and Tabor AB. Stabilised Integrin-Targeting Ternary LPD (Lipopolyplex)
Vectors for Gene Delivery Designed To Disassemble Within the Target Cell. (To be submitted to the Journal of the American Chemical Society)
Summary of Work:
A number of structurally related cationic lipids have been synthesised and tested for their transfection efficiency. Surprisingly, despite very small structural changes, such as the
positioning or the conformation of the double bonds, large differences in their transfection efficiencies were observed. In order to understand why some of those lipids transfected
better than others, the fate of the lipoplexes inside MDA-MB-231 cells has been examined. By fluorescently labelling the cationic lipids and the plasmid DNA with green BODIPY and
rhodamine labels respectively, it was possible to examine the intracellular fate of both the lipid and the DNA over a period of time using confocal fluorescence microscopy. It is
thought that the lipoplexes enter the cells via endocytosis, and are transported close to the nucleus via endosomes soon after internalisation. Over time, the fluorescence usually gets
more diffuse, probably indicating endosomal escape. Very little fluorescence was detected in the nucleus, however, indicating that nuclear localisation is the main barrier towards
achieving good transfection efficiency with those lipids. Cytotoxicity, measured using the MTT assay and microscopy, was another factor that could have contributed towards the low
transfection efficiency of some of the lipids.
Subsequent Post: Teaching Fellow in Pharmaceutics at the Pharmacy Department, KCL
Dr Catherine Lane (SoP) – 1 October 2004 to 1 May 2006
"Pharmaceutical proteomics of cytochromes P450; towards patient individualised anticancer therapy"
Reports: June 2005, March 2006
Publications:
C. S. Lane (2005) Mass spectrometry-based proteomics in the life sciences. Cell. Mol. Life Sci., 62: 848-869.
C. S. Lane, L. H. Patterson and W. J. Griffiths. Quantitative analysis of cytochrome P450 enzymes using SDS-PAGE and 18O stable isotope labelling. Manuscript in
preparation.
Catherine Lane, Yuqin Wang, Richard Betts, William Griffiths and Laurence Patterson. Comparative Cytochrome P450 Proteomics in the Livers of Immunodeficient Mice using
180 Stable Isotope Labeling. 2007 The American Society for Biochemistry and Molecular Biology, Inc.
21
Summary of Work:
The cytochromes P450 (P450s) are a superfamily of enzymes with a central role in the metabolism of drugs and xenobiotics. P450s can also be responsible for the conversion
of chemical toxins and procarcinogens to their toxic forms; this ability has been exploited in cancer chemotherapy, where several anticancer drugs require P450 activation to
exert their cytotoxic effects. A method has been developed that allows the relative quantification of P450 proteins from complex samples. Two samples for comparison are
analysed by SDS-PAGE and pairs of gel bands in the P450 molecular weight region excised and subjected to in-gel digestion with trypsin. The resultant peptides are
suspended with immobilised trypsin in either 18O-labelled water or normal-isotopic water. The trypsin catalyses the incorporation of the water oxygen at both peptide C-terminal
oxygen atoms, resulting in a differential mass labelling of peptide samples by 4 Da. Differentially labelled sample pairs are combined and analysed by mass spectrometry;
relative quantification is achieved via comparison of the abundance of 16O and 18O-labelled peptides. The method was validated using recombinant P450s and human liver
microsomes, applied to SDS-PAGE gels in known amounts. Comparative P450 proteomics was performed using an animal model, in which human tumours are grown on
immune-deficient mice. The compound 1,4 bis 2-(3,5-dichloropyridyloxybenzene) (TCPOBOP) has been shown to induce profound expression of P450s in this model. Liver
microsomes from control and TCPOBOP-dosed mice were analysed. To ensure no bias was introduced from the labelling procedure, two labelling experiments were
performed: in the first experiment the treated sample was labelled with 18O, and in the second the untreated sample was labelled with 18O. Relative quantification was
performed on peptides from 17 mouse P450 proteins; the levels of almost all the P450s quantified were affected by treatment with TCPOBOP. For each of the two labelling
experiments protein standard deviation values indicate a high level of reproducibility; in addition, agreement between the two labelling experiments is high. The ability to
quantify expression levels of P450s in healthy and diseased tissue on an individual patient basis prior to treatment would provide invaluable information regarding the fate of
selected drugs, and could ultimately lead to patient-individualised therapy.
Subsequent Post: Proteomics and Small Molecules Support Specialist, Applied Biosystems, Warrington, UK.
Miss Khuloud Al-Jamal (SoP) – 1 September 2005 to 31 August 2007
"Heparin-binding dendrimers as anti-angiogenic therapeutics in vivo and in vitro delivery"
Reports: August 2006, August 2007.
Publications:
Al-Jamal, K T; Buddle, J and Kostarelos, K. Dendrimer Nanocontainers Complex Doxorubicin and Enhance its Tumour Spheroid Penetration and Cytotoxicity. Full paper is in
preparation.
Al-Jamal, KT; Kanthou, C; Vargesson, N; Tozer, G M; Kostarelos, K: Dendrimeric Nanocontainers as Anti-angiogenic Therapeutics. Full paper is in preparation
Summary of Work:
The purpose of the project is to evaluate the therapeutic and tumor anti-angiogenic efficacy of newly synthesized, heparin-binding dendrimers alone, and in combination with other
therapeutic modalities such as chemotherapeutic drugs and gene delivery. Dendrimer-based anti-angiogenic therapeutics was developed and their activity was tested using various
models such as tubule formation assay, CAM assay, dorsal window chamber and finally their therapeutic efficacy was evaluated in vivo. Cationic polysine dendrimers have shown,
and at a relatively non-toxic concentration, to inhibit tubule formation, reduce tumour vascularity and led to tumour size reduction after subcutaneous or intravenous administration.
Subsequent Post: I am joining the School of Pharmacy as a Research Fellow in Nanomedicine, starting in September 2007.
Bosquillon, Dr Cynthia (KCL) – 1 October 2005 to 31 July 2007
“Adaptation of the isolated perfused lung (IPL) technique for screening inhalation products”
Reports: August 2006, July 2007.
Publications:
Book Chapters
- A.Tronde, C.Bosquillon, B.Forbes, The isolated perfused lung for drug absorption studies, in: Pharmaceutical Cell Biology- in situ, in vitro and in silico tools to study drug
absorption, K-J.Kim, C.Ehrhardt (eds.), Springer, New York, in press.
Conference abstracts
- C.Bosquillon, M.Madlova, N.J.Clear, B.Forbes, Can a simplified isolated perfused lung model be a useful drug absorption screen? J.Aerosol.Med 20, 190/P086 (2007)
- C.Bosquillon, N.J.Clear, J.Bennett, B.Forbes, Drug permeability in airway cell lines correlates with absorption from the rat lung, J.Aerosol.Med 20, 186/P071 (2007)
22
-
M.A. dos Santos, C.Bosquillon, T.Russomano, F.Falcão, C.Marriott, B.Forbes, Development of a diffusion chamber to evaluate in vitro respiratory epithelial cell layer
permeability in gravity. Aviation, Space, and Environment Medicine 78, 259 (2007).
- L.Belhadj Salem, C.Bosquillon, L.Delattre, G.P.Martin, B.Evrard, B.Forbes, Kleptose Crysmeb is a biocompatible solubiliser for pulmonary delivery of Ro 28-2653, a new
synthetic matrix metalloproteinase inhibitor, 3rd Pharmaceutical Sciences World Congress, Amsterdam, April 2007.
- M.Madlova, C.Bosquillon, B.Forbes, In vitro study into P-gp transport in airway epithelium, APS inhalation 2007, Bath, March 2007.
- C.Bosquillon, L.Belhadj Salem, L.Delattre, G.P.Martin, B.Evrard, B.Forbes, Effect of ß-cyclodextrin derivatives on bronchial epithelial cell layer permeability, Drug Delivery to the
Lung 17, 90-93 (2006)
- J.Patel, L.Belhadj Salem, L.Delattre, G.P.Martin, B.Evrard, B.Forbes, C.Bosquillon, Use of the MTT assay to evaluate the biocompatibility of β-cyclodextrin derivatives with
respiratory epithelial cells, J. Pharm. Pharmacol. Suppl 1: A64 (2006)
Presentations at Scientific Meetings
- C.Bosquillon, M.Madlova, N.J.Clear, B.Forbes, Can a simplified isolated perfused lung model be a useful drug absorption screen? Poster presentation at the 16th International
Congress of the International Society for Aerosols in Medicine, Tours, France, June 2007.
- C.Bosquillon, N.J.Clear, J.Bennett, B.Forbes, Drug permeability in airway cell lines correlates with absorption from the rat lung, Poster presentation at the 16th International
Congress of the International Society for Aerosols in Medicine, Tours, France, June 2007.
- C.Bosquillon, L.Belhadj Salem, L.Delattre, G.P.Martin, B.Evrard, B.Forbes, Effect of ß-cyclodextrin derivatives on bronchial epithelial cell layer permeability, Poster presentation
at Drug Delivery to the Lung 17, Edinburgh, December 2006.
Summary of Work:
The recent development of inhalation as a route for systemic drug delivery arises the need for simple and reliable methods to study pulmonary drug absorption. The ability of the two
airway cell lines Calu-3 and 16HBE14o- to predict drug absorption from the lungs was evaluated and for the first time, directly compared with that of normal human bronchial epithelial
(NHBE) cells. The permeability of a selected panel of compounds was measured in those three models and compared with absorption data from the rat lung. Similar log-linear
relationships were obtained (R2 ≈ 0.85), indicating airway cell layers can predict pulmonary absorption in vivo and NHBE do not offer any advantage over cell lines. In parallel, a
simple rat isolated perfused lung (IPL) model in which tissue viability is maintained for 90 min was developed under the UK Schedule One Programme. The absorption of the panel of
compounds was studied in this set-up and was shown to strongly correlate with absorption data from the rat lungs (R2 = 0.96). The relationship was as strong as that reported in more
complex IPL techniques, demonstrating a simple model is a useful tool for inhaled drug screening as well as for cell culture data validation.
Subsequent Post: Lecturer in Drug Delivery, University of Nottingham.
Hitch, Dr Geeta (SoP) – 1 October 2005 to 30 September 2007
“Treating anthrax by selective removal of the protective poly-D-glutamic acid capsule”
Reports: September 2006, August 2007.
Publications:
Summary of Work:
Poly gamma-glutamic acid polymer was purified from Bacillus licheniformis ATCC9945a following incubation at 37oC for 72 hours in medium containing 0.6mM Mn2+. SDS-PAGE
analysis showed this PDGA to have a molecular weight of ~104kDa. CapD depolymerase was expressed in Escherichia coli M15 containing pQEDPE1 plasmid and the enzyme
examined for its capacity to degrade the PDGA in vivo. The enzyme was found to be unstable on storage over periods greater than 24 hours. Bacillus anthracis lytic phage γd’Herelle
was propagated on host Bacillus cereus ATCC 4342 and was used to test for putative capsule degrading activity by incubating with the PDGA. The PDGA was slowly hydrolysed over
a 24 h period. On concentration of the proteins from this phage, SDS-PAGE analysis indicated the presence of putative enzymes of molecular weights putative enzymes of 37, 27,
25, 20 and 15kDa. Current research has also shown presence of putative enzymes from the IM phage of ~35kDa and from FAH phage of 73 and 70kDa. Studies are now focused
on isolation and concentration and further degradative activities of these enzymes from all three γd’Herelle, FAH and IM Bacillus anthracis bacteriophages on PDGA.
Subsequent Post: Intends to take up a post in Community Pharmacy
23
Jones, Dr Matthew (SoP) – 1 October 2006 to 30 September 2008
“Comparison of atomic force microscopy and inverse gas chromatography for the prediction of dry powder inhalation performance”
Reports: July 2007; September 2008.
Publications:
• Jones, M.D., Beezer, A.E. and Buckton, 0. (2008). Determination of outer layer and bulk dehydration kinetics of trehalose dihydrate using atomic force microscopy, gravimetric
vapour sorption and near infrared spectroscopy. Journal of Pharmaceutical Sciences, accepted, in press, p. DOT: 10.1 002/jps .21318.
•
Chiou, H., Tang, P., Chan, H.K., Adi, H., Jones, M.D., Buckton, G., Prud’homme, RH. and Raper, J.A. Production, characterisation and aerosollsation of elongated mannitol
particles.
Summary of Work:
The ability of the cohesive-adhesive balance approach to atomic force microscopy (AFM) and the measurement of Hansen partial solubility parameters by inverse gas
chromatography (ICC) to predict the performance of carrier-based dry powder inhaler (DPI) formulations \vas compared using all possible combinations of five drugs and three
carriers. Comparison of the AFM and IGC results suggested that a weak relationship might exist between these two sets of data, but this was not consistent for different drugs.
Comparison of the AFM and ICC data with the in vitro fine particle delivery of appropriate DPI formulations revealed a previously observed pattern for the AFM measurements, with a
slightly cohesive AFM CAB ratio being associated with the highest fine particle fraction. However, no consistent relationship between formulation performance and the ICC data was
observed. Therefore, the measurement of cohesive-adhesive balances by AFM is the more appropriate technique for the prediction of DPI performance. Other projects demonstrated
that: i) AFM could be used to monitor the surface kinetics of a physical process which correlated with data obtained from bulk measurements; ii) the most appropriate technique for
the calculation of specific free energies of adsorption from ICC data is a little-used method based on topological indices; iii) complex interactions exist between drug concentration,
blending order, mixing time and the in vitro performance of ternary DPI formulations which explain discrepancies between the results of previously published studies.
Subsequent Post:
Senior Medicines Information Pharmacist at the Royal United Hospital in Bath (January 2009)
Eriksson, Dr Andre (KCL) – 1 January 2007 to 31 December 2007
“The role of transporters and metabolism in the disposition of inhaled salbutamol”
Reports: November 2007
Publications:
André Huss Eriksson, Manthena V. S. Varma, Everett J. Perkins and Cheryl L. Zimmerman: The intestinal absorption of a prodrug of the mGlu2/3 receptor agonist LY354740 is
mediated by PEPT1.
Manuscript in preparation for Journal of Pharmaceutical Science.
Manthena V S Varma, André H Eriksson, Everett J Perkins and Cheryl L Zimmerman: Absorptive transport of the mGlu2/3 receptor agonist LY354740 and its prodrug LY544344:
Involvement of uptake and efflux transporters.
Manuscript in preparation for Pharmaceutical Research.
Summary of Work:
The absorption of drugs across biological membranes has traditionally been considered to be by passive diffusion. Recently, attention has also been drawn to the contribution of
active transporters. Drugs delivered orally are absorbed in the intestine and many active transporters in the gastro-intestinal tract have been characterised and identified as implicated
in oral drug delivery. However, very little is known about absorption mechanisms in the lung, the functional significance of transporters that may be expressed and their implications
for local and systemic drug delivery. Culture of epithelial cells is exploited in in vitro characterisation of absorption processes. In vitro studies have shown that the pulmonary
absorption of salbutamol may occur by way of active transporters. The current study aimed to investigate this further using the pulmonary epithelial cell line Calu-3. However, no
involvement of active transporters in the absorption of salbutamol was observed. This discrepancy could be due to a number of factors such as type of cell used, inter-laboratory
variation and culture time and conditions. The characterisation of the pulmonary absorption of salbutamol in particular and the impact of transporters on pulmonary absorption in
general remains an area of importance. However the paucity of consistent models and specific knowledge of the expression levels and patterns in native tissue warrants further
investigations into the field.
Subsequent Post:
Research scientist at the Danish Drug Development Center, Bioneer:FARMA, University of Copenhagen
24
Lam, Dr Jenny Ka Wing (KCL) – 1 October 2007 to 31 August 2009
“Smart polymeric materials; pH responsive polymers as vectors for gene delivery”
Reports: August 2008,
Publications:
Strategic Difference in Plasmid DNA and siRNA Delivery (Review paper, in preparation, 2008)
Summary of Work:
A novel approach to deliver nucleic acids for gene therapy using zwitterionic polymer is introduced here. The major advantage of using zwitterionic polymers instead of cationic
polymers or lipids is their low toxicity. The hypothesis behind the work is that the interaction between zwitterionic polymers and negatively charged nucleic acids can be mediated by
divalent cations such as calcium or magnesium. Preliminary studies indicated that p(DADMAC-co-AA), which is a random copolymer containing quaternary amines and acrylic acids,
demonstrated some degree of in vitro DNA transfection. However other zwitterionic polymers synthesised including p(DMAPMA-co-MA) and p(MAPTAC-co-MA), failed to bind to
DNA in the presence of calcium or magnesium ions. The structure of the zwitterionic polymer needs to be re-designed and this approach for gene delivery is worth further
investigation.
A series of histidine-rich amphipathic peptides was also investigated as vector for nucleic acid delivery. These peptides have pH responsive membrane destabilizing activity. Amongst
all peptides tested in this study, LAH6-X1L which contains the highest number of histidine (6) and lysine (13) residues, showed the greatest DNA and siRNA transfection efficiency.
This result is thought to be due to the high content of histidine which promotes endosomal escape and the high content of lysine that increases the nucleic acid binding efficiency. A
further investigation on the structural-activity relationship on these peptides will help us to gain an insight of the strategic differences in delivering DNA and siRNA.
Subsequent Post: Assistant Professor in Pharmaceutics, University of Hong Kong.
Modarai, Dr Maryam (SoP) – 1 April 2008 to 30 April 2010
“Safety assessment of herbal medicinal products: development of novel procedures based on the new regulatory framework”
Reports: February 2009, March 2010.
Publications:
Safety of herbal medicinal products: Echinacea and selected alkylamides do not induce CYP3A4 mRNA expression.
M. Modarai, E. Silva, A. Suter, M. Heinrich and A. Kortenkamp
th
eCAM ( Epub ahead of print: 2009 (November 11 ) DOI: 10.1093/ecam/nep174)
Metabolomic profiling of liquid Echinacea Medicinal products with in vitro inhibitory effects on Cytochrome P450 3A4 (CYP3A4).
M. Modarai , M. Yang, A. Suter, A. Kortenkamp and M. Heinrich
Planta Medica (Epublication (September 2009 online) ahead of print: DOI: 10.1055/s-0029-1186152).
Herbal extracts used for upper respiratory tract infections. Are there clinically relevant interactions with the cytochrome P450 enzyme system?
M. Heinrich, M. Modarai and A. Kortenkamp
Planta Medica (2008), 74:657-660
Cytochrome P450 inhibitory action of Echinacea preparations differs widely and covaries with alkylamide content.
M. Modarai , J. Gertsch, A. Suter, M. Heinrich and A. Kortenkamp
Journal of Pharmacy and Pharmacology (2007), 59(4):567-573.
Papers in preparation:
A safety platform for assessing interactions between herbal medicinal products and conventional medicines: A safety study on Devils claw .
M.Modarai, A.Suter, Andreas Kortenkamp and Michael Heinrich
The mixture effects of alkylamides – CYP inhibitory activity of Echinacea liquid preparations.
M. Modarai, M. Heinrich and A. Kortenkamp
A metabolomic analysis of Ranunculus species and their association with equine grass sickness.
M. Modarai, J. Michl, S, Edwards and M. Heinrich
25
Summary of Work:
A screening panel for assessing interactions between herbal medicinal products (HMP) and the CYP P450 system was successfully set up and evaluated. It consists of
luminescence-based inhibition assays for CYP1A2, 2C9, 2C19, 2D6 and 3A4 and a reporter gene (luciferase) assay for pregnane-X-receptor activation and CYP3A4 induction. The
resistance of the system to interference was demonstrated by successfully assaying caftaric and chlorogenic acid. These two compounds could not be fully tested with a fluorescence
based assay due to strong fluorescence quenching. This panel was used to evaluate the interaction potential of devil’s claw, by assaying ten commercial preparations. Five
preparations were found to weakly inhibit CYP3A4 (IC 124.2 - 327.6 μg/ml) and five were found to weakly activate PXR (EC 10.21-169.3 μg/ml), but harpagoside (main active
50
50
component) and harpagide showed no inhibition. In addition a Hedera helix (ivy) preparation but not Hederacoside C (main active component), was found to weakly inhibit CYP3A4.
Bergamottin, a natural product known to both inhibit and induce CYP3A4, was shown to inhibit with an IC of 13.63 μM and activate PXR with an EC of 6.7 μM. Overall, the panel
50
50
proved effective at predicting CYP inhibition and induction effects and is suitable for the high throughput screening of HMP prior to marketing.
Subsequent Post:
I will be working as a Visiting fellow, at the Centre for Hepatology, in the Department of Medicine, University College London with Prof. James Owen’s group at the Royal Free
Hospital.
Abbate, Dr Vincenzo (KCL) - 20 October 2008 to 19 October 2010
“Molecularly Imprinted Polymers for Hepcidin Quantification”
Reports: August 2009, August 2010
Publications:
Vincenzo Abbate*, Dareen Jaiash, Sukhi Bansal (2009). Poly (N-isopropylacrylamide) Gel Beads Prepared by Water-in-Oil Suspension Polymerization for the Adsorption of
Fluorescently Tagged Hepcidin. E-polymers, no. 113, 1-9
Vincenzo Abbate*, Nunzianda Frascione and Sukhvinder S. Bansal (2010). Preparation, Characterization and Binding Profile of Molecularly Imprinted Hydrogels for the Peptide
Hepcidin. Journal of Polymer Science Part A: Polymer Chemistry, 48, 1721-1731
Sukhi Bansal, Vincenzo Abbate, Adrian Bomford, John Halket, Iain McDougal, Swee Lay Thein, Robert C. Hider (2010). Quantitation of Hepcidin using Ultra High Pressure
Chromatography and a Linear Ion Trap Spectrometer. Rapid Communications in Mass Spectrometry, 24, 1251-1259
Yun Lan, Bérangère Langlet-Bertin, Vincenzo Abbate, Louic S. Vermeer, Xiaole Kong, Kelly E. Sullivan, Christian Leborgne, Daniel Scherman, Robert C. Hider, Alex F. Drake,
Sukhvinder Bansal, Antoine Kichler and A. James Mason (2010) Incorporation of 2,3-diaminopropionic acid in linear cationic amphipathic peptides produces pH sensitive vectors.
ChemBioChem, 11, 1266-1272
Vincenzo Abbate*, Nunzianda Frascione, Carola Mckenna, Thy Hoang and Sukhvinder S. Bansal (2010). Novel Biohybrid Hydrogels Prepared by One-Pot Photopolymerization of
Chemically Modified Gelatin and Acrylate Co-Monomers. Polymer Chemistry (submitted)
Lan,Y., Abbate,V., Drake, A.F., Bansal, S.S., Mitchell, G.H. & Mason, A.J. Targeting intracellular Plasmodium falciparum with D-enantiomer linear cationic peptides. Antimicrobial
Agents and Chemotherapeutics (manuscript in preparation)
Abbate, V., Iacobucci,V., Kudsiova, L., Chaudhuri, P., Drake, A.F., Bansal, S.S, & Mason, A.J. Rational design of linear cationic peptides containing derivatives of 2,3diaminopropionic acid produces highly effective and non-toxic nucleic acid vectors. Journal of Medicinal Chemistry (manuscript in preparation)
Regpaul, R., Abbate, V., Gaffney, P. R. J., Bansal, S.S and Harvey R. D. Synthesis and biophysical properties of novel lysilphosphatidylglycerol analogues. Journal of Biological
Chemistry (manuscript in preparation)
Presentation at scientific meetings
Vincenzo Abbate*, Dareen Jaiash, Sukhi Bansal (August 2009) - Poster Presentation, “Poly (N-isopropylacrylamide) Gel Beads Prepared by Water-in-Oil Suspension Polymerization
for the Adsorption of Fluorescently Tagged Hepcidin”, 42nd IUPAC World Chemistry Congress, Glasgow, UK
Vincenzo Abbate*, Nunzianda Frascione, Sukhi Bansal (July 2010) – Poster Presentation, “Molecularly imprinted hydrogels for the peptide-hormone hepcidin prepared by the epitope
via water-in-oil suspension polymerization under mild conditions”, Macro2010: 43rd IUPAC World Polymer Congress, Glasgow (UK).
Vincenzo Abbate*, Nunzianda Frascione, Carola Mckenna, Thy Hoang and Sukhvinder S. Bansal (August 2010) – Poster Presentation, “Development of Novel Biohybrid GelatinBased Copolymers with High Peptide Encapsulation Efficiency”, 3rd EuCheMSChemistry Congress, Nürnberg (Germany)
Vincenzo Abbate*, Nunzianda Frascione, and Sukhvinder S. Bansal (August 2010) – Poster Presentation, “Molecularly imprinted hydrogels for the peptide-hormone hepcidin
26
prepared by the epitope approach via water-in-oil suspension polymerization under mild conditions”, EuCheMSChemistry Congress, Nürnberg (Germany)
Summary of Work:
Hepcidin is a peptide hormone which has a major role in maintaining iron homeostasis in mammals. It acts as a negative regulator of iron entry into the systemic circulation from
macrophages and duodenal enterocytes and provides the predominant mechanism for controlling plasma iron levels. Given the complex structure of hepcidin and because it is a
highly conserved peptide, raising antibodies is a difficult task. Therefore, synthetic sorbents towards hepcidin are highly desirable. Moreover, serum levels of hepcidin are generally
very low; it is thus important to develop novel affinity matrices which are able to selectively capture small peptides while excluding larger peptides and proteins during sample
preparation. During the tenure of the Maplethorpe Fellowship, I have been investigating the synthesis of hepcidin analogues and of fluorescent hepcidin, the quantification of human
hepcidin from body fluids using LC-MS/MS, and the development of novel affinity matrices for hepcidin with particular attention to molecularly imprinted polymers in the form of gel
beads prepared via novel suspension polymerization technique. Spherical and relatively uniformly sized gel beads were routinely obtained after optimization of the synthetic
methodology. The polymers were analyzed by Fourier Transform infrared spectroscopy, optical microscopy and scanning electron microscopy. Although the imprinted materials
exhibited higher affinity towards the epitope template (hepcidin N-terminus) than their corresponding blank polymers, the full length target peptide strongly bound to all the beads
tested. However, by using whole fluorescent hepcidin as the print species, the imprinting effect was more pronounced. Moreover, bovine serum albumin did not bind to poly Nisopropylacrylamide-based polymers. Thus, polymeric “sponges” for biomacromolecules with size-exclusion effect were developed, useful for peptide concentration, immobilization
and purification from serum samples. Finally, this research has led to the development of novel biohybrid hydrogels prepared by one-pot photopolymerization of chemically modified
gelatin and acrylate co-monomers, which were shown to bind the fluorescently-labeled hepcidin as a model peptide through electrostatic interactions in an aqueous medium.
Subsequent Post: Postdoctoral Research Associate (King’s College London).
In addition, I have applied for an EPSRC Postdoctoral Fellowship in Cross-Disciplinary Interfaces (application submitted to the Council in August 2010).
Sparapani, Dr Silvia (SoP) – October 2008 to 30 September 2010
“Novel medicinal chemistry approaches to the treatment of human cancers”
Reports: August 2009, September 2010.
Publications:
Two manuscripts are in preparation (for PNAS and Organic & Biomolecular Chemistry).
Rational Design of Acridine-Based Ligands with Selectivity for Human Telomeric Quadruplexes. Sparapani, S.; Haider, S. M.; Doria, F.; Gunaratnam, M.; Neidle, S. J. Am. Chem.
Soc. 2010, 132, 12263-12272.
Bis-guanylhydrazone diimidazo[1,2-a:1,2-c]pyrimidine as a novel and specific G-quadruplex binding motif. Sparapani, S.; Bellini, S.; Gunaratnam, S.; Haider, S.M.; Andreani, A.;
Rambaldi, M.; Locatelli, A.; Morigi, R.; Granaiola, M.; Varoli, L.; Burnelli, S.; Leoni, A.; Neidle, S. Chem. Commun., 2010, 46, 5680-5682.
Poster presentation.‛240th ACS National Meeting & Exposition’, Boston, MA, USA, August 22-26, 2010
Summary of Work:
Besides Watson-Crick duplex structures, DNA can adopt other topologies during replication, transcription, recombination and damage repair processes. Of particular interest are the
higher-order topologies, termed G-quadruplexes, formed by the planar arrangement of four hydrogen bonded guanine bases. These G-rich sequences are found at the ends of
telomeric DNA in eukaryotic chromosomes and have the potential to cause instability at chromosome ends, leading to senescence and apoptopsis in cancer cells. A number of small
molecules are capable of trapping and stabilizing the 3’ single-stranded overhang of telomeric DNA, as these G-quadruplex structures disrupt the binding of telomerase and
associated telomere maintenance. G-quadruplex forming motifs are also found in the promoter regions of many oncogenes (for example in the genes c-myc and c-kit) and they may
be involved in the control of cell growth and proliferation. A number of small molecules, based on planar chromophores, have been examined and shown to be effective stabilisers of
quadruplex DNA and inhibitors of telomerase activity in vitro. Circular dichroism studies have been carried out as a proof of concept experiment, to support preliminary structural
information and have been evaluated for quadruplex binding selectivity by a FRET-based binding assay. Their effects on a panel of cancer cell lines, known to express telomerase,
and a normal fibroblast cell line have also been reported.
Subsequent Post: Looking for new research opportunities.
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Dabkowska, Dr Aleksandra (KCL) 1 October 2009 to 30 September 2011
“Protein-containing phospholipid bilayer models for study of structure and function of membrane receptor proteins.”
Reports: September 2010
Publications:
Couzon, C., D. Banks, J. Jonkman, A. Dabkowska, R. Nutiu, Y. Li, and C. Fradin. (2004) Characterization of the conformational changes of a tripartite molecular beacon. In
Multiphoton Microscopy in the Biomedical Sciences IV (P.T. So, Editor), 184-194.
Hughes, A.V., J.R. Howse, A.P. Dabkowska, R.A.L. Jones, M.J. Lawrence, and S.J. Roser. (2008) Floating lipid bilayers deposited on chemically grafted phosphatidylcholine
surfaces. Langmuir, 24, 1989-1999.
Dabkowska, A.P., G. Fragneto, A.V. Hughes, P.J. Quinn, and M.J. Lawrence. (2009) Specular neutron reflectivity studies of the interaction of cytochrome c with supported
phosphatidylcholine bilayers doped with phosphotidylserine. Langmuir, 25 (7), 4203–4210.
Dabkowska, A.P., A.V. Hughes, R.A. Campbell, D.J. Barlow, P.J. Quinn, and M.J. Lawrence. (2011) The effect of neutral helper lipids on the structure of cationic lipid monolayers
studied by neutron reflection. Journal of the Royal Society Interface. E-published ahead of print. (doi:
Dabkowska, A.P., F. Foglia, M.J. Lawrence, C. Lorenz and S.E. McLain (2011) On the solvation structure of DMSO/water around the phosphatidylcholine head group in solution.
Submitted to Journal of Chemical Physics.
Dabkowska, A.P., R.A. Campbell, A.V. Hughes, D.J. Barlow, P.J. Quinn, and M.J. Lawrence. The effect of helper lipids on the complexation of DNA with cationic lipid monolayers
studied by neutron reflection. Submitted to Journal of the Royal Society Interface.
Dabkowska, A.P., D.J. Barlow, R.D. Barker, A.V. Hughes, F. Campbell, H. Hailes, J. Keddie, and M.J. Lawrence. The formation of a floating supported phospholipid bilayer using
vesicle deposition suitable for the incorporation of membrane proteins. (In preparation)
Dabkowska, A.P., R.D. Barker, J. Lam, A.V. Hughes, D.J. Barlow, and M. J. Lawrence. Floating lipid bilayers as a model for lipoplexes: the calcium-mediated interaction between
zwitterionic phospholipids and DNA and siRNA. (In preparation, intended submission to Langmuir)
Dabkowska, A.P., M. J. Lawrence, D.J. Barlow, P.J. Quinn, R. Green, and A.V. Hughes. The interaction between DNA and zwitterionic lipid monolayers at the air-water interface
studied by neutron reflectivity and external-reflection FTIR. (in preparation, intended submission Langmuir)
Dabkowska, A.P., G. Fragneto, A.V. Hughes, D.J. Barlow, P.J. Quinn, and M.J. Lawrence. The effect of substrate cleaning protocol and lipid unsaturation on the binding of
cytochrome c to single bilayer model membranes studied by neutron reflectivity. (in preparation)
Dabkowska, A.P., A.V. Hughes, D.J. Barlow, P.J. Quinn, and M.J. Lawrence. Specular neutron reflectivity studies of the interaction of cytochrome c with monolayers containing
anionic lipids. (in preparation)
Summary of Work:
Membrane receptor proteins (MRPs) are potential drug targets that require the membrane environment to preserve their function and structure. The reconstitution of high amounts of
purified MRPs into a suitable membrane-like environment is imperative to obtaining information about the target MRP using biophysical techniques. Floating supported bilayers (FSB)
are promising model membranes as they are fluid and can be tailored to mimic a wide range of natural membranes. In order to facilitate the formation of MRP-containing FSB on
silicon support, a novel positively charged acryloyloxy-terminated lipid was synthesized. Using Langmuir-Blodgett-Schaeffer methodology the cationic lipid was deposited onto a selfassembled methylacryloyl-silanized surface and subsequently grafted onto it with radical polymerization. Attenuated total reflectance FTIR and specular neutron reflection (SNR)
techniques were used to characterize the molecular architecture at the surface. The grafted cationic lipids were found to be stable as a monolayer with a ~4 Å head group and a tilted
chain of ~20 Å. Phospholipid vesicles doped with anionic phosphatidylserine were found to deposit on the cationic surface as a single bilayer, with a thickness of ~ 26 Å. E. coli
glycerol-conducting channel protein, selected as the model MRP for incorporation into the FSB, was expressed and purified with its structure intact. Further SNR studies have been
scheduled to determine the structure of these MRP-containing FSBs. The MRP-containing FSB is a promising system for in vitro studies of purified membrane proteins using surface
sensitive techniques.
Subsequent Post:
Brucoli, Dr Federico (SoP) 4 January 2010 to 3 January 2012
“Total synthesis and biological evaluation of Proximicins A, B and C as novel potential antitumor agents.”
Reports: November 2010, January 2012
Publications: Brucoli, F., Natoli, A., Marimuthu, P., Borrello, M.T., Stapleton, P., Gibbons, S., Schätzlein, A. (March 2012) Efficient synthesis and biological evaluation of proximicins
A, B and C. Bioorganic and Medicinal Chemistry, 20(6), 2019-2024
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Abstract highlighting the proximicins anti-proliferative and anti-apoptotic properties was submitted to the American Association for Cancer Research (AACR) meeting for poster
presentation. The conference will be held in Chicago in April 2012.
Summary of Work:
The proximicins A, B and C are a family of antitumor-antibiotics isolated from marine actinomycetes of the genus Verrucosispora. The distinctive features of these compounds are
the structural similarity to the DNA-binding agents distamycin and netropsin, and the presence in their molecular framework of the unusual γ-amino acid, 4-aminofuran-2-carboxylic
acid.
In this project, the proximicins were successfully synthesized and then tested for their biological activity. The characteristic repetitive unit of these molecules, the methyl 4-Bocaminofuran-2-carboxylate, was prepared in three synthetic steps using an optimised copper-catalysed amidation method. The proximicins were evaluated for their antitumor activity
using cellular methods. Proximicin B induced apoptosis in both Hodgkin’s Lymphoma and T-cell leukemia cell lines and proximicin C exhibited significantly high cytotoxicity against
glioblastoma and breast carcinoma cells. The proximicins were also screened against Escherichia coli, Enterococcus faecalis and several strains of methicillin-and multidrugresistant Staphylococcus aureus. Proximicin B showed noteworthy activity against antibiotic-resistant Gram-positive cocci (MRSA). These interesting results will be presented at an
international meeting and published in a medicinal chemistry journal.
Given their interesting molecular framework and their significant anticancer and antimicrobial activity, proximicins represent an attractive synthetic target and could be potentially
developed into either anticancer or antimicrobial drugs of broad utility.
Subsequent Post:
Ali-Boucetta, Dr Hanene (SoP) 1 October 2010 to 31 October 2012
“Engineering carbon nanotubes as multimodal modalities for cancer therapy.”
Reports: August 2011, September 2012
Publications: C. Bussy, H. Ali-Boucetta, K. Kostarelos. Safety considerations for Graphene: Lessons learnt from carbon nanotubes. Accounts of Chemical Research, (2012), in
press. [Acknowledgment to Maplethorpe Fellowship included].
H. Ali-Boucetta , D. Bitounis, R. R. Nair, A. Servant, J.van den Bossche, K. Kostarelos. Purified graphene oxide dispersions do not show in vitro cytotoxicity and in vivo pathogenicity.
Advanced Healthcare Materials, (2012), in press [Acknowledgment to Maplethorpe Fellowship included].
C. Fabbro, H. Ali-Boucetta, T. Da Ros, K. Kostarelos, A. Bianco, M. Prato. Targeting carbon nanotubes against cancer. Chemical Communications (2012), 48, 39113926. [Acknowledgment to Maplethorpe Fellowship included].
H. Ali-Boucetta, A. Nunes, R. Sainz, A. M. Herrero, B. Tian, M. Prato, A. Bianco, K. Kostarelos. Can asbestos-like pathogenicity of long carbon nanotubes be alleviated by chemical
functionalization? Angewandte Chemie International Edition, (2012), Submitted.
D. Bitounis, H. Ali-Boucetta, B. H. Hong, D-H. Min, K. Kostarelos. Prospects and Challenges of Graphene in Biomedical Applications, Advanced Materials, 2012, Submitted.
K.Kostarelos, H. Ali-Boucetta, C. Bussy, H. Meng, T. Xia, A. Nel. A Toxicology and Toxicokinetic Framework for Nanoparticles in Medicine. Nature Biotechnology, 2012, to be
submitted.
H. Ali-Boucetta, K. Kostarelos. Toxicokinetics, excretion, and tissue accumulation of carbon nanotubes. Advanced Drug Delivery Reviews,2012, (invited).
H. Ali-Boucetta, I. Del Rey, E.Venturelli, A.Bianco, K. Kostarelos. The binding and diffusion of CNT-antibody constructs within avascular tumour spheroids, 2012, (in preparation).
H. Ali-Boucetta, L. Newnham, E.Venturelli, I. Del Rey, A. Bianco, M. Robinson, K. Kostarelos. Towards a novel intracellular targeting: Carbon nanotubes- Intrabody Biology and
Pharmacology, 2012, (in preparation).
C. Menard, H. Ali-Boucetta, K. Kostarelos. A. Bianco. Triple Functionalised Carbon Nanotubes for Therapy, Targeting and Imaging, 2012, (in preparation).
Co-editor of special issue on “Carbon Nanotubes in Medicine & Biology" to be published with the journal Advanced Drug Delivery Reviews.
Summary of Work: Functionalised carbon nanotubes were engineered with targeting ligands of different sizes against mucin-1 glycoprotein (MUC1). It was interestingly found that
the size of antibody is the driving force in the cellular translocation of the CNT-Antibody construct inside 3D multicellular tumour spheroids. The latter represents the bridge between
2D monolayer and in vivo tumour model. It was found that if the antibodyis of large enough molecular weight such as full IgG then the binding and the interaction of CNT with the
spheroid become dependent on it; otherwise if the antibody is of a smaller molecular weight (e.g. a fragment like Fab’) then the CNT seem to determine the intratumoral interaction
and trafficking instead. Moreover, the development of new targeting constructs with an intracellular target was achieved using CNT linked to novel intrabodies (INT). Surface plasmon
resonance studies showed that CNT-INTT (with therapeutic activity) have strong binding to its target protein compared to negative control CNT- INTC. The CNT-INT showed clear
cellular uptake and translocation inside cancer cells and signs of therapeutic activity in vivo with the CNT-INTT showing the longest survival of animals. The above discoveries are
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interesting and warrant further research in the use of CNT as a nanovector for therapeutic and targeting ligands for cancer therapy.
Subsequent Post: joining University of Birmingham as a Lecturer in Pharmaceutical Nanoscience.
Benaouda, Dr Faiza (KCL) 1 January 2011 to 31 December 2012
“The treatment of acute respiratory crisis using novel lung-targeting drug ion pairs.”
Reports: October 2011, December 2012
Publications: F. Benaouda, M.B. Brown, S. Ganguly, S.A. Jones, G.P. Martin. Discriminating the molecular identity and function of discreet supramolecular structures in topical
pharmaceutical formulations. 2012. Mol. Pharm. 9 (9), 2505-2512.
F. Benaouda, M.B. Brown, B. Shah, G.P. Martin, S.A. Jones. The influence of self-assembling supramolecular structures on the passive membrane transport of ion-paired
molecules. 2012. Int. J. Pharm. 439, 334-341.
F. Benaouda, M.B. Brown, G.P. Martin and S.A. Jones. Triggered in situ drug supersaturation and hydrophilic matrix self-assembly. Pharm Res. 2012. DOI: 10.1007/s11095-0120838-x
M. Reid, F. Benaouda, R. Khengar, S.A. Jones, M.B. Brown. Topical corticosteroid delivery into human skin using hydrofluoroalkane metered dose aerosol sprays. Int. J. Pharm
(planned submission- Dec 2012).
H.X. Ong, F. Benaouda, D. Traini, D. Cipolla, I. Gonda, B. Forbes and P.M. Young. Inhaled liposomal ciprofloxacin enhances residency time of the antibiotic in the respiratory tract:
in vivo, ex vivo and in vitro Methods Comparison. J. Control Release. (Planned submission Dec 2012).
F. Benaouda, MB. Brown, G.P. Martin and S.A. Jones. Understanding the role of macromolecules to control self-assembly of supersaturated drug suspensions within
supramolecular structured propylene glycol-water vehicles. Mol. Pharm. (Planned submission- Jan 2012)
F. Benaouda, S.A. Jones, G.P. Martin and M.B. Brown. Controlling drug deposition in human skin through in situ drug re-crystallisation in superstructured co-solvents. J. Control
Release. (Planned submission- Jan 2012).
F. Benaouda, C. Page, B. Hider, S. A. Jones, B. Forbes. Novel lung-targeting drug ion-pairs for improved theophylline lung disposition. (In preparation).
F. Benaouda, C. Page, B. Hider, S. A. Jones, B. Forbes. Characterisation of theophylline ion-pairing in binary mixtures. (In preparation).
F. Benaouda, B. Forbes, C.P. Page and S.A. Jones. The characterisation of theophylline ion-pairing in the solution state. PharmSciFair. 103: P-149 (2011).
F. Benaouda, M.B. Brown, G.P. Martin and S.A. Jones. Manipulation of in situ supersaturation kinetics using superstructured hydrophilic matrices. 2011. UK-PharmSci. 12: P-8.
F. Benaouda, G.P. Martin, M.B. Brown, and S.A. Jones. The influence of supramolecular structuring of topical delivery vehicles on drug transmembrane penetration. 2012.
Perspectives in Percutaneous Penetration. La Grande Motte, France.
B. Shah and F. Benaouda. Characterising ion-pair formation in binary solvents. 2012. UKPharmSci, Nottingham, UK.
H. X. Ong, F. Benaouda, D. Traini, D. Cipolla, I. Gonda, B. Forbes and P.M. Young. Absorption profiling of inhaled liposomal ciprofloxacin nanoparticles using isolated perfused lung.
2012. Drug Delivery to the Lung. Edinburgh, UK.
Summary of Work: Modifying the biodistribution of theophylline, a systematically delivered bronchodilator, to improve pulmonary targeting would localise pharmacological activity to
the site of action and reduce systemic toxicity. Polyamine active transporter uptake is one possible means to achieve this. As such, the aim of this project was to investigate the effect
of formulating theophylline as an ion-pair with spermine, a substrate for the polyamine transporters, on the extraction of theophylline from the pulmonary circulation into the lung.
Theophylline ion-pair formation was confirmed in the solution state using high performance liquid chromatography (HPLC) and Fourier transform infrared spectroscopy (FTIR). When
passed through isolated perfused lung, the theophylline-spermine ion-pair showed good stability which resulted in a 2-fold increase in pulmonary tissue uptake compared to
theophylline. In-vivo lung function studies indicated that the theophylline-spermine ion-pair was as effective as theophylline in reducing the histamine-induced bronchoconstriction.
Accordingly, the active polyamine transporter pathway appears to provide a promising strategy to achieve pulmonary targeting for the treatment of acute bronchoconstriction.
Subsequent Post:
Negus, Dr David (UCL SoP) 1 October 2011 to 30 September 2013
“Identification, cloning and characterization of a Poly-γ-D-glutamic acid depolymerase; a novel therapeutic for the treatment of anthrax infections”
Reports: September 2012, August 2013
Publications: Negus D, Burton J, Sweed A, Gryko R, Taylor PW. Poly-γ-D-glutamic acid capsule interferes with lytic infection of Bacillus anthracis by B. anthracis-specific
bacteriophages. Appl. Environ. Microbiol. 2013. 79(2):714-717
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Stabler R, Negus D, Pain A, Taylor PW. Draft genome sequences of Pseudomonas fluorescens BS2 and Pusillimonas noertemannii BS8, soil bacteria that cooperate to degrade the
poly-γ-D-glutamic acid anthrax capsule. Genome Announc. 2013. 1(1):e00057-12
Witcomb L, Dalgakiran F, Johansson M, Negus D, Hansson G, Taylor PW. Repair of gastrointestinal lesions by oral and parenteral administration of recombinant trefoil factor 2.
Manuscript in preparation for Proc. Natl. Acad. Sci. U.S.A.
Negus D, Stabler R, Pain A, Taylor PW. Identification, cloning and characterization of a novel poly-γ-D-glutamic acid depolymerase from Pusillimonas noertemannii BS8. Manuscript
in preparation for Molecular Microbiology
Negus D, Celejewski-Marciniak P, Stabler R, Pain A, Fouet A & Taylor PW. 2013. Isolation and identification of novel capsule depolymerase from Pusillimonas noertemannii BS8
with the capacity to degrade poly-γ-D-glutamic acid. 113th Annual Meeting of the American Society for Microbiology. Denver, Colorado, USA.
Summary of Work: Experimental infections due to bacteria expressing a polysaccharide or polypeptide capsule can be resolved by administration of depolymerases that selectively
hydrolyze the external protective layer. As all isolates of Bacillus anthracis elaborate a poly-γ-D-glutamic acid (PDGA) capsule that is essential for pathogenesis, anthrax represents
an attractive target for this therapeutic approach. We set out to identify a PDGA depolymerase with characteristics suitable for development as a therapeutic for the treatment of
systemic anthrax infections. Using standard soil enrichment techniques we identified a rapidly growing bacterial consortium with the capacity to degrade PDGA as a sole source of
carbon and energy. Analysis of cell lysates by 2D-zymography revealed a single enzymatically active protein of approximately 30 kDa that could not be identified by LC-MS/MS and
peptide matching using publically searchable databases. The genomes of the two isolates were sequenced and the bacteria designated Pseudomonas fluorescens BS2 and
Pusillimonas noertemannii BS8 based on 16 S rRNA homology. Re-analysis of LC-MS/MS data using the newly inferred proteomes identified the depolymerase as a putative
dienelactone hydrolase encoded by P. noertemannii BS8 which we have termed EnvD. Preliminary characterization of recombinant EnvD has established the enzyme is stable,
specific and potent. The characteristics of the depolymerase permit further study in vivo.
Subsequent Post: Currently searching for post-doctoral positions within academia.
Heikal, Lamia (KCL) 1 October 2011 to 30 September 2013
“Is the endogenous tetrahydropbiopterin regulatory system a feasible new therapeutic target for endothelial dysfunction?”
Reports: September 2013
Publications: Lamia Heikal, Anna Starr, Gary P. Martin, Manasi Nandi and Lea Ann Dailey. In Vivo Pharmacological Activity and Biodistribution of SNitrosophytochelatins after
Intravenous and Intranasal Administration in Mice. Nitric oxide. (Planned submission October 2013)
Lamia Heikal, Anna Starr, Dania Hussein, Jesus Prieto-Lloret, Phil Aaronson, Lea Ann Dailey and Manasi Nandi. Restoration of vascular function in spontaneously hypertensive rats
through activation of GCH1- GFRP complex . Circulation research (Planned submission November 2013)
IUPS at Birmingham (21-27 July 2013) Poster title: “Investigating the effects of L-phenylalanine on an endogenous tetrahydrobiopterin regulatory system in spontaneously
hypertensive rats”
Young life scientists’ symposium (YLS 2013) on 6th September 2013 at Queens Mary University , London. Presentation title: “L-Phenylalanine and endogenous regulation of vascular
tetrahydrobiopterin levels”
Summary of Work: Tetrahydrobiopterin (BH4) is an essential cofactor for the synthesis of nitric oxide (NO), a key regulator of the cardiovascular system. Intravenous BH4 improves
vascular function in patients with underlying vascular dysfunction but lacks efficacy following oral administration, possibly due to oxidative inactivation during absorption. Strategies
that can raise BH4 at the level of the endothelium are thus highly desirable. GTPcyclohydrolase
1 (GCH1) performs the rate limiting step of BH4 biosynthesis and this represents a rational target to raise endothelial BH4. L-phenylalanine can activate GCH1 when complexed to an
allosteric regulatory protein (GFRP). We therefore used L-Phe as a tool to investigate its effects in an established model of vascular dysfunction
Spontaneously Hypertensive Rats (SHR). SHR displayed endothelial dysfunction, reduced BH4 and nitrite levels and elevated superoxide levels compared to wild type
(WKY) rats. Oral L-Phe given acutely (100 mg/Kg) or chronically (2% L-phe in drinking water over 10 weeks) restored BH4 and NO levels and reduced superoxide levels back
towards WKY values and restored vascular function ex vivo. This study suggests that small molecules mimicking allosteric changes within the GCH1-GFRP complex induced by LPhe have the potential to improve vascular function through restoration of BH4 levels within the endothelium itself.
Subsequent Post: Postdoctoral Research Fellow in Vascular Biology, Division of Medical Education at Brighton and Sussex Medical school.
Morgan, Anna (KCL), 1 September 2012 to 31 August 2014
“Conventional and non-invasive longitudinal assessments of murine lungs following instillation of nanoparticle suspensions”
Reports: August 2013, August 2014
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Publications: Marie-Christine Jones, Stuart A. Jones, Yanira Riffo-Vasquez, Domenico Spina, Ewelina Hoffman, Anna Morgan, Aateka Patel, Clive Page, Ben Forbes, and Lea Ann
Dailey. Quantitative assessment of nanoparticle surface hydrophobicity and its influence on pulmonary biocompatibility. 2014. Journal of Controlled Release. 183:94-104.
Anna Babin Morgan, Domenico Spina, Yanira Riffo-Vasquez, Aateka Patel, Hanpeng Chen, Stuart Jones, Ben Forbes, Kavitha Sunassee, Stephen Clark, Rafael Torres Martin De
Rosales, Clive Page, and Lea Ann Dailey. Longitudinal assessment of lung health following oropharyngeal administration of poly (vinyl acetate) nanoparticles in mice. In preparation
for 2014. Nanotoxicology.
Aateka Patel, Lea Ann Dailey, Domenico Spina, Yanira Riffo-Vasquez, Arcadia Woods, Anna Babin Morgan, Stuart Jones, Ben Forbes, Kavitha Sunassee, Stephen Clark, Rafael
Torres Martin de Rosales, and Clive Page. The clearance and biodistribution of lipid nanocapsules from the lungs. In preparation for 2014. Nanomedicine.
Arcadia Woods, Domenico Spina, Yanira Riffo-Vasquez, Aateka Patel, Anna Babin Morgan, Helen Collins, Ken Bruce, Rafael Torres Martin de Rosales, Kavitha Sunassee,
Stephen Clark, Lea Ann Dailey, and Ben Forbes. Albumin Nanoparticles as a Pulmonary Drug Delivery Vehicle: Pro-inflammatory Potential, Clearance Kinetics and Biodistribution
of In-111 labelled Albumin Nanoparticles Delivered to the Mouse Lung. In preparation for 2014. Journal of Controlled Release.
Ewelina Hoffmann, Anna Babin Morgan, Stuart A. Jones, Ben Forbes, Clive Page, and Lea Ann Dailey. In vitro investigation of a polymeric nanoparticle-induced ‘foamy’
macrophage phenotype. In preparation for 2014. Toxicology In Vitro.
Abhinav Kumar, Elif Melis Bicer, Anna Babin Morgan, Marco Monopoli, Kenneth Dawson, Lea Ann Dailey, Ben Forbes, and Ian Mudway. The evolution of nanoparticle protein
corona in the human lung. In preparation for 2014. Nature Nanotechnology.
Sackler Pulmonary Institute, Research Seminar Series 2014, London, UK. Presentation title: The Use of Non-Invasive Imaging for Preclinical Pulmonary Models in Drug Development
and Drug Delivery.
American Thoracic Society, Conference May 2015, Denver, CO, USA. Abstract to be submitted for 2014: Influence of Hydrophobicity on Nanoparticle-Induced Lung Injury.
Respiratory Drug Delivery, Conference 2015, Nice, France. Psoter to be submitted: Investigation of Longitudinal Nanotoxicity with a View to Safer Formulation Development.
Drug Delivery to the Lungs, Conference December 2015, UK. Poster to be submitted: Longitudinal Potential for Lung Fibrosis: Assessing Relative Nanotoxicity of Drug Delivery
Vehicles.
Summary of Work: Nanoparticle carriers could improve pulmonary drug delivery, if shown free of toxicity. Whereas it was previously demonstrated that increased nanoparticle
hydrophobicity was associated with acute lung inflammation, in the current study, the long-term effects of nanoparticle hydrophobicity were investigated using invasive or non-invasive
methodologies.
Mice were administered 300 µg low (PVAc60%) or high (PVAc80%) hydrophobicity nanoparticles to the lungs. As positive controls, a separate cohort received crystalline silica
particles or bleomycin-A5 HCl. Animals were sacrificed at days 1, 7, and 28 post-treatment, to assess lung inflammation, injury, and oedema. Non-invasive micro computedtomography (CT) was employed to assess structural changes to the lungs.
Lung inflammation was observed following treatment with PVAc80% nanoparticle suspension, but not PVAc60% (alveolar neutrophils: 1.46 ±0.51 vs 0.25 ±0.05, x105 cells/mL,
respectively). Lung injury and oedema were notable only in the bleomycin group. CT assessment yielded elevated high density (‘water’) signal volumes in the lungs of bleomycintreated animals, characteristic of oedema. Signal volumes for silica-treated mice were minimally elevated compared to naïve mice, and remained unresolved throughout the study.
In contrast, minimally elevated signal volumes in nanoparticle-treated mice resolved with time. Results suggested that CT detected structural changes in addition to oedema, which
were not detected by invasive methods. Naïve mice showed no change in high density signal volumes over time.
This study confirmed that the low hydrophobicity PVAc60% nanoparticle suspension was less toxic than the high hydrophobicity PVAc80% nanoparticle suspension. Furthermore this
study provided support for the use of non-invasive imaging in the assessment of longitudinal changes to the lungs following inhaled treatments.
Subsequent Post: Postdoctoral Research Scientist, Sackler Institute of Pulmonary Pharmacology
Knyght, Ivana (KCL), 1 September 2012 to 30 September 2014
“Novel biosurfactants for pharmaceutical formulation”
Reports: August 2013, October 2014
Publications: Knyght I, Rose RS, Lawrence MJ, Panaretou B Barlow, DJ (2014): Functional expression of dimeric plant defensin SPE10 in Escherichia coli. Biochemical Journal;
m/s submitted
Knyght I, Saaka Y, Clifton L, Lawrence MJ, Barlow DJ (2014) Neutron reflectivity studies of the interaction of θ-defensin and the protegrin with anionic phospholipid monolayers.
Langmuir; m/s submitted
Knyght I, Lawrence MJ, Barlow DJ (2013) Novel peptide biosurfactants for pharmaceutical formulation. Eur Biphys J 42 (Supp 1): S56
Knyght I, Kudsiova L, Zhai C, Parsons R, Lawrence MJ, Panaretou Barlow DJ (2012). Fusion protein expression and insertion of human mitochondrial uncoupling protein 1 in the
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inner membrane of Escherichia coli. Journal of Biological Research 1(1): 12-16
Knyght I, Lawrence MJ, Barlow DJ (2013) Novel peptide biosurfactants for pharmaceutical formulation: poster presentation at the 9th European Biophysics Congress 2013 Lisbon
organised by the European Biophysical Societies Association (EBSA)
Summary of Work: With the aim to develop biosustainable, environmentally friendly surfactants for pharmaceutical and biomedical use, we have successfully designed and
produced two novel peptidyl biosurfactants based on the dimeric plant defensin SPE10 (an antifungal protein naturally occurring in seeds of Pachyrhizus erosus). Molecular modelling
studies were used to guide amino acid mutations in the defensin sequence such that the new disulphide-stabilised peptides would fold so as to possess distinct hydrophobic and
hydrophilic surfaces and thus present as amphiphilic surfactants.
We report the first successful functional expression, purification and refolding of plant defensin SPE10 from inclusion bodies of a well-established expression host, the Gram-negative
bacterium Escherichia coli (E. coli) together with a biosurfactant templated on the defensin peptide backbone. The identity and integrity of the peptide products were verified by mass
spectrometry and their biological activities determined by fungal growth inhibition assays. Encouraging yields of up to 17 mg of pure lyophilised protein per L of bacterial culture were
achieved by a straightforward, quick, versatile and reliable methodology. Preliminary studies of the physical properties of the designed peptides indicate that they have characteristics
consistent with their presentation as surfactants.
Subsequent Post:
Gilmartin, Julia (UCL SoP), 1 December 2013 to 30 November 2015
“Monitored dosage systems in care homes: improving safety or increasing hazard?”
Reports: September 2014, August 2015
Publications: GILMARTIN JF-M, Jani Y, Smith FJ. Exploring the past, present and future of care home medicine management systems: Pharmacists’ perceptions of multicompartment compliance aids. Journal of Pharmaceutical Health Services Research (accepted, 2015)
GILMARTIN JF-M. Informed, tailored and targeted pharmacy support for nurses administering medicines in care homes. Integrated Pharmacy Research and Practice (accepted,
2015)
GILMARTIN JF-M. Using ethnographic approaches to explore care home medicine management systems. Research in social & administrative pharmacy: RSAP (In press, 2015). doi:
http://dx.doi.org/10.1016/j.sapharm.2015.01.005
GILMARTIN JF-M, Raimi-Abraham BT, Jani Y, Smith FJ, Orlu-Gul M. Taking the guesswork out of supplying multi-compartment compliance aids: do pharmacists require further
guidance on medication stability? The International journal of pharmacy practice (epub ahead of print, 2015). doi: 10.1111/ijpp.12183
Edirisinghe S, Raimi-Abraham BT, GILMARTIN JF-M, Orlu-Gul M. Multi-compartment compliance aids (MCAs): Application to the geriatric community. European geriatric medicine
2015; 6(1): 65-68. doi: 10.1016/j.eurger.2014.05.004
GILMARTIN JF-M, Jani Y, Smith FJ. The benefits of using qualitative research methods to understand care home medicine management systems. BMJ open (Proceedings of the
UCL Qualitative Health Research Symposium) 2015; 5(4): A6. doi:10.1136/bmjopen-2015-UCLSymposiumAbstracts.16
Alsaeed D, Davies N, GILMARTIN JF-M, Jamieson E, Kharicha K, Liljas AEM, Raimi-Abraham BT, Aldridge J, Smith FJ, Walters K, Orlu-Gul M. Older people's priorities in health and
social care research and practice: A public engagement workshop. Research Involvement and Engagement (submitted, 2015)
Raimi-Abraham BT, Bordos E; GILMARTIN JF-M, Orlu Gul M. The potential of mobile healthcare technologies to enhance pharmacy medicine services for older persons and chronic
medicine users. International journal of medical informatics (submitted, 2015)
Summary of Work: Multi-compartment compliance aids (MCAs) are used to organise medicines into dosing schedules and are used by UK care home (CH) staff to administer
medicines. Limited literature supports perceptions of increased safety associated with administering medicines from MCAs compared to original medicine packaging (OP). This study
aimed to understand how MCAs and OP affect the way in which medicines are managed in CHs.
Semi-structured interviews with eight pharmacists identified that MCAs were originally introduced into UK CHs to address unsafe medicine administration practices and because of
pharmacy commercial interest. It was predicted that MCAs will continue to be used in the future due to their perceived benefits of improved safety and efficiency, although some
pharmacists recommended removing MCAs and training CH staff to administer medicines from OP.
Medicine administration was observed in 10 Greater London CHs which used MCAs or OP. Up to four days was spent at each CH, observing up to three medicine administration
rounds per day, to achieve a total study sample size of approximately 2,250 observed medicine doses. Medicine administration discrepancies, barriers and facilitators were observed.
Semi-structured interviews with 23 CH staff who administered medicines ascertained perspectives on the advantages and disadvantages of MCAs and OP.
Subsequent Post: Applied for an Australian early career researcher postdoctoral fellowship to refine my pharmacy research and teaching skills. Also exploring how to facilitate
continued collaboration with UCL School of Pharmacy in areas of pharmacy research and teaching.
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Lexmond, Anne (KCL), 1 October 2014 to 30 September 2016
“Is the delivery of adenosine as dry powder the answer to shortcomings in the bronchial challenge test?”
Reports: August 2015
Publications: Lexmond, AJ (2015). Kinderinhalator: Middelhoge weerstand best geaccepteerd. Pharmaceutisch Weekblad, 150(6): 16–17.
Summary of Work:
Subsequent Post:
Smith, Kimberley (UCL SoP), 1 October 2014 to 30 September 2016
“Zebrafish dopamine transporter mutants: disease modelling and drug discovery in infantile parkinson-dystonia”
Reports: August 2015
Publications: Oral presentation at joint UCL and KCL fish meeting. Talk entitled ‘Zebrafish dopamine transporter mutants: disease modelling and drug discovery in infantile onset
parkinsonism-dystonia’. 2015.
Poster presentation at UCL neuroscience symposium. Poster entitled ‘Zebrafish dopamine transporter mutants: disease modelling and drug discovery in infantile onset parkinsonismdystonia’. 2015.
Accepted onto the prestigious and competitive ISN Advanced Summer School on Fitzroy Island, Australia. I was also awarded a scholarship for which to attend. The topic of the
Advanced School is ‘synaptopathies’. I will be giving a poster presentation entitled ‘Dopamine Transporter Deficiency Syndrome: New clinical findings and disease modelling in
zebrafish’. 2015.
Poster presentation at ISN biennial meeting in Cairns Australia. Poster presentation entitled ‘Dopamine Transporter Deficiency Syndrome: New clinical findings and disease modelling
in zebrafish’. 2015.
Smith KM, Meyer E, Kurian M, Harvey RJ (2015) The role of Na+/Cl- dependent SLC6 family transporters in human disease. Invited review in preparation for Trends in Genetics.
J Ng, J Chen, K Erreger, E Meyer, S Mohammed, K Smith, S Barral, JP Linn, NC Øien, D Horn, MR Schweiger, R. Harvey, S Pope, SJ Heales, A Galli, ME Reith, MA Kurian (2015)
Novel mutations in Dopamine transporter deficiency syndrome. In preparation for Neurology
Smith KM, Pernaute B, Rodriguez T, Cobb BS (2015) miRNAs in the role of the canonical stress response and calcium homeostasis. In preparation for Genes and Development.
Pernaute B, Spruce T, Smith KM, Sanchez-Nieto JM, Manzanares M, Cobb B, Rodriguez TA (2014) MicroRNAs control the apoptotic threshold in primed pluripotent stem cells
through regulation of BIM. Genes and Development,28:1873-1878
Smollett KL, Smith K, Kahramanoglou C, Arnvig KB, Buxton RS, Davis EO (2012) Global analysis of the regulon of the transcriptional repressor LexA, a key component of SOS
response in mycobacterium tuberculosis. Journal of Biological Chemistry 287:22004-22014.
Summary of Work:
Subsequent Post:
Herrero Zazo, Maria (KCL), 1 October 2015 to 30 September 2017
“Repurposing of Drugs for Treatment of Psychiatric Conditions”
Reports:
Publications:
Summary of Work:
Subsequent Post:
Gomes Pina, Fatima (UCL SoP), 1 July 2015 to 30 June 2017
Compartamentalised Nanoparticles to Overcome Antibacterial Resistance: A Novel Electrohydrodynamic Manufacturing Approach
Reports: June 2016
Publications: L. Modica de Mohac, M. F. Pina, B. T. Raimi-Abraham (2016) Solid Microcrystalline Dispersion Films as a New Strategy to Improve the Dissolution Rate of Poorly
34
Water Soluble Drugs: A Case Study Using Olanzapine. International Journal of Pharmaceutics. In press
M. F.Pina, K. Lau, K. M. Scherer, M. Parhizkar, M. Edirisinghe, D. Q.M.Craig, The Generation of Compartmentalized Nanoparticles Containing SiRNA and Cisplatin using a MultiNeedle Electrohydrodynamic Approach, Submitted to Small
M. F. Pina, N. F. Kamaruzzaman, L. Good, M. Edirisinghe, D. Q.M. Craig; A Novel Therapeutic Approach for MRSA using Compartmentalised Nanoparticles prepared via
Electrohydrodynamics, poster submitted to UK PharmSci (2016), Glasgow (UK).
M. F. Pina, M. Parhizkar, W. K. Lau, M. Edirisinghe, D. Q.M. Craig; Preparation of Layered Nanoparticles for Synergistic Drug Delivery Applications Using Multi-Needle Coaxial
Electrohydrodynamics, poster for: Pharmaceutics, Biopharmaceutics and Pharmaceutical Technology (2016), Glasgow (UK).
M. F. Pina, W. K. Lau, M. Parhizkar, M. Edirisinghe, D. Q.M. Craig; An Innovative Co-axial Multi-needle Electrohydrodynamic Approach for the Preparation of Compartmentalised
Nanoparticles, poster for University of North Carolina Research Meeting (2015), North Carolina (USA).
Summary of Work:
Subsequent Post:
Brauer, Ruth (UCL SoP), 1 October 2016 to 30 September 2018
“Psychotropic drug safety in pregnancy”
Reports:
Publications:
Summary of Work:
Subsequent Post:
Woods, Arcadia (KCL), 1 October 2016 to 30 September 2018
“From lead candidate molecule to inhalable formulation: Can biocompatible nanoparticles provide a controlled-release platform suitable for delivery of a novel antibiotic compound to
the lung?”
Reports:
Publications:
Summary of Work:
Subsequent Post:
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