FELLOWS

ESR1: Characterization of protein solution structure using light scattering techniques and SAXS (NZ)

Main supervisor: Senior formulation Scientist Jens Bukrinski, Novozymes A/S.

Protein flexibility and disorder are affected by the physicochemical conditions, and solving and understanding the solution structure under these conditions is the first step towards controlling their behaviour. This PhD project (ESR1) focuses on determination of structure of single domain proteins. The impact of various excipients on the structure will be analysed to find conditions leading to a mono disperse protein solution. The monodispersity of the samples will be ensured by using techniques such as dynamic and static light scattering and asymmetrical flow field flow fractionation-multi angle light scattering and the solution structure will be determined using Small-angle X-ray Scattering. If single crystals are obtained of protein-excipient complexes X-ray crystal structure determination will be used to identify and characterize the molecular interactions. Training will also be gained through extended secondments to Technical University of Denmark, MAXIV and University of Manchester.

DTU Chemistry - PippiSowmya Indrakumar - ESR 2
Project title:
Host:
Main supervisor:


After completing my Bachelor of Science (Research ) degree with major subject as biology and minor in Mathematics, I continued to pursue Master’s degree in Biology from Indian Institute of science, Bangalore. I was a recipient of INSPIRE-DST fellowship throughout my under-graduate studies.

My PhD research focuses on the determination of protein-excipients interaction sites (‘hot spots’) at different physicochemical conditions such as pH and ionic strength using computer simulation techniques. For instance, bioinformatics tools, molecular docking and molecular dynamics simulations will be used to study the effect of excipients on protein stability and to identify excipient hot spot regions on proteins and protein-protein complexes with the aim to correlate these results with experimentally determined structural and stability data generated in the consortium.


DTU Chemistry - PippiSujata Mahapatra - ESR3
Project title:
Host: Novozymes A/S
Main supervisor:


I did my Bachelor and Master of Science in Chemistry at National Institute of Technology, in India. In my Ph.D., I will work on the molecular basis for protein-protein interactions from measurements of static structure factors for proteins that are available at high protein concentrations. I will purify proteins and do the necessary characterization of protein behavior in different pH and ionic strength followed by Small-Angle X-ray Scattering studies for determining the structure factor and thereby the nature of the intermolecular interactions. The experiments will be supplemented by in-silico modeling to elucidate the nature of protein-protein interactions in concentrated solution structure. The primary goal of my research is to determine the intermolecular solution structure of proteins for determining the formulation attributes.

ESR4: Coarse grained modeling of proteins for characterizing structure (LU) 

ESR12: Electrostatic Feature Extraction using Statistical Thermodynamic (LU) 
Main supervisor: Associate professor Mikael Lund, Lund University.
Protein-protein interactions control formulation attributes such as rheology, phase separation, and aggregation.  Interpolating between solution conditions or extrapolating behaviour to concentrated protein solutions requires the use of molecular models and in the two PhD projects we will develop coarse grained (colloidal or semi-colloidal) protein models that can accurately capture protein intermolecular solution structure as determined by i.e. experimental scattering techniques. We will further develop a predictive theoretical tool to identity fundamental protein-protein interaction mechanisms using single protein structural information. This is done by a statistical thermodynamic model, linking molecular features, such as electrostatic anisotropy and shape of single proteins, to intermolecular interactions in solution. The main tools for both projects are statistical thermodynamics and simulations techniques such as Metropolis Monte Carlo and Molecular Dynamics.

ESR5: Experimental characterization of protein-excipient interactions and impact on protein conformation and aggregation using NMR spectroscopy (UoM)
Main supervisor: Dr. Alexander Golovanov, University of Manchester. 
Experimental characterization of protein-excipient interactions and impact on protein conformation and aggregation using NMR spectroscopy. Excipients are additives, which can modulate stability and aggregation properties of protein-based pharmaceuticals. The best excipients are often found by trial-and-error in the course of formulation for the end usage, and currently there is a lack of understanding of the mechanistic principles of their action, and their effect on protein structure, dynamics, transient self-association and aggregation. This PhD project will use advanced high-field high-resolution NMR spectroscopy to look at the structure and dynamic properties of proteins of different sizes (both in unlabelled and isotopically-labelled forms) and develop techniques for characterizing protein-excipient interactions. The data gained from NMR will complement the SAXS, HD-exchange mass spectrometry and studies conducted by other ESRs. The successful candidate will visit both academic and industrial partners (MedImmune) during the project. This will ensure the build-up of a portfolio of interdisciplinary techniques relevant for the applicant’s project and PIPPI objectives.

ESR6: Rational design of aggregation breaking excipients (LMU)
Main supervisor: Professor Gerhard Winter, Ludwig-Maximilians-Universitaet Munchen.
ThisPh.D. project (ESR6)focuses onthe rational design of aggregation-breaking excipients in protein formulations. We want to test a novel concept for designing excipients through modelling of aggregation sites, binding sites and excipient/protein interaction studies for general applicability.Up to now excipients for protein formulations are selected empirically from a few substance classes. The interaction of excipients with proteins is either neglected or studied on a basis that does not consider the excipient binding sites nor the individual molecular sites on the protein where aggregation starts. If one could selectively block protein – protein interaction by adding an excipient binding to that “hot spot”, aggregation could be reduced. This concept was already presented in literature for one specific antibody. The general applicability of the concept is unknown and the right degree of binding or balance between destabilisation through structural interaction and aggregation breaking is unknown. Model proteins and molecular variants with different structures and aggregation behaviour shall be studied. Aggregation hot spots need to be found by data mining and modelling plus characterization via NMR and SAXS. Aggregation-breaking small molecules are then modelled onto the aggregation site. Testing of excipient candidates with all necessary methods in short term aggregation studies shall be carried out and finally the compatibility of excipient candidates needs to be evaluated.

DTU Chemistry - PIPPI - Maria Laura Greco - ESR7Maria Laura Greco - ESR7:
Project title: Effects of excipients on protein behaviour in dilute to concentrated solutions (MEDI) 
Main supervisor: Shahid Uddin, MedImmune.

After receiving her combined BSc ans MSc degree in “Pharmaceutical chemistry and technologies” at the University of Siena, Maria Laura obtained an additional II level master in “Clinical pharmacy” from the University of Milan, Cagliari and Granada. She is also registered at the Pharmacist Society in Italy. Currently she is a candidate PhD student at the University of Manchester, but mainly based in MedImmune (Cambridge, UK) whithin the Formulation team. The research project is focused on the elucidation of the molecular mechanisms responsible of non specific protein protein interactions (nsPPIs) through orthogonal methods: DLS, SLS, SAXS and AUC. In addition, from the development of a novel method for characterizing kD based on Taylor dispersion analysis (TDA) the analyse of solutions containing concentrated excipients will be possible. Final aim, is to develop models for elucidating how nsPPIs depend on excipients and solvent conditions.

ESR14: Characterizing effects of excipients on local protein flexibility and behaviour of partially folded proteins (MEDI) 
Main supervisor: Dr. Shahid Uddin, MedImmune.

About the two posts:
 The two posts will undertake PIPPI project references ESR 7 and ESR 14 in ‘Protein formulation, analytics and delivery’.The projects will focus on monoclonal antibodies (mAbs) and related proteins with novel architectures such as peptide-protein fusions, bispecific antibodies, and self-assembling multimers. Therapeutic mAbs are core to delivering the biologics portfolio of several pharmaceutical companies, including AstraZeneca. MAbs are multidomain proteins structurally comprised of one Fc and two Fab regions connected by a flexible disulphide-bridged hinge. Non-mAb architectures may be more complex and inevitably retain interdomain mobility. Maintaining the physical stability of these proteins at high solution concentrations is paramount but complex and demands rigorous methodology. Research will be conducted in the MedImmune laboratories and have access to state-of-the-art equipment. Through advanced characterization of the protein formulations, the projects will address protein-protein and protein-excipient interactions, delivering new strategies to overcome instabilities such as protein aggregation, phase separation and high solution viscosity.



DTU Chemistry - PippiLorenzo Gentiluomo - ESR 8:
Project title: 
Host: Wyatt Technology
Main supervisor: Dr. Dierk Roessner, Wyatt Technology.



Lorenzo Gentiluomo was born in 1991, studied Chemistry at La Sapienza Univerisity of Rome. During his studies he worked as Formulation Scientist at Henkel and as young Scientist at the University of Jyväskylä. He finished his Master in Organic and Bimolecular chemistry in 2016 with a thesis in Crystal engineering. In 2016 he joined the PIPPI project as PhD studentunder the supervision of Prof. Dr. Frieß at Wyatt Technology Europe.The aim of his project is to develop novel methods, with a focus on light scattering and separation techniques, for assessing aggregation propensity and kinetics of pharmaceutical proteins.Aggregation kinetics (WYT)


DTU Chemistry - PIPPI - Aisling RocheAisling Roche - ESR9:
Project title: 
Host: University of Manchester
Main supervisor:


Aisling is an Irish student currently carrying out her PhD research at University of Manchester, UK. Aisling studied Chemical and Bioprocessing Engineering at University College Dublin, before embarking on the PIPPI project in August 2016. Her project 'ESR 9: Concentrated Solution Rheology' focuses on using multiple rheology and light scattering techniques to assess the viscosity of protein solutions and the role of protein-protein interactions in the increase in viscosity of some protein solutions at high concentrations.

ESR10: Interfacial aggregation kinetics (LMU)
Main supervisor: Professor Dr. Wolfgang Frieß, Ludwig-Maximilians-Universitaet Munchen.
The current PhD project aims at developing an understanding of the role of protein structure, physicochemical properties and formulation characteristics on the aggregate formation as a consequence of exposure of proteins to the air - liquid and packaging material – liquid interface. Aggregation at the interface involves multiple pathways with protein adsorption, dynamic change of the interface by compression and decompression, displacement of protein film from the interface into the bulk and aggregate formation depending on structural change of the protein and protein-protein interaction. Consequently, the protein behaviour at the interface of a series of protein molecules and formulations will be tested which includes fluorescence based methods at the interface, Langmuir-trough-measurements, automated repeatedly cycling Langmuir trough, Langmuir-Schäfer transfer and FTIR-microscopy. Predictive models will be developed which integrate knowledge of all individual steps in the aggregation pathway, unfolding, crowding and stress at the interface as well as protein-protein interactions at the interface and in the bulk.

DTU Chemistry - PippiDillen Augustijn - ESR 11:
Project title: Multivariate data analysis in protein design (UCPH) 
Host:  University of Copenhagen
Main supervisor: Associate professor Åsmund Rinnan, University of Copenhagen.


I’m a Dutch chemometrician and Analytical Scientist educated at the Radboud University Nijmegen. During my education I worked with autofluorescence flow cytometry data of algae. In another project I measured and modelled MALDI-TOF-MS data of polymers. During my master degree I received a COAST MSc+ grant which included over 200 hours of additional teaching in analytical chemistry at various Dutch universities and institutions.

Since August 2016 I’ve been employed at the UCPH. Under supervision of dr. Åsmund Rinnan I will handle the collection, storage and modelling of the data generated within PIPPI.

The first part of my project focuses on gathering the information, standardizing the data input, and storing it. As PIPPI will produce many data, it is vital for the outcome of the project to analyze these data from a multivariate perspective. The main part of my research will therefore be in the field of chemometrics, which is the multivariate data analysis for chemical data. Relational Database Design and Management are a large part of the project and interfacing these with chemometric applications will be one of my aims. I am very much looking forward to collaborating with my fellow ESRs and the companies involved. I personally believe the questions the PIPPI project will answer are very interesting and will aid the development of more stable, safer, and more efficacious medicines.

DTU Chemistry - PippiHristo Svilenov - ESR 13:
Project title: Protein folding stability and storage stability
Host: Ludwig-Maximilians-Universität München
Main supervisor: Professor Gerhard Winter

Hristo Svilenov joined PIPPI in June 2016 as Early Stage Researcher 13. He works on a project focused on the prediction of long-term protein stability.

Briefly, we will use various stability indicating methods (e.g. DSF, chemical denaturation, DLS, SLS) to study how different excipients and formulation conditions affect protein stability. Next, the prediction quality of the different methods will be evaluated by comparison with the outcome of long-term (real-time) stability studies for as much proteins as possible. Finally, we will try to determine general rules when predictivity is high and when it is low. Recommendations for a rationale use of such predictive methods shall conclude the project.

Before joining PIPPI Hristo studied Pharmacy from 2016 to 2012 at the Medical University – Sofia, Bulgaria, where he also worked as Assistant after his graduation. In 2015 Hristo joined Bioskinco GmbH and worked on a project related to cryopreservation and freeze-drying of human cells.

DTU Chemistry - PippiHristo Svilenov - ESR 13:
Project title: Protein folding stability and storage stability
Host: Ludwig-Maximilians-Universität München
Main supervisor: Professor Gerhard Winter

Hristo Svilenov joined PIPPI in June 2016 as Early Stage Researcher 13. He works on a project focused on the prediction of long-term protein stability.

Briefly, we will use various stability indicating methods (e.g. DSF, chemical denaturation, DLS, SLS) to study how different excipients and formulation conditions affect protein stability. Next, the prediction quality of the different methods will be evaluated by comparison with the outcome of long-term (real-time) stability studies for as much proteins as possible. Finally, we will try to determine general rules when predictivity is high and when it is low. Recommendations for a rationale use of such predictive methods shall conclude the project.

Before joining PIPPI Hristo studied Pharmacy from 2016 to 2012 at the Medical University – Sofia, Bulgaria, where he also worked as Assistant after his graduation. In 2015 Hristo joined Bioskinco GmbH and worked on a project related to cryopreservation and freeze-drying of human cells.

DTU Chemistry - Pippi

Alina Kulakova - ESR 15:
Project title: Characterization of solution structure of multi-domain proteins and protein complexes (DTU)
Host: Department of Chemistry, Technical University of Denmark
Main supervisor: Associate professor Pernille Harris, Technical University of Denmark


I had completed my bachelor in Biochemistry and my master in Biochemistry for Health at New University of Lisbon in Portugal. My master thesis consisted in membrane protein production and purification, complemented by crystallization, data collection and structural modelling of several variants of thiosulfate dehydrogenase. 

My PhD project will be focused on structural characterization of multi-domain proteins and protein complexes using primarily Small Angle X-ray Scattering (SAXS) complemented with X-ray crystallography and in-silico modelling tools, that will provide information about protein-protein and protein-excipient interactions. Protein purification and biochemical characterization will be performed in Novozymes A/S, SAXS data collection and analysis in MAXIV and in silico modelling in University of Copenhagen. 

DTU Chemistry - PippiHristo Svilenov - ESR 13:
Project title: Protein folding stability and storage stability
Host: Ludwig-Maximilians-Universität München
Main supervisor: Professor Gerhard Winter

Hristo Svilenov joined PIPPI in June 2016 as Early Stage Researcher 13. He works on a project focused on the prediction of long-term protein stability.

Briefly, we will use various stability indicating methods (e.g. DSF, chemical denaturation, DLS, SLS) to study how different excipients and formulation conditions affect protein stability. Next, the prediction quality of the different methods will be evaluated by comparison with the outcome of long-term (real-time) stability studies for as much proteins as possible. Finally, we will try to determine general rules when predictivity is high and when it is low. Recommendations for a rationale use of such predictive methods shall conclude the project.

Before joining PIPPI Hristo studied Pharmacy from 2016 to 2012 at the Medical University – Sofia, Bulgaria, where he also worked as Assistant after his graduation. In 2015 Hristo joined Bioskinco GmbH and worked on a project related to cryopreservation and freeze-drying of human cells.

DTU Chemistry - PippiHristo Svilenov - ESR 13:
Project title: Protein folding stability and storage stability
Host: Ludwig-Maximilians-Universität München
Main supervisor: Professor Gerhard Winter

Hristo Svilenov joined PIPPI in June 2016 as Early Stage Researcher 13. He works on a project focused on the prediction of long-term protein stability.

Briefly, we will use various stability indicating methods (e.g. DSF, chemical denaturation, DLS, SLS) to study how different excipients and formulation conditions affect protein stability. Next, the prediction quality of the different methods will be evaluated by comparison with the outcome of long-term (real-time) stability studies for as much proteins as possible. Finally, we will try to determine general rules when predictivity is high and when it is low. Recommendations for a rationale use of such predictive methods shall conclude the project.

Before joining PIPPI Hristo studied Pharmacy from 2016 to 2012 at the Medical University – Sofia, Bulgaria, where he also worked as Assistant after his graduation. In 2015 Hristo joined Bioskinco GmbH and worked on a project related to cryopreservation and freeze-drying of human cells.

DTU Chemistry - PippiHristo Svilenov - ESR 13:
Project title: Protein folding stability and storage stability
Host: Ludwig-Maximilians-Universität München
Main supervisor: Professor Gerhard Winter

Hristo Svilenov joined PIPPI in June 2016 as Early Stage Researcher 13. He works on a project focused on the prediction of long-term protein stability.

Briefly, we will use various stability indicating methods (e.g. DSF, chemical denaturation, DLS, SLS) to study how different excipients and formulation conditions affect protein stability. Next, the prediction quality of the different methods will be evaluated by comparison with the outcome of long-term (real-time) stability studies for as much proteins as possible. Finally, we will try to determine general rules when predictivity is high and when it is low. Recommendations for a rationale use of such predictive methods shall conclude the project.

Before joining PIPPI Hristo studied Pharmacy from 2016 to 2012 at the Medical University – Sofia, Bulgaria, where he also worked as Assistant after his graduation. In 2015 Hristo joined Bioskinco GmbH and worked on a project related to cryopreservation and freeze-drying of human cells.

DTU Chemistry - PippiHristo Svilenov - ESR 13:
Project title: Protein folding stability and storage stability
Host: Ludwig-Maximilians-Universität München
Main supervisor: Professor Gerhard Winter

Hristo Svilenov joined PIPPI in June 2016 as Early Stage Researcher 13. He works on a project focused on the prediction of long-term protein stability.

Briefly, we will use various stability indicating methods (e.g. DSF, chemical denaturation, DLS, SLS) to study how different excipients and formulation conditions affect protein stability. Next, the prediction quality of the different methods will be evaluated by comparison with the outcome of long-term (real-time) stability studies for as much proteins as possible. Finally, we will try to determine general rules when predictivity is high and when it is low. Recommendations for a rationale use of such predictive methods shall conclude the project.

Before joining PIPPI Hristo studied Pharmacy from 2016 to 2012 at the Medical University – Sofia, Bulgaria, where he also worked as Assistant after his graduation. In 2015 Hristo joined Bioskinco GmbH and worked on a project related to cryopreservation and freeze-drying of human cells.

DTU Chemistry - PippiHristo Svilenov - ESR 13:
Project title: Protein folding stability and storage stability
Host: Ludwig-Maximilians-Universität München
Main supervisor: Professor Gerhard Winter

 

Hristo Svilenov joined PIPPI in June 2016 as Early Stage Researcher 13. He works on a project focused on the prediction of long-term protein stability.

Briefly, we will use various stability indicating methods (e.g. DSF, chemical denaturation, DLS, SLS) to study how different excipients and formulation conditions affect protein stability. Next, the prediction quality of the different methods will be evaluated by comparison with the outcome of long-term (real-time) stability studies for as much proteins as possible. Finally, we will try to determine general rules when predictivity is high and when it is low. Recommendations for a rationale use of such predictive methods shall conclude the project.

Before joining PIPPI Hristo studied Pharmacy from 2016 to 2012 at the Medical University – Sofia, Bulgaria, where he also worked as Assistant after his graduation. In 2015 Hristo joined Bioskinco GmbH and worked on a project related to cryopreservation and freeze-drying of human cells.

Contact

Pernille Harris
Associate Professor
DTU Chemistry
+45 45 25 20 24

Contact

Mette Hansen
Coordinator, PhD, HR, Economic
DTU Chemistry
+45 45 25 23 82