Wednesday 21/03/18

On a brilliantly sunny day in Pembroke College, I made my way around (not over) a perfectly mowed lawn towards the grand hall for breakfast. Jumping straight into the spirit of the immersive three-day spring school, I sat next to new faces and introduced myself, encountering a variety of nationalities and research interests within the field of gene and cell therapy. For the first session, I had signed up for the career progression talk however the other sessions included public engagement and presentation workshops. Angela Bradshaw from the University of Glasgow gave an extremely helpful introduction into fellowships, particularly the steps we can take to best prepare ourselves for the competitive, and ever daunting application process.

Pembroke College, Oxford

After lunch we were all warmly welcomed to the 2018 ESGCT & BSGCT Spring School by ESGCT vice-president, Hildegard Büning from Hannover Medical School and BSGCT president, Uta Griesenbach from Imperial College London. To begin the talks, ESGCT president, Robin Ali from UCL gave a history of the ESGCT since its conception in 1992 as well as the field of gene and cell therapy itself. Important breakthroughs in diseases such as X-linked severe combined immunodeficiency were highlighted as important landmarks in the growth and advancement of the field as well as Robin’s own research area, AAV vectors used for inherited retinal conditions.

Next a session focusing on delivery tools. Stuart Nicklin from the University of Glasgow gave an elegant description of adenoviral vectors; their structure and interactions in the body as well as techniques to change vector tropism through capsid modification. Similarly, Hildegard Büning described capsid shuffling in the context of adeno-associated virus (AAV), redirecting AAV vectors away from the liver and towards target receptors. Zoltán Ivics from the Paul Elrich Institute in Langen gave a non-viral perspective using ‘cut and paste’ transposons, specifically Sleeping Beauty transposon, for cell engineering and gene delivery. Continuing on the theme of delivery tools, Vincenzo Cerullo from the University of Helsinki encouraged finding creative solutions; dressing oncolytic adenoviruses in tumour ‘clothing’ to enhance the body’s anti-tumour immune response. To introduce retro/lentiviral vectors, Luigi Naldini from HSR TIGET institute in Milan spoke from his wealth of knowledge and experience in the field, describing improvements in vector design, such as self-inactivating vectors, to enhance vector safety following genome integration.

Moving on to more recent advances, Rafael Yáñez from Royal Holloway in London spoke of life before CRISPR (who would have thought!), introducing other nucleases used in gene editing (meganucleases, zinc-fingers and TALENs) and the advancement of CRISPR-Cas systems which can now make single-strand base changes without introducing double-strand breaks. UCL’s Simon Waddington animatedly described the future challenges faced by scientists in the field of gene and cell therapy followed by some amusing examples of the widely inaccurate portrayal of ‘gene therapy’ by Hollywood. To close the first day, Luigi Naldini walked us through his transition in interest from gene replacement to gene editing of haematopoietic stem cells (HSCs), including instructing novel functions and encoding microRNA target sites for regulated expression.

After a jammed-pack day of fascinating talks, it was time to relax and enjoy dinner. To encourage new introductions and informal chatter, students and experts alike were allocated to house ‘colours’ which denoted different tables. To finish the meal, Oxford’s own Len Seymour spoke of Oxford’s and particularly Pembroke College’s history and accolades, relaying the story of the great Oxford polymath, Robert Hooke, who probably described gravity before Newton but his (Cambridge) rival took all the glory. The moral: don’t trust anyone from Cambridge!

Thursday 22/03/18

On the Thursday the attendees gathered in full force for the 8am session delivered by Jean Davoust from the Necker Hospital in Paris. As he put it, the immune response against vectors like AAV resembles Cerberus, the mythical guard dog. All three heads, representing innate, adaptive humoural and cellular immunity, must be evaded. Strategies such as deliberately targeting the liver which has an important tolerogenic role in eliminating naïve activated CD8 T-cells have shown success in various disease. The theme was continued in the final session of the day when Amit Nathwani from UCL discussed the successful gene therapy trials in Haemophilia A and B. Transient liver inflammation early on in Haemophilia B trials, probably due to hepatocyte destruction by T-cells, was linked to reduced transgene expression later. Patients are now treated with prednisolone during the risk period with encouraging results.

Toni Cathomen from the University of Freiburg then took us through progress to date in gene editing technologies, highlighting the successes of zinc-finger nucleases which have been used clinically since 2001. Although easier to design and more customisable, CRISPR technology still carries the risk of off-target activity which can lead to potentially dangerous deletions and for which we still lack a good readout.

After a much needed coffee break the focus shifted to stem cells. Alessandro Aiuti from the SR TIGET institute in Milan outlined the processes of harvesting HSCs including the relative strengths of obtaining these from umbilical cord blood, bone marrow, and mobilised peripheral blood. He highlighted the cost barrier to using HSCs in treatment of common diseases, suggesting that this will become less of a problem as more centres develop expertise in harvesting, transducing and reengrafting HSCs in addition to large-scale vector production.

Sally Cowley from the University of Oxford discussed the paradigm shift that followed the discovery of the four ‘Yamanaka factors’ sufficient to reverse engineer skin fibroblasts into induced Pluripotent Stem Cells (iPSCs), obviating the need for embryonic stem cells which are difficult and controversial to obtain. She showed astounding timelapse footage of these cells organising into an embryoid body in vitro and explained how they have been used to replace dopaminergic neurons which are absent in Parkinson’s disease as well as many other applications.

After lunch in the great hall we received guidance on public speaking by Paul Charlton outlining many common pitfalls (resist the temptation to use laser pointers!) before being treated to an overview of progression in the field of ocular disease from TIGEM’s Albert Auricchio. There has been huge progress since the first successful transduction of a mouse eye by Robin Ali’s group in 1996 and CRISPR/Cas9 gene editing may now allow us to address dominant inherited diseases by knocking out genes containing toxic gain-of-function mutations.

Nathalie Cartier from MIRCen spoke about gene therapy in neurodegenerative diseases. The two main approaches here are in vivo transduction of neurons using AAV and ex vivo transduction of HSCs using lentiviruses. In the severe form of spinal muscular atrophy severe disability and death usually occur before the age of four but after a single AAV infusion, dramatic rescue of motor function was observed with some children even walking independently.

A similar theme was taken up later in the day by George Dickson from Royal Holloway University who discussed gene therapy for muscular dystrophies. Professor Dickson showed sequential footage of dogs with Duchenne muscular dystrophy that showed huge motor improvements following treatment with AAV vectors containing canine microdystrophin, providing the foundations for a first-in-man clinical trial.

In the final session before the break, Uta Griesenbach explained the challenges of gene therapy for cystic fibrosis where the lining of the lung provides a tantalisingly accessible target for viral vectors but one which is heavily defended by the immune system. Early iterations of the group’s non-viral vector caused flu-like symptoms before design optimisations led to a better tolerated vector. The resulting trial in 2015 raised thorny issues such as whether to involve young children who are theoretically most amenable to gene therapy. This trial heralded the first sustained improvements in lung function following repeat-dosing with gene therapy.

In the final session of the day Sian Harding, also from Imperial, ran through the challenges of AAV gene therapy to ameliorate heart failure. After encouraging safety and efficacy outcomes in the initial CUPID trial, the failure of CUPID2 to meet its endpoints compromised the programme and led to a further trial involving patients with severe heart failure and ventricular assist devices being halted.

That evening experts and early career researchers from across Europe again mingled for dinner and drinks in a ‘meet the expert’ session from which many interesting discussions could be overheard.

Sitting down for dinner in our house ‘colours’ in the grand hall at Pembroke College.

Friday 23/03/18

After two days of enthralling sessions, we started Friday with one of the most important applications of gene therapy at present; recognition and lysis of cancer cells. Waseem Quasim from UCL gave a clear introduction into the well-known CAR-T cell therapies and how the field may be working alongside cell banks and gene editing in the future. Alan Parker from Cardiff University and Len Seymour spoke about an alternative approach; using oncolytic adenoviruses which target and enter cancer cells before replicating and causing cell lysis, eliciting an anti-tumour immune response. Alan Parker explained what the film ‘How to Train Your Dragon’ can teach us about entraining your oncolytic virus to target cancer cells. Len Seymour described the impressive remodeling of the tumour microenvironment by encoding bi-specific monoclonal antibodies within oncolytic viruses to potentiate viral activity by encouraging T-cell recognition of immunosuppressive fibroblasts.

Juan Bueren from CIEMAT in Madrid maintained the excitement of the morning sessions by introducing the use of gene therapy in immunodeficiencies, focusing specifically on bone marrow failure syndromes such as Fanconi anaemia. Both Professor Bueren and Claire Booth from UCL presented impressive translational research which resulted, for instance in the context of ADA-SCID, in the approval of a gene therapy drug called Strimvelis in 2016. Giving a unique perspective, Jean-François Briand from AFM in Paris explained how patient associations can positively impact the progress of gene therapy both economically and by increasing society’s awareness of the importance of studying rare diseases.

After the lunch break, Thierry VandenDriessche from the Free University of Brussels gave a wonderful presentation on the rather complex combination of iPS cells and CRISPR technology to treat non-dominant muscle dystrophies. Changing tack towards vector production, Steven Hyde from the University of Oxford drew on his experience in clinical grade manufacturing to outline the methods for the production of high quality lentiviral vectors and non-viral formulations in lab scale, encouraging us all to manufacture vectors in-house. On a similar note, Steve Howe from GSK further discussed the process of manufacturing vectors in a commercial setting, specifically the bottlenecks associated with transforming gene and cell therapies into medicines. For our final talk, Adrian Thrasher from UCL emphasized the resounding success of gene therapy for immunodeficiencies. His talk focussed on congenital forms of immunodeficiency and the use of lentiviral vectors to transduce haematopoietic cells ex vivo, highlighting the difficulty of transducing the progenitor forms (CD34+ cells).

To end the hugely successful 2018 ESGCT and BSGCT Spring School, Uta Griesenbach and Hildegard Bünning prepared a lovely closure expressing their gratitude to everyone that participated and announcing the winner of the riddle prize which included registration to the ESGCT congress is Lausanne this autumn. Those that remained until the end were rewarded with a guided tour around historical Oxford before heading home for a much needed weekend off. With any luck, some of the relationships formed at the 2018 Spring School in Oxford may lead to international collaborations in the future.

The end of a great three days!

With thanks to co-authors Helena Lund-Palau and Aarash Saleh for their contributions to this blog.