A little bit about biotechnology and biopharmaceuticals

Biotechnology and the biopharmaceutical industry are a global enterprise, with academic research and industrial manufacturing taking place in much of the developed world. What we mean by biopharmaceuticals are therapeutic compounds produced by living systems, organisms, or enzymes and most commonly, therapeutic proteins produced by cultured animal cells or bacteria. If you know someone who had a heart attack or stroke and received tissue plasminogen activator (t-PA), or someone with breast cancer who was treated with Herceptin, these are examples of biopharmaceuticals. There are currently over 200 approved biopharmaceuticals on the market today and they represent approximately 25% of all new drug approvals. These drugs provide revolutionary, life-saving and life-changing treatments for a variety of conditions including cancer, autoimmune diseases, infectious diseases, and clotting disorders. Despite huge advances since the development of the first recombinant protein drug, human insulin, in the 1970s, many scientific and technical challenges remain as we create new types of therapeutic molecules and attempt to reduce the costs of these drugs to make them available to the developing world.

A little about me and what I do

I am a Professor of Nanobioscience at SUNY Polytechnic Institute, in Albany, New York. The focus of my research program is in mammalian cell biotechnology, primarily the use of cultured mammalian cells to produce therapeutic proteins (primarily antibodies) and carbohydrates (particularly Heparin and Heparin-like molecules). Unlike traditional pharmaceuticals (e.g. aspirin, Tylenol), biopharmaceuticals cannot be easily synthesized in the laboratory. Instead, the cellular machinery of cultured cells is “hijacked” to produce the desired compounds, using the tools of modern molecular biology. While this is conceptually straightforward, there are many fundamental questions about the role of culture conditions and cell physiology on the yield and quality of recombinant proteins produced in mammalian cell cultures. This lack of understanding is exacerbated by the short time period for cell line and process development in the biopharmaceutical industry, leaving little time for fundamental research in process science. Thus, despite the tremendous advances in the techniques available for biological characterization in the last decade, including genome sequencing, discovery-based technologies (e.g. microarrays, proteomics), and silencing RNA, and their application to Chinese Hamster Ovary (CHO) cells (the workhorse of the biotechnology industry), there are tremendous gaps in our understanding of industrially relevant cell lines.

With limited fundamental understanding of how process conditions affect production, the majority of process development is done empirically. Similarly, a lack of understanding of the ideal characteristics for an industrial cell line results in extensive, labor-intensive clone screening to pick the “best clone”. My laboratory has made significant advances in addressing a number of relevant questions such as the effects of on osmolarity recombinant protein production, identification of characteristics of highly productive cell lines, and understanding the role of protein sequence on expression levels. In addition, we have made a number of technological advances in related areas, including metabolic engineering of CHO cells to produce a bioengineered heparin and development of a novel cell-scaffold system for screening glaucoma therapeutics, which one of my former students, Dr. Karen Torrejon, turned into a successful company - Glauconix. In addition to my research program, I have a strong focus on curriculum and course development at the interface between engineering and the life sciences.

Why come to Ireland?

I am in Ireland, supported by a Fulbright Global Scholar Award, collaborating with faculty at the National Institute for Cellular Biotechnology (NICB) located at Dublin City University (DCU) to advance the science of bioproduction of therapeutic proteins. After spending fall in Ireland, the second half of my fellowship will be spending spring (their fall) at the University of Queensland in Brisbane, Australia. The objectives of this research are to improve production of therapeutic proteins by trying to engineer the cells to make proteins more efficiently; identifying characteristics of good producers so it is easier to pick a good productivity cell line out of a bunch of cells (kind of like looking for a needle in a haystack); determining why some therapeutic proteins are easier to make than others.

I selected these institutions for several reasons, the strength of their research programs, their relevance to my research interests (particularly in the areas of proteomics and bispecific antibodies), and for the unique collaboration opportunities they provide. In addition, both institutions have strong training programs and academic-industrial collaborations in biotechnology and mammalian cell bioprocessing. My reason for choosing to visit both these countries in one year is to observe and compare their training programs, with a goal of bringing expertise back to my home institution (SUNY Poly) to develop a training program in Nano-biomanufacturing.

A little more about NICB at DCU and Biotechnology in Ireland

As a result of significant government investment in life-science research and development, combined with a favorable tax policy, Ireland’s biotechnology industry has grown dramatically in the last decade. Nine of the world’s top 10 pharmaceutical companies have facilities in Ireland, with a total of 75 pharmaceutical companies operating in Ireland and 33 US FDA-approved pharmaceutical and biopharmaceutical plants, making Ireland the 7th largest exporter of medicinal and pharmaceutical products worldwide with ~ €39 billion in annual exports.

NICB is a leading research institute located on the DCU campus in Dublin, established in 2000. Building on the emphasis in Ireland on biotechnology and the biopharmaceutical industry, NICB’s mission is to provide targeted and applied solutions to challenges facing the biopharmaceutical industry and translational medicine, based on multi-disciplinary research and clinical and industrial collaboration. To support the growth of the biotechnology industry in Ireland and development of international best practices in biomanufacturing, the National Institute for Bioprocessing Research and Training (NIBRT) was established in 2010. NIBRT is a global center of excellence for training and research in bioprocessing with a new, world class facility in Dublin, replicating a modern bioprocessing plant with state of the art equipment. NIBRT is based on an innovative collaboration between multiple universities in Ireland, including DCU, with funding by the Government of Ireland through IDA Ireland.

Our collaborative research: NICB at DCU has an extensive research program in biopharmaceutical processing, particularly focusing on CHO cells. The recent sequencing of the CHO genome has permitted many novel studies of CHO cell physiology, particularly proteomic studies. For several years, my laboratory has been interested in how cell lines with high productivity differ from cell lines with low productivity. Through a long-standing collaboration with a biopharmaceutical company, we have a unique collection of CHO cell clones, producing the same recombinant monoclonal antibody with varying productivities. We have characterized a number of aspects of the cell physiology including gene copy number, mRNA levels, and protein expression. We observed that cell lines with higher gene copy number exhibited a disproportionate increase in protein expression. We hypothesize that there are changes in transcription factor expression and/or activity in the higher productivity clones. As many transcription factors are activated by phosphorylation, we propose to study these changes using phosphoproteomics, in which the phosphorylated proteins in the cell are identified and quantified to determine their levels of expression. Phosphoproteomic analysis is a challenging technique; however, Dr. Paula Meleady at DCU has extensive expertise in this area, particularly focusing on CHO cells.

Other activities

In addition to the collaborative research, I plan to develop and deliver a short course on the use of mammalian cells for production of non-protein therapeutics NICB, building on our unique research on producing a bioengineered heparin. I also intend to interact with the training facility at NIBRT to observe their facilities and training programs and participate in training activities. Finally, I intend to give guest lectures on my research outside of my host institutions to share my expertise and build collaborations. In Ireland, because of the small size of the country and widespread interest in biotechnology, I will visit several academic institutions and biopharmaceutical companies.

What I do besides work

I have been in Ireland for about a month and half with my entire family. My daughter is going to an all-girls secondary school in Raheny, and my son is taking classes at UCD. My husband is taking a 3 month break from his job at GE. Since being in Ireland, we have taken advantage of the saying that “in Europe, 100 miles is a long distance and in the US, 100 years is a long time” and seen many interesting historical (both ancient and modern) sites. My daughter and I visited the monastery at Glendalough; we went to Malahide Castle; my husband and son went to the General Post Office, and we have lots more to do before we leave. We also had the opportunity to go to the pub and watch the men’s Gaelic football championship (congrats to Dublin), which I am told may be the best (and only) example of Gaelic football I have ever seen. My son celebrated his 18^th birthday with his first legal pint and is looking forward to a visit to the Jameson distillery, and my daughter was introduced to “Taytos” and Chippies (though I think since she does not eat fish, putting chips in a sandwich is a little strange). And we are getting used to the idea that every day you need your raincoat and if you are lucky, your sunglasses as well. Like many folks in Ireland, we are going to spend the midterm break in Spain to get a little more sun.

A little more about Fulbright

Supporting the late Senator J. William Fulbright’s goal of developing international understanding through open communication and cooperative partnerships, the Fulbright program provides bilateral exchange opportunities between the US and 140 countries worldwide. Distinguished participants in the program have become heads of state, judges, ambassadors, cabinet ministers, CEOs, and university presidents, as well as leading journalists, artists, scientists, and teachers. They include 58 Nobel Laureates, 82 Pulitzer Prize winners, 31 MacArthur Fellows, 16 Presidential Medal of Freedom recipients, and thousands of leaders across the private, public, and non-profit sectors. Since its inception in 1946, more than 370,000 “Fulbrighters” have participated in the Program. I am delighted to join their ranks.

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Cancer research at National Institute for Cellular Biotechnology (NICB) aims to understand human cancer and how it might be better detected, treated and monitored. The NICB is fortunate to collaborate with several leading clinical researchers in Ireland including Prof. Michael Moriarty (St Luke’s Oncology group) and Prof. John Crown (St Vincent’s University Hospital) among others.

I am a final year PhD student in the NICB and I am funded by the Irish Cancer Society under the Breast Predict programme. I believe that scientists must engage with the public and provide feedback to them of the progress that is being made in our research field. During my PhD, I have presented at several public events and this year organised a public research seminar for ‘Cancer Week’ to promote the cancer research on-going in the NICB.

NICB staff and students hosted a seminar showcasing some of their ongoing cancer research. The aim of the seminars was to increase awareness of cancer in Ireland and explain to DCU staff and the General Public how the NICB is undertaking studies to further understand and treat the disease. The seminars took place at 2pm on Tuesday 26th September 2017 in the NICB seminar room. All of the speakers invited currently work within the NICB in DCU and we hope to use these events to raise awareness of the research that is being conducted in the NICB with members of the public, patients, and their families.

Neil Conlon (Funded by the Irish Cancer Society)

My talk presented work which highlighted the current treatment options for women with HER2-positive breast cancer in Ireland. I also explained how in certain cases women with HER2-positive develop resistance to their therapy and their cancer spreads.

I highlighted the work I have done, as part of my PhD. thesis, to identify novel treatment options for these patients. I described how I have been able to model treatment resistance by developing laboratory based models of resistance to current therapies. These models have been used to not only examine the resistance mechanisms that emerge, but to identify therapies which can overcome resistance that the cancer can develop to these therapies. I then presented his results which demonstrated several therapeutic strategies which could be used to improve current treatment options and prevent drug resistance from developing.

Nicola Gaynor (Funded by Cancer Clinical Research Trust)

Nicola Gaynor is a final year PhD research student and is supervised by Dr. Denis Collins (see more below), a Principal Investigator in Molecular Therapeutics for Cancer in Ireland (MTCI), funded by Cancer Clinical Research Trust (CCRT).

Nicola’s project focuses on investigating immune checkpoint inhibition in breast cancer. During her presentation, Nicola described how immune checkpoints (which are proteins expressed on the surface of cancer cells) can be inactivated by circulating immune cells. The effect of inactivating these immune checkpoints results in cancer cells being able to evade immune mediated destruction. Nicola presented data which demonstrated how her work is being targeted towards combining immune relevant therapies such as the monoclonal antibody Herceptin with novel checkpoint inhibitors. Her goal is to see if these therapies can work together to provide a better treatment for women with HER2-positive breast cancer.

Dr. Alex Eustace (funded by CCRT, Breast Cancer Now)

Dr. Alex Eustace is a research lead for Molecular Therapeutics for Cancer in Ireland (MTCI) and works closely with Prof John Crown, a consultant oncologist based in St Vincent’s University Hospital.

Alex presented results demonstrating how pre-clinical work conducted in his lab was translated into an early phase clinical trial in Irish women with HER2-positive breast cancer. Alex also mentioned how the clinical trial incorporated a large translational study which enabled clinicians to collect samples including tissue, blood and serum which can be analysed in the lab. Alex further explained that by performing pan-omic profiling, where each patient’s sample can be analysed for mutations, gene changes and proteomic changes, allows us to try to identify biomarkers which can determine which patients are likely to respond to a therapy and which patients unfortunately are likely to relapse.

Dr. Fiona O’Neill

Dr. O’Neill is a Post-Doctoral Scientist supported by St Luke's Radiation Oncology Network and her research focuses on the importance of radiation therapy in the treatment of pancreatic cancer, focusing on the use of conventional as well hypo-fractionated regimes in the treatment of pancreatic tumours.

Hypo-fractionation radio-therapy differs from conventional radiotherapy treatment, as it delivers higher doses of radiation over a much shorter period of time. This method has been shown to increase the therapeutic effectiveness in certain cancer types. Fiona presented results demonstrating the importance of stromal cells and how these cells can protect a patient’s tumour from their radiation treatment. To further understand the role of the local tumour environment in cancer resistance, Fiona is generating radio-resistance in vitro models of pancreatic cancer, which can be used as laboratory models to investigate mechanisms of resistance to radiotherapy. What Fiona and her group are trying to understand is can scientists and clinicians determine a better course of treatment for use in patients with pancreatic cancer using conventional and hypo-fractionated therapy in combination with chemotherapy.

Dr. Sandra Roche

Dr. Roche explained to the audience that pancreatic cancer is associated with very poor survival, and that current chemotherapeutic treatment advances have failed to yield significant clinical improvement.

Here in DCU, in collaboration with SVUH, Sandra has developed Ireland's only panel of patient derived tumour models that may help in understanding the biology of the tumour as well as being more suitable and relevant models in which to test to combinations of drugs. Sandra hopes that these models can help us in two ways: to understand why the pancreatic tumour is so aggressive (using tools in the lab to tell potentially what genes and what proteins may be linked with tumour aggressiveness) and secondly to help work out if it is possible to improve the delivery of the therapeutic agents and novel drugs to the cancer itself. Pancreatic tumours are made up of many different types of cells. In collaboration with University at Buffalo (NY, USA), Sandra hopes to break down some of the other components of the tumour so that the drugs get better access to the tumour cells. If the drug can get to the cells it has a better chance of killing the cancer cells. In this case, we would be using these models as surrogates for patients.

Dr. Denis Collins

Dr. Collins is a Principal Investigator funded by Clinical Cancer Research Trust and works for the Cancer Bio-therapeutics group as part of MTCI.

Denis’ work focuses on how, in recent times, the immune response to cancer has become an area of major interest as a treatment strategy. Denis presented results showing how novel treatment strategies using new anti-cancer drugs called Immune Checkpoint Inhibitors (ICIs) are being investigated in the clinic and explained to the audience how these ICIs prevent immune cells (that can potentially kill cancer) from being turned off by the cancer cells they are targeting. In Denis’ lab, the Cancer Bio-therapeutics group use breast cancer cells grown in the laboratory along with white blood cells and plasma from patients and healthy volunteers to investigate the white blood cell reaction to antibody therapies like Herceptin. The goal of Denis’ research is to discover the combinations of existing drugs and new immune checkpoint inhibitors that gives the best anti-cancer immune response in breast cancer patients.

The Cancer Week Research Showcase offered the public a close view of the cutting edge research that is taking place in the NICB.  Whilst all the presenters acknowledged that advanced cancer is hard to treat, they all presented novel ways in which their research groups can work to further understanding the disease, and potentially finding better ways to treat it.

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The world population is projected to increase to 9.7bn by 2050, placing increasing demands on food production. To meet this requirement, agricultural and animal health industries need to develop innovative solutions for sustainable food production, while considering animal wellbeing, energy and resource limitations. Trace minerals such as copper, iron, selenium and zinc are essential in the diet but both deficiency and over exposure can both result in deleterious consequences and can be detrimental to health (for example immune function impairment). Much has yet to be learned regarding uptake and processing these minerals, especially at the primary site of absorption: the intestinal epithelium.

To address this challenge as an opportunity, the National Institute for Cellular Biotechnology and Alltech Ireland have formed a partnership to develop models which allow us to examine how intestinal cells process trace minerals. Have a look at the video below for more information:

The NICB-Alltech partnership proposal was initiated with the primary goal to achieve a greater understanding of the regulation of mineral levels, the impact of manipulating these levels in mammalian tissues and whether this is influenced by the type of mineral, a study referred to as nutrigenomics. Our mission is to develop and optimise a number of in vitro cell models which track changes related to micronutrient exposure, for minerals in isolation and combination, incorporating genetic and proteomic analysis following uptake and transport of the micronutrients. Current focus is on the impact of inorganic and organic trace minerals on cellular health and function.

Key members of the project are Professor Martin Clynes (head of the DCU research team), Dr. Richard Murphy, the Director of Research at Alltech and Associate Research Director, Dr. Karina Horgan. The DCU team includes Dr. Finbarr O’Sullivan (Associate Director of the NICB) and Dr. Joanne Keenan (Research Fellow), and two PhD students, Ali Coyle and Charles O’Doherty (author). Other members of the NICB-Alltech team include Dr. Niall Barron, Dr. Paula Meleady, Dr. Padraig Doolan and Mr. Michael Henry.

In his role as research director Dr. Murphy is currently involved in multiple areas including peptide bio-marker detection, nutrigenomics, antimicrobial resistance and regulation of protein production. Dr. Horgan is currently responsible for Alltech Life Sciences research in Europe and as part of this she has formed a cellular biology research group focused on developing cellular-based bioactivity assays and profiling yeast products.

My research background is in cell biology, having completed a four year biotechnology degree in DCU in 2011 and working in various roles in the pharmaceutical and orthopaedic industries in the years prior to securing my PhD studentship. While I enjoyed my experience in these positions, I knew my academic life was not over yet.

I began searching for research positions combining my primary interests: cell culture and nutrition. I was fortunate enough to find and secure one in both these interests, at the NICB in DCU. I am currently doing a Ph.D. along with my colleague Ali Coyle (supported by Alltech Ltd. and Enterprise Ireland) under the mentorship of Dr. Keenan and Dr. O’Sullivan.

Left: Members of DCU team; Ali Coyle, Charles O’Doherty, Dr. Finbarr O’Sullivan and Dr Joanne Keenan
Right: Director of Research for Alltech, Dr Richard Murphy presenting at the business matchmaker event on 11th May in the Helix, DCU

Cellular conditions within the small intestine are critical for health and performance in humans and animals, due to its crucial role in controlling uptake of essential minerals and nutrients and protecting the body from pathogenic organisms. This is evidenced by genetic mutations related to intestinal nutrient uptake such as the debilitating disorders: Menkes disease for copper and hemochromatosis for iron. Our team use a multi-targeted approach to assess interactions between micronutrients and the intestinal epithelium, such as mineral micronutrient competition, toxicity, transport and uptake.

When conducting in vitro intestinal studies, Caco-2 cells are differentiated over a 21-day period on permeable supports to generate enterocyte-like cells that mimic the small intestine. Confirmation of differentiation is established by Western blotting for markers of mature enterocytes (e.g. Sucrase isomaltase) and confocal microscopy establishing the appropriate localisation of structural proteins (e.g F-actin, ZO1) as well as measuring trans-epithelial electrical resistance (TEER).

Schematic diagram and photo of Transwell® insert setup

Following optimisation of the models, differentiated cells are studied by altering their environment, such as exposure to different forms and combinations of inorganic and organic minerals, generating a knowledge-base on the effect of micronutrients on small intestine-like tissue. The tools employed to complete these analyses include microarray analysis for gene expression, mass spectrometry for global proteomic expression analysis, confocal microscopy to study protein localisation and ICP-MS for micronutrient content analysis. Complex bioinformatics tools are used to interpret large scale data sets. The combination of these techniques helps to dissect the intricate effects micronutrients have on the cells and the pathways involved in micronutrient exposure and how best to optimise these interactions.

Capturing images of cell cultures using microscope with Olympus DP70 camera

In order to identify the key issues facing the agricultural industry the Alltech Global Alliance Conference took place on 23rd and 24th March 2015, in Beijing, China. Dr. O’Sullivan and Prof. Clynes presented research for the NICB-Alltech team at the conference.

Group photo of the attendees at the Alltech Global Alliance in 2015

The event saw international academic and industry research groups come together to discuss the latest advancements and issues concerning the agricultural industry. One of the key pending developments highlighted during this conference was “gut health management tools and disease models for defining immunity and productivity”. Following this meeting collaborations were organised between research teams to strategically tackle these issues.

The original proposal between the NICB and Alltech signified an important opportunity to engage a growing multinational with Irish academia. Dr Richard Murphy acknowledged that the level of expertise and research facilities at Dublin City University (DCU) were the primary factor in developing the collaboration, following the original partnership. To recognize the group’s efforts, DCU's President Professor Brian MacCraith presented our team with an award for the partnership with Alltech Ireland at the Invent Commercialisation Awards ceremony on 21st April 2016.

The author with Dr Joanne Keenan receiving the Invent Commercialisation award from DCU President Professor Brian MacCraith at the Invent Commercialisation Awards ceremony on 21st April 2016

The agricultural industry will need to overcome great difficulties in the coming years, as constraints on resources increase alongside population growth. To reach the aim of sustainable food supply within these limitations, scientific advancements surrounding nutrient delivery to both animals and humans is essential.

The Alltech-team at the NICB is comprised of post-graduate and post-doctoral researchers led by Professor Martin Clynes and through engagement with a leading industry partner we are using a range of cell manipulation techniques to decipher and optimise micronutrient supply to animals and humans.

Charles O’Doherty

PhD student at the NICB, DCU

NICB-Alltech Alliance group

This industry-academic collaboration is supported by both Alltech LTD. and Enterprise Ireland’s Innovation Partnership Scheme, as part of the National strategy for Science, Technology and Innovation.

The NICB team investigate human and animal cells and tissues at the molecular level in order to uncover biological mechanisms and develop new treatments and diagnostics for a variety of human diseases with particular emphasis on cancer, diabetes, infectious diseases, ocular diseases and nutrient uptake.

Alltech is a global leader in the animal health and nutrition industry and one of the top animal health companies in the world. As part of Alltech’s research program a collaborative research initiative was formed with NICB to investigate the role of micronutrient products in cellular health. ONE 17: Alltech Ideas Conference was completed recently and the highlights can be seen in this video.

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This post is about my career and experiences as a research postdoc, followed by my decision to pursue the dream of running my own business and how that worked out. If you want the short version – just watch the video below!

In this post, I’m going to try and give a quick overview of the last 5 years; so it’s going to be necessarily brief on individual events! I’m hoping to follow this up with a series of smaller, individual posts, discussing in more detail some of the issues I experienced and which should hopefully provide more context for this post.

Every startup journey is different and I don’t think mine is any more representative than anyone else’s. That said, I hope this post is instructive and maybe (if I’m lucky) this will be a fun read for anyone thinking of taking the same leap.

My Background in Research: Cancer, Biopharma and a host of others

In June 1996, I had just finished a 4-year degree program in biotechnology and I was looking at opportunities to do a PhD. A senior postdoc who was interviewing me gave me some great advice: “Don’t do one”. Given that I went ahead and completed a PhD. (with her as my supervisor), why do I think that was good advice today? Because the kind of single-minded attitude that accepts advice like that and thinks (“I’m gonna do one anyway”) has just the type of bloody-minded determination to get them through the tough times that every PhD. student experiences.

I think that advice is good to give to budding entrepreneurs as well: “don’t do it”, because it’s more difficult than probably anything else you’ll ever do; it’s fantastic and heart-breaking and exciting and lonely, all in equal measure, and the ones that hear that advice and plough ahead regardless are the ones who are more likely to succeed.

I initially started out as a cancer researcher at the National Institute for Cellular Biotechnology (NICB), located in Dublin City University (DCU). However, over several years, I moved away from cancer and branched more into biopharmaceutical research, particularly working in collaboration with Wyeth Biopharma (now Pfizer), where we focused on improving their cell line development processes.

Biopharmaceuticals are drugs that are manufactured from living cells that are grown in large vats – the majority of cells used are from the Ovary of a Chinese Hamster isolated in the 1950s (no joke) and improving the productivity of this “CHO” cell is the overall goal.

Biopharma has a large presence in Ireland, with companies like Pfizer, Eli Lilly, Biomarin and Sanofi Genzyme all having extensive operations. The sector is vital for the Irish economy, making up ~50% of Irish exports per annum and is expected to grow to $386.9 billion by the end of 2019, making it the fastest growing sector in Ireland [1].

We conducted this research principally as part of an 8-year research program with Pfizer (initially Wyeth Biopharma), which was funded to the tune of over €6M by Science Foundation Ireland (SFI) and involved Pfizer’s Bio-Manufacturing Sciences Group at Grange Castle, Clondalkin and their Bioprocess R&D Group, located in Andover, an hour’s drive north of Boston, USA.

One of the most interesting bioinformatics projects I worked on for Pfizer was developing a predictive model of productivity for their CHO cell lines, which we published in 2011 in the academic Journal of Biotechnology ("Predicting cell-specific productivity from CHO gene expression"). This was what we could call a “dimension reduction” problem – where you have lots of data points (in this case, gene expression data) that you’re trying to link to a single parameter (in this case “productivity”). Using a combination of the microarrays and our bioinformatics techniques, we could predict how productive a cell line was going to be to within 4 pg units (out of a typical range of 1-50 pg/cell/day).

This was early-stage stuff; we didn’t see this translated into being used for selecting individual cells for cell line development, but it did provide some proof of an important concept - that we could predict, at an early point, which individual cells were likely to exhibit the desired characteristics for a good-quality bioprocess – the emphasis being that these should be the ones to be taken further to eventually deliver a product that would be used as a medical product.

 Stage 2: Escaping the Ivory Tower!

This “predictive-productivity” study got me thinking; what other possibilities could this type of “big data-small predictions” be used for? (if  a picture of Brad Pitt doesn't keep you interested, I don't know what will....)

My combined interests of sports and statistics led me inexorably to books like “Soccernomics”, “The Numbers Game” and the peerless “Moneyball” and from that to my own “eureka!” moment (for my more sensitive readers, no baths were involved…) - I could use my bioinformatics skills to build a predictive model for football players, by using player performance information already collected by sports data firms.

Thus was born the “Beat Your Manager! Fantasy Football” game, which I pitched (in early 2012) to Enterprise Ireland, who were running a new Entrepreneur Development programme (called New Frontiers), in tandem with DCU’s Technology Transfer office Invent and Dundalk Institute of Technology. They loved the idea and accepted me onto the program, beginning in March 2012.

It was under New Frontiers that I had my 2^nd major insight: by tracking user activity, the game would be the first product to deliver real-time consumer information on what fans are thinking about the live game (or “fanalytics”).

With this, I felt I had a new business model; instead of trying to make money from game users, I would sell their behavioral data (most popular lineups, substitutions, formations, etc.) to key stakeholders in the sports industry; including broadcasters, betting companies, clubs/franchises, sporting associations, sportswear companies and drinks companies.

When I started New Frontiers, I had to leave my job to work on the business full-time. During this stage (and until the course finished in Feb. 2013), my main business location was in Invent's startup incubator space. As a combination, I can’t speak highly enough of E.I., the New Frontiers program or Invent. On a simple level, just being given the time and space to develop your business idea and your confidence in a new endeavor was critical. Making a jump like this is so daunting; having sound supports, good advice and realistic criticism is vital to making your dreams a reality.

I started my company (“Beat Your Manager! Fantasy Games Ltd.”) in Sept.2012 and successfully received Feasibility funding from Fingal County Enterprise Board (CEB) to develop the scoring algorithms and a Minimum Viable Product (MVP), which included a “Fanalytics Dashboard” for analysing fan data. I hired the excellent software application & development company NearForm to develop the game as a web-hosted smartphone app which we launched for testing.

On the back of the MVP test, I successfully achieved €50,000 Competitive Start seed funding from Enterprise Ireland; this put me squarely on E.I. commercialisation route as a pre-HPSU (High Potential Start Up).

This allowed me (with NearForm again doing the coding) to develop and launch an improved version of the Game (ver1.0) for iPhone, Android and web, as well as an updated Dashboard for the following 2013/4 English Premier League season.

With assistance from the games marketing company ScragglyDogGames, the launch was well-covered in the regular media, with very supportive reviews from Silicon Republic, RTE News and The Irish Examiner, as well as a host of games-related media outlets.

With the Game operational, I expanded the userbase to over 15,000 registered players and attracted the national radio station NewstalkFM as a broadcast partner, as well as 14 fan websites (including EPL Index, Chelsea in America and LFC Daytrippers), with a combined total of 1 million (monthly) website visitors.

Additionally, former Ireland/USA Rugby coach Eddie O’Sullivan joined my company’s advisory board and I also recruited a full-time Chief Technical Officer (CTO) to oversee product development and improve the company website (which was initially built by a family friend). I also received additional mentorship from Dublin Business Innovation Centre (DBIC) under their “Client Assistance Program for Early-Stage Business Startups and Developing Businesses”:

The Game was also a runner-up in the 2013 AIB Jumpstart competition, Vodafone Startup Awards 2013 and The Irish Times 2013 FUSION program. In Dec. 2013, the game was also profiled on George Hook’s “Kickstart Your Business” radio show – you can listen to a recording of the interview here:

Stage 3: The Road Back

To cut a very long story short; after that, not a lot happened for a long time. I struggled to find a paying customer and without a defined income, I didn’t feel I could reasonably approach VCs for investment. I operated the game until the end of May 2014 and then had to shut it down. I continued to seek commercial clients, as well as contracts for other sports events (the game format works for any team-based sport) and I kept in touch with a few of the venture capital funds that had expressed an interest, but in the absence of a compelling source of revenue, I didn’t feel ready to take the business to a stage where I was willing to ask for investment.

While I was pursuing my startup business, I also completed a Graduate Certificate in Digital Marketing with DCU Business School, my first formal business qualification. I had additionally received lots of training during the 15months of the New Frontiers program (topics covered included: Building an Investible Team, Business Models, Business Planning, Business Valuation, Competitor Analysis, Customer Acquisition Techniques, Customer Development, Financial Basics and Key Metrics, Intellectual property, Legal Issues, Market Research, Market Validation, Negotiating, Presentation Skills, Pricing, Product Design & Development, Marketing Plans, Route to market, Sales Leads Generation and Sales Process).

My new qualification and training (as well as the experience of trying to get a startup business off the ground) gave me a very different perspective on research – I now started to look at science much more from a commercial standpoint; examining its utility and I became interested in the best way to translate basic research into usuable commercial products.

By January 2015, I had returned to the NICB part-time as a senior researcher, with a renewed focus on research projects and industrial collaborations that had a specific commercial element. I felt that over the previous 2.5 years, I had developed immeasurably as a scientist and had acquired business insights that I would have been unlikely to have attained if I had just progressed on a standard postdoc career path.

The first big change I helped implement was the redevelopment of the NICB’s website, incorporating a blog (that you’re reading right now) and also the creation of a range of social media assets (Twitter, Youtube, Linkedin, Google Plus, Flickr) which are aimed to help spread the message of the NICB’s activities, mission and values.

The projects I am engaged on have also tended to be more commercially-focused: for example we are developing a new Conditioned Medium (CM)-based growth supplement for cloning of CHO cells for production of biopharmaceuticals – I can’t think of a better example of coming full circle!

I’m also working on promoting sales of NICB’s flagship hybridoma media product Briclone as well as developing a serum-free version, so as to access customers/markets that demand animal-free products.

I’m also commercialising NICB’s really exciting microRNA technology DECOY-7, which has application in improving upstream cell line development for biopharmaceutical production.

So, it’s all going well right now – even my startup has had a happy ending, as my game was recently (end Jan. 2017) licensed by the sports marketing company Sportego, who will operate the product (with a commercial partner) as a fantasy game for the 2017/8 English Premier League season.

Sportego are a Kilkenny-based fan engagement startup that specialises in sports analytics and social media activation for clubs, fans and sponsors with a client base that includes the SSE Airtricity League, British & Irish Lions, Bournemouth FC and West Bromwich Albion FC. They were previously profiled in Silicon Republic, SportTechie and StartUs magazine and their prospects have undergone a huge upward trajectory, as more recent reviews in The Irish Independent, Silicon Republic and Sport for Business demonstrate.

I’m delighted to be working with them and I hope that there’s a bright future for the game yet – watch this space!

When I think back over the last 5 years of my life and career, I can't but be reminded of  the T. S. Eliot line: "We shall not cease from exploration, and the end of all our exploring will be to arrive where we started and know the place for the first time".

And maybe I don't really agree with the advice I gave in the beginning of this post; if you have a business idea, even if its not in the area you usually work, and you really feel passionately about it; DO try to make it happen. The journey will be long and challenging, there'll be times you'll feel on top of the world and others when you'll wish you'd never taken that first step. The future will be risky and the road unclear.

But, most likely, the experience will be worth the sacrifice and you'll never be the same person again.

Update: This blog post was profiled in an article in the Irish Times Innovation section on Apr. 27th 2017

The post From Biopharma…to Fantasy Football…and back again appeared first on National Institute for Cellular Biotechnology.

First, let’s go back in time……

The 15 year human genome project that cost a massive €3 billion has been one of the greatest milestones of the century, unveiling genetic information that can now be used for personalised medicines and targeted therapies in the clinic. Advances over the last decade in sequencing technology has ushered in the next generation of sequencing platforms such as Illumina, making the cost of sequencing an entire genome go from ~€3 billion to ~€1,000. This has widespread implications for academic research groups, in that this once expensive technology can now be accessed for an array of projects in every corner of academia. The accessibility of whole genome sequencing has now gone beyond the release of the 1,000 genomes project and has moved onto the ambitious 100,000 genomes project.

This is a large-scale study which seeks to sequence the genome of ~70,000 patients in the National Health Service and identify novel recurrent genetic abnormalities associated with a plethora of diseases which could be used to both diagnose and treat.

Image sourced from the National Human Genome Research Institute

Biopharmaceutical drug production

A primary focus of research here at the National Institute for Cellular Biotechnology is in the area of biopharmaceutical production of recombinant therapeutic proteins. The Chinese hamster ovary (CHO) cell is the primary mammalian cell line used in the biopharmaceutical industry for the production of recombinant therapeutic proteins. This cell line is heralded as the “work horse” of biopharma due to its capability to produce high quality biologics with post-translational modifications (PTMs) similar to humans which ensure a fully functional and bioactive protein drug for the treatment of various diseases such as cancer and rheumatoid arthritis.

The various parental CHO cell lines such as CHO-K1 or CHO-S used today were originally isolated in 1957 from the ovary of a Chinese hamster. Today, over 70% of recombinant therapeutic monoclonal antibodies are produced in these cell lines with a long and rigorous cell line development process of ~12 months required to generate a single clone that maintains and exhibits desirable bioprocess characteristics such as fast growing, reaching high cell density, surviving long in culture and producing high amounts of protein per cell.

Unstable genetics – A double edged sword

By nature and owing to their genetic plasticity as a desirable feature for genetic engineering, CHO cells are genetically unstable. This genetic instability was highlighted in a previous study by Lewis and colleagues which revealed there to be more than 4 million mutations across 6 CHO cell lines and 3 lineages in addition to sequence variation that arose during the cell line development process alone. Understanding the genomic landscape of CHO has given insights into genetic variation in genes associated with bioprocess relevant phenotypes such as apoptosis and cell growth. By better understanding these genetic variations that could impact on CHO cell behaviour within the bioprocess, a better selection criteria could be adopted that would see the selection of genetically predisposed CHO cells that perform better in the bioreactor. On top of this, the unpredictable genetic heterogeneity that can arise within multiple CHO cell clones derived from the same parental line means that mutations can occur throughout the cell line development (CLD) process that often results in producer cell lines with sub-optimal performance characteristics.

All of the above means that the process of developing a CHO cell line to reliably produce a therapeutic product that meets the stringent safety regulations of regional drug authorities such as the Food and Drug Administration (FDA) or the European Medicines Agency (EMA) is a very costly and time-consuming one. At the end of this process, the results may still not be desirable. Although it is common to achieve 2-5 g/L production yields in the bioprocess, the high-throughput development of new and sophisticated therapies means that the already existing production cells lines may not produce these therapeutic proteins as efficiently. The CHO group at the NICB harnesses the power of genetic engineering to modify CHO cell lines using a variety of molecular tools such as CRISPR-Cas, microRNAs (DECOY-7) and inducible expression systems to enhance their performance within the bioprocess and ultimately boost drug production.

Multiple Ploidy Disorder

Paramount to the efficiency of production CHO cell lines is the presence of a balanced diet. That is, the perfect amount of critical nutrients within the culture media that will allow the cells to grow fast, survive longer and ultimately produce large quantities of therapeutic protein. Media development, process optimization and basic understanding of CHO cell metabolism has paid the largest dividends in relation to advances in protein drug production. At the centre of all this on a cellular level is the mitochondria; an organelle that is the powerhouse of the cell and generates the majority of cellular energy through a process called respiration. We have carried out studies in our own group that have shown enhanced mitochondrial activity to be associated with increases in recombinant protein production when microRNA-23 (miR-23) was stably depleted using a microRNA sponge.

The unique part of the mitochondrion is that, unlike all other cellular organelles, it carries its own genetic material in the form of a small circular DNA plasmid. The mitochondrion plasmid encodes 37 genes that play a critical role in mitochondrial function and cellular respiration. Interestingly, genetically, a single mitochondrion is polyploidy which means numerous copies of plasmid DNA exist in the same space. Additionally, the mitochondrial genome is 10 times more susceptible to DNA damage than the nuclear DNA which can usher a variety of mutations that can impeded mitochondrial activity. In humans, >250 variants have been associated with metabolic disorders and disease so it’s not surprising that genetic mutations in the mitochondrion could affect CHO cell behaviour.

Image sourced from Optimal Living Dynamics 

Another fascinating thing about the mitochondria is that the presence of genetic mutations are quite common and widespread throughout nature and in a lot of cases are not harmful to the organism. This is due, in part, to a phenomenon called heteroplasmy. Mitochondrial heteroplasmy is where each mitochondrial organelle can contain multiple copies of DNA but each/all/some copies may have a small genetic difference. This genetic difference in a small subset of mtDNA copies that amount to a dysfunctional protein often goes unnoticed because the number of wild type copies prevail. However, there is only so much a cell can take. As the number of mutated copies increase, a biochemical threshold is reached where by normal cellular metabolism cannot be maintained.

Image sourced from Stewart and Chinnery, 2015

With the emerging potential for heteroplasmic variations to occur within the mitochondrial genome on top of the already existing nuclear genomic instability which contributes to the unpredictable behaviour of CHO cells during development, we sought to explore the genomic landscape of the CHO cells mitochondrial genome using next-generation deep sequencing. To get a good picture of this, we sequenced the mitochondrial genome of 22 CHO cell lines in collaboration with Dr. Colin Clarke from the National Institute for Bioprocessing Research and Training (NIBRT). This panel of 22 cell lines was quite expansive covering publically available parental lines such as CHO-K1 and CHO-S, recombinant protein producing cell lines including CHO-DP12, industrially developed production clones from our partners at Biogen in Boston, Massachusetts, our own in-house transgenic cells and a family of clones all derived from two cell line development programmes. Firstly, we built a reliable reference sequence for Cricetulus Griseus by sequencing the mitochondrial genome of a liver tissue sample directly from an outbred Chinese hamster, gifted to us by Prof. Michael Betenbaugh from Johns Hopkins University.

Of the 22 CHO cell lines that were sequenced, it was evident that there was widespread heteroplasmy present within the mitochondrial genome of all cell lines with each cell line possessing a mutation that changed the amino acid sequencing of a protein-coding gene.

Furthermore, various transfer RNA (tRNA) were found to have heteroplasmic variants which could have huge repercussions on the ability of the mitochondria to efficiently translate its own protein-coding genes. CHO cell metabolism and nutrient requirements has been a central theme of biopharmaceutical research, however, up until now, the genomic architecture of the CHO mitochondria has remained unexplored. Our work recently published in Metabolic Engineering details the genetic variability in the CHO cells mitochondrial genome. By understanding the level of heterogeneity in CHO cells, the rate of its progression and the potential impact on the CHO cell performance, it could be possible to predict and select for clones whose production attributes and metabolic programmes are best suited to producing high quality therapeutic proteins.

The post Exploring the unknown – Sequencing the CHO mitochondrial genomic landscape appeared first on National Institute for Cellular Biotechnology.

Ireland has now placed itself as the international home for pharmaceutical companies with 9 of the 10 leading biopharmaceuticals having manufacturing sites here. The Irish Development Authority (IDA) constantly showcases the leadership the emerald isle possess in biopharmaceutical drug production with companies like Bristol-Meyers Squibb (BMS) investing a further $900 million at their site in Cruiserath in Blanchardstown. This is an exciting area of research to be involved in and as so, I am involved in various research programmes which have real translational impact on the current climate of bio-manufacturing.

The industry is under constant pressure to evolve and develop novel therapies for the treatment of various diseases such as rheumatoid arthritis and cancer. CHO cells have become such a key player in the production of recombinant biotherapeutic proteins because of the complex, human-like, post-translation modifications added to the protein during its synthesis which ensures high drug activity and low immunogenicity. This mammalian cell line has served the biopharmaceutical industry for the past 3 decades and because so has earned itself a good track record for safety with the Food and Drug Administration (FDA) and the European Medicines Agency (EMA).

CHO Group, NICB, DCU

The Mammalian Cell Engineering group under Director Dr. Niall Barron at the National Institute for Cellular Biotechnology (NICB) located in Dublin City University focuses its research on improving the performance of CHO cells for the production of biopharmaceuticals.

With the increase in the incidence of various diseases such as cancer and the swift development of novel therapies moving through clinical trials, the biopharmaceutical industry is met with a bottleneck in relation to meeting the global demand for certain drug products and keeping up with emerging products, given the extensive time lines associated with cell line development. Despite g/L yields being routinely achieved, CHO cells are grown in vast 10,000 litre bioreactors to ensure that product demand is met. The sheer scale of these manufacturing facilities bring with it a high cost of production for these therapeutic drugs and as a consequence, the cost of these life-saving therapies is high.

Engi-miRs – microRNA-based CHO cell engineering

One area of particular interest in the mammalian cell engineering group is microRNA (miRNA or miRs) and their potential as engineering tools to genetically modify CHO cells to alter and improve their bioprocessing traits such as growth, ability to survive longer and production capacity. miRNAs are attractive molecular tools as they are small non-coding RNAs which have the capacity to regulate entire pathways through simultaneous target interaction. This regulation is at the post-transcriptional level thereby inhibiting protein translation and the viability of miRNAs as “engi-miRs” has grown over the last few years (see Review from our group). The non-coding nature of miRNAs minimises the translational burden on the protein producing CHO cell allowing for cellular resources to be committed to protein drug production.

Two miRNAs have gained considerable interest by industry, miR-7 and miR-23. Both of these miRNAs, when depleted using a microRNA sponge construct where observed to prolong cell survival and product yield in the case of miR-7 and triple product yield in the case of miR-23. DECOY-7 was developed at the NICB, culminating in a multitude of primary research articles which showcase its utility as an attractive engineering target for improving CHO cell production characteristics. Since its development, we’ve had considerable interest from industrial manufacturers such as our strong collaboration with Biogen Idec from Boston, Massachusetts.

miR-23 sponge graphical abstract

Single-use plastic reactors – Remodelling the Biopharmaceutical industry

The Mammalian Cell Engineering Group strives to adapt to meet the current needs of the biopharmaceutical industry. The Irish Pharmaceutical Healthcare Association (IPHA) plays a key role in driving translational and targeted research programmes in the pharmaceutical industry to meet Ireland's growing healthcare needs. As part of a Science Foundation Ireland (SFI)-funded collaboration in association with the Synthesis and Solid-State Pharmaceutical Centre (SSPC), I am part of a dedicated research team exploring the potential risks associated with single-use plastic bioreactors. There is considerable interest in the biopharmaceutical industry to move away from the use of large 10,000 litre stainless steel bioreactors which are costly to initially build and subsequently clean between production runs and adopt the use of smaller scale disposable plastic bioreactor bags.

Reservations for full implementation of these systems however ripple throughout the industry because of potential toxic leachable compounds from these polyethylene-based films that can contaminate the bioprocess posing a risk to both the process itself or the patient. One such reported leachate is bis(2,4-di-tert-butylphenyl)-phosphate (bDtBPP) which we have characterised to reduce CHO cell growth by 50% at concentrations as low as 0.035-0.1 mg/L in a recent Biotechnology Progress publication.

This observation is significant because it has previously been shown that this concentration can leach over the course of a bioreactor run which can open the bioprocess to risk in relation to considerably reducing the yield of therapeutic protein produced. Working in concert with our academic collaborators in the National Institute for Bioprocessing Research and Training (NIBRT) and Trinity College Dublin (TCD) and with support from our seven industry partners (Allergan Pharmaceutical Ireland, BioMarin Manufacturing Ireland Ltd., Eli Lilly and Company, Genzyme Ireland Ltd. – A Sanofi Company, Janssen Biologics, Merck Sharp & Dohme (MSD), Pfizer Ireland Pharmaceuticals), this project aims to fully characterise the risks, if any, associated with disposable bioreactors which will settle any concerns related to their safety and allow for the widespread adoption of this technology within the biopharmaceutical industry.

The Next-Generation of production cells

Whole genome sequencing has come a long way since the release of the human genome which cost a staggering $3 billion and 13 years to complete. Now, entire genomes can be sequenced for as little as €1,000 in just a few days making this technology more accessible to academic research groups like ours.

(Reproduced from the NIH National Human Genome Research Institute)

Sequencing efforts in the biopharmaceutical research community have performed whole genome sequencing using next-generation platforms such as Illumina. This has unveiled vast differences in the genomic landscape across various CHO cell lines revealing mutational differences in genes related to glycosylation and cell death, traits of particular importance within CHO cell biology and recombinant protein production.

We have undertaken a project in collaboration with Dr. Colin Clarke at the National Institute for Bioprocessing Research and Training (NIBRT) to identify genetic differences in the mitochondrial DNA of a panel of CHO cell lines. The mitochondria itself is the epicentre of cellular energy production and mitochondrial disruption can be a source of disease. The mitochondria is a cellular organelle which arose around 3 billion years ago through a symbiotic relationship with bacteria. As a result the mitochondria carries its own DNA template separate from nuclear DNA. This study has revealed widespread heteroplasmy throughout the CHO cell mitochondrial genome i.e. multiple mitochondrial DNA copies per CHO cell, each of which have unique genetic differences. This work will be used to identify CHO clones whose energy capacity best suit them for biopharmaceutical therapeutic production. We have also published the first Chinese Hamster Mitochondrial reference sequence, thereby making it available to the scientific community. I have won a bursary to present this work at the upcoming ESACT-UK conference this January in Oxford where leaders in the animal cell culture community meet to discuss the most recent innovations and promising directives in the field.

Biopharmaceutical drug production is a continuously evolving field with more advanced and complex therapies being developed. To keep up with this pipeline, a better understanding of the host CHO cell line is critical to ensure the efficient production of these emerging sophisticated therapeutic proteins. The CHO group at NICB is a multidisciplinary group comprised of post-doctoral and post-graduate researchers in the areas of molecular cell biology, cell engineering, proteomic analysis, transcriptomic analysis and sequencing capabilities. We have a long standing relationships with academic collaborators both nationally and internationally in addition to strong ties to industry. As a centre of excellence and innovation, the CHO group in the NICB under the leadership of Director Dr. Niall Barron strives to understand the biological mechanics of the hardest working cell on the planet.

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ESACT Frontiers Retreat 2016 Logo

The Scientific Meeting

The more conventional part of the meeting included Oral Scientific Sessions. All of them including many presentations from young researchers, PhD students or Postdocs, from a variety of topics related to Animal Cell Technology. It was interesting to have presentations of finished projects as well as presentations about ongoing work, which, despite not showing a lot of very conclusive data, showed very interesting ideas that are being investigated in the field. It is worth to highlight that a member of the eCHO Systems ITN, Nuša Pristovšek, did a great job during her talk! (see photo of Nuša below)

The Non-Conventional Meeting

In addition to the Oral Sessions, we had the opportunity to attend up to four Career Development Sessions: a Public Speaking Workshop, a Collaboration Workshop, a Think Tank Workshop and a Career Development Workshop. These workshops were the differential trait of the meeting, and I believe they were the sessions that people enjoyed the most.

Source: Tom Kost for ESACT Fromtiers Retreat 2016

The Public Speaking seminar was held by Tom Kost, former Director of Biological Reagents and Assay Development at GlaxoSmithKline and aimed to provide the audience with “key tips” to Prepare, Organize and Present their scientific work. My favourite tips were: show enthusiasm! If you are not passionate about your presentation, how can you expect to inspire your audience? Keep it simple and focus your message and edit, edit, edit yours slides and use them as a support to your presentation, not as the presentation itself.

Source: MH Consulting - Collaboration Workshop - ESACT Frontiers Retreat 2016

The Collaboration Workshop, driven by Michael Heath from MH Consulting, was such an experience for me as I had never done something like this before.

The first part of the workshop was  a talk about cultural differences. It was very interesting to learn how people from different cultural environments have different ways to communicate, which need to be understood in order to have fruitful collaborations. Those differences are neither intrinsic to human nature, nor specific personality traits of a given individual, but they belong to each person's cultural background and are often overseen. However, these existing cultural differences have a very important influence on how people understand each other and how a collaboration can develop. Concepts such as low context or high context cultures, Direct Negative Feedback or Indirect Negative feedback, were completely unknown to me before the workshop. The two first refer to how communication carried out in specific cultures. In low context cultures, people use precise and simple messages versus the more sophisticated and “read between lines” messages that people from high context cultures use. The two last are related to how people from different cultures provide negative feedback to each other. For instance, direct and honest negative feedback in a group as opposed to negative feedback wrapped with positive messages to soften critics which are only given in private. Now I know I will have to pay more attention to those details, and actively make efforts to understand the cultural context of my peers if I want to improve our communication.

The second part of the workshop involved the participation of the attendants in a collaborative competition. It would be unfair to go into details of what we did there, as the main exercise actually works best if people never have done it or heard about it before. What I can share is the take home message we all got from this exercise: the importance of the synergy effect of collaborations, as when developed in a good and effective collaborative environment, the final result will be greater that that sum of the parts.

The Think Tank workshop was also very interesting. Participants distributed in teams, and had to prepare a poster about specific research-related topics, including, for example, IP versus open source research or crowd-funded research. To promote participation and debate inside the teams, we only had access to a paper board, colourful post-its and pens! It was very nice to dive into a new topic with nothing else than a group of young scientists and to have to build a poster with our own hands. I must say that I really enjoyed sharing ideas with my peers and I travelled back to my childhood when building the poster!

Finally, participants attended a Career Development session. The format of this session was again very good. Three speakers, Kerstin Otte, Professor at University of Applied Sciences Biberach, Hitto Kaufmann, currently Global Vice-President of BioPharmaceutics Development & Platform Innovation within Sanofi R&D and Marcel Tigges, Co-founder of BioVersys, shared their experience with the audience. In their talks, they highlighted the main skills to succeed as well as the decisions, good ones and bad ones, that lead them to success in their respective careers in an academic, industrial or start-up environment.

An eCHO meeting point

Several of my fellow eCHOs from the eCHO Systems ITN attended the meeting as well. This made it even a better experience for me, as we had the chance to get updated on our respective projects and promote our network at the event.

eCHO Systems crew explaining their work at the ESACT Frontiers Retreat 2016 Poster Sessions

eCHOs sightseeing in Lyon before attending the ESACT Frontiers Retreat 2016

In Summary

I would say the organizers were right and my expectations were fullfilled. The 1^st ESACT Retreat 2016 was a great forum for young researchers to present and promote their work. We could learn about science and other important soft skills, as well as grow our network in a very friendly, relaxed, and professional environment. I highly recommend you to attend future meetings and hope I can do it myself! I would also like to thank again the organizing committee for their wonderful job and invite you to watch the great video they recently released.

https://www.youtube.com/watch?v=AgcTT7gnNkA

In addition, you can access the Abstract book of the meeting here.

Ricardo Valdés-Bango Curell
Marie-Curie PhD fellow at NICB, Dublin City University
eCHO Systems International Training Network (ITN)

My research focuses on the investigation of genetic switches, endogenous or exogenous, for controlled transgene expression in mammalian cells. My final aim is to develop potential applications of these in the biopharmaceutical industrial context. I am funded by an Early-Stage Researcher Marie Skłodowska-Curie action from the European Comission, as part of the  Innovative Training Network eCHO-systems,

The post ESACT Frontiers Retreat in Lyon 2016 – A Great Unusual Scientific Meeting appeared first on National Institute for Cellular Biotechnology.

When people ask me what I do and I reply, I’m a PhD candidate, they don’t always get it. The best example I can think of is my Nan, she thinks I’m a bit dim because I’m still in “school” at the age of 24. It can be a struggle to relate what you do, not only to the non-scientific community, but also scientists in different fields. When friends ask, I tell them a PhD is a 300- to 400-page book that only a handful of people will ever read and my day to day involves adding small volumes of liquid to other small volumes of liquid, with documentation of the results. Why, you may ask?

The biopharmaceutical industry is one of Irelands largest, with capital investment exceeding €8 billion in the last 10 years. Currently 9 of the world’s top 10 pharmaceutical companies have a base in Ireland, with annual exports of €39 billion, as recently published in The Irish Times. This industry not only exists, but thrives due to the inevitability that people get sick. A limitation of the biopharma sector, by no fault of their own, is the inability to provide affordable treatment options for many debilitating and life threatening diseases. In some instances, this is not because a bio-therapeutic does not exist, but simply the drug cannot be manufactured in reliable, robust and cost-effective manner. That’s where we come in… kind of!

CHO cells have become a favourite of the biopharma industry due to their uncanny ability to synthesise complex human-like proteins and their adaptiveness regarding culture format. Numerous factors contribute to the generation of a stable recombinant CHO (rCHO) cell line. CHO cells are enabled  to produce therapeutic drugs by the stable integration of DNA, encoding for the therapeutic, into their genome. Traditional cell line development is essentially a numbers game; rCHO cells are screened by the thousand for what is termed high producers, single cells from an original pool are isolated and their drug production capabilities monitored, with the best candidates progressing to process optimisation. The most pressing drawbacks to current clonal screening strategies include the vast number of clones that must be vetted, and the naivety of this practice, we don’t know why one cell outperforms another.

Figure 1: Schematic representation of recombinant CHO cell line development. Genetically engineered CHO cell lines are used to make stable transgenic mixed pools expressing the drug of choice. Clonal isolation and screening is used to find the top candidates. Subsequent process development is required to maximize product output and CHO cell production efficiency.

The biopharmaceutical industry requires robust, reliable and reproducible bioprocesses, yielding a high level of good quality product to ensure, above all else, patient safety. Reproducibility is achieved and enhanced by understanding and controlling bioprocess variables. Evolution of process analytical technology (PAT), has aided this greatly at experimental, pilot and industrial scales, involving both on-line, in situ, and off-line monitoring of culture variables. Real-time assessment of the bioprocess enables the manufacturer to identify issues or irregularities and act on them, saving any potential loss in yield.

This is all great but we still don’t know what is going on inside the cell and why some cells make good quality homogeneous product while so many others fail in this regard. This is where genetic engineering of CHO cells comes in.  In the simplest of terms, my research, which is funded by Science Foundation Ireland, aims to improve the productive capability of a CHO cell culture. This is achieved by altering natural cellular pathways to improve the growth of these cells. The idea being, if we can increase the number of cells present in the culture with no negative effects on how they make a product, it is back to a numbers game - more cells equates to more product. To do this I am altering specific gene regulatory elements, microRNAs (miRNAs), at defined points in the culture in the hope to maximize the process yield.

The post miR-acle Grow: Improving CHO cell Growth by miRNA Dysregulation appeared first on National Institute for Cellular Biotechnology.

The eCHO Systems ITN is a consortium of  four academic partners; Denmark Technical University (DTU), Copenhagen (Denmark), University of Natural Resources and Life Sciences (BOKU), Vienna (Austria), University of Kent (UKent), Canterbury (UK), and Dublin City University (DCU), Dublin (Ireland), together with several industrial partners, including the Austrian Center for Industrial Biotechnology (ACIB) and UCB Pharma.

The main aims of the network are providing training to 15 PhD students and seeking to significantly advance the Chinese Hamster Ovary (CHO) cell platform for production of biopharmaceuticals. The projects in the consortium are all focused on developing and applying synthetic and systems biology tools to create the next generation CHO cell factory.

Currently, three Early Stage Researchers (ESRs)  are enrolled on the PhD program at Dublin City University: Prashant Kaushik and Ricardo Valdés-Bango Curell, as part of the CHO engineering group at the National Institute for Cellular Biotechnology (NICB), under the supervision of Dr. Paula Meleady and Dr. Niall Barron, and Krishna Motheramgari, as part of the Bioinformatics group at the National Institute for Bioprocessing Research and Training (NIBRT), under the supervision of Dr. Colin Clarke.

During this first year, the students participated in a series of  Network Training Activities (NTAs), which took place at the different partner sites.

Copenhagen: The CHO cell and the 'CHOmics' world 

In September 2015, the first two NTAs took place at DTU in Copenhagen and were organized by Dr. Mikael Rørdam Andersen, Associate Professor at the Department of Biotechnology and Biomedicine, Technical University of Denmark (DTU). These two courses focused on Genome Engineering and Cell Culture Methods and Genome-Scale Science and Modelling, with a special focus on the CHO cell factory and the production of recombinant therapeutic proteins. The students learned about the latest advances in cell engineering and genome-scale science related to the CHO cell platform. The practical part of the course allowed them to get hands-on experience handling CHO cell cultures in bioreactors, monitoring critical culture parameters such as pH and metabolites, and evalute their impact on productivity.

In addition, the students had the opportunity to visit several manufacturing sites from various biopharmaceutical companies located in the Copenhagen Area, such as Biogen, CMC Biologics and Novo Nordisk A/S. At each site, the students met with industrial experts and discussed the current challenges related to CHO cell factories and recombinant prtein production.

Dublin: How do we make sense of the data?

Later, during the first two weeks of June 2016, the NICB in Dublin hosted the members of the eCHO Systems ITN to attend the second NTA, focused on Advanced Bioinformatics and organized by Dr. Colin Clarke, Principal Investigator in the Bioinformatics group at NIBRT. The topics covered included the acquisition and analysis of large transcriptomic, proteomic and glycomic datasets obtained from CHO cells using a broad range of technologies, from microarrays, Next-Generation Sequencing (NGS), miRNA and proteomics profiling.  During the practical sessions of the course, the students had the chance to work on data from real CHO-based experiments and were able to perform their own bioinformatic analysis. Case studies always aimed to address industry-relevant problems related to production of biopharmaceuticals in CHO cells using a multi-omics bioinformatics approach. In addition, the students had to work in teams on small projects, which required putting into practice all the learned concepts.

During their stay in Dublin, the eCHO ESRs also had the opportunity to visit the Pfizer manufacturing site in Grange Castle (Dublin, Ireland) and discuss the present and future challenges in recombinant protein production at an industrial scale with on-site experts.

Alongside with their training, the eCHO crew had the opportunity to get a taste of Irish culture through several social fun activities, such as a day trip to the Wicklow mountains and Glendalough or the visit of the Guinness Storehouse in Dublin.

Kent: How do we characterize our valuable products..?

The third NTA took place at the University of Kent, in Canterbury (UK), and was organized by Prof. Dr. Mark Smales, Professor of Industrial Biotechnology, and his collaborators Dr. James Budge and Dr. Tanya Knight. On this occasion, the course was focused on Process Analytical Technologies such as chromatography, mass spectrometry, fluorescence spectroscopy and circular dichroism, applied to recombinant protein production and characterization. During this course, the students also had an extensive amount of laboratory training involving the purification of a monoclonal, its characterization using SDS-PAGE and mass spectrometry, as well as specific applications of the above-mentioned techniques.

In addition to all the training, social and team building activities were also planned during the stay in Canterbury. Examples of those were a lovely dinner at a traditional English Pub, a guided walking tour around the Canterbury Old Town and a half-day trip to the neighbouring Cliffs of Dover.

Vienna: ...and our cells?

Finally, the last NTA was held in Vienna early September 2016. On this occasion the course was organized by Prof. Dr. Nicole Borth, Dr. Jennifer Schoberer and Dr. Markus Schosserer from BOKU. This time, the main topics were Cell Imaging, Flow Cytometry and Presentation Skills. The ESRs had a comprehensive overview of the current techniques for live cell imaging, immunofluorescent staining and flow cytometry applications to characterize and sort cell populations. As usual, a special emphasis was given on the potential of such techniques to analyse CHO cells for specific improved phenotypes such as high producers or fast growing cells.

The practical sessions on fluorescence microscopy, confocal microscopy, gSTED microscopy and RAMAN microscopy allowed the students to prepare live and fixed biological samples for fluorescent staining and operate by themselves the different types of equipment. The Presentation Skills training was organized by Wolfgang Kainz from Business Theater. Here, the students had the opportunity to be recorded while presenting their Annual Meeting slides. The videos were then analyzed by the group to provide corrections and advice to improve body language and help the students improve their presentations.

Once more, social activities were not missing in the schedule and the ESRs were given a taste of the rich Viennese lifestyle. Some examples are a guided tour around the city center in Vienna and inside the Schönbrunn, a lovely dinner in the oldest Viennese restaurant and a day trip to the region of Wachau, famous for its vineyards.

This project has received funding from the European Union’s Horizon 2020 research and innovation programme under the Marie Sklodowska-Curie grant agreement No 642663

Ricardo Valdés-Bango Curell
Marie-Curie PhD fellow at NICB, DCU.
eCHO Systems International Training Network (ITN)

Funded by an Early-Stage Researcher Marie Skłodowska-Curie action from the European Comission, as part of the  Innovative Training Network eCHO-systems, my research focuses on the investigation of genetic switches, endogenous or exogenous, for controlled transgene expression in the CHO cell factory and their potential applications in the biopharmaceutical industrial context.

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