Optimizing quality & compliance for cord blood-derived cellsPublished: February 15, 2019
Cell Therapy Quality/ CMC & Analytics
Roger Horton & Christopher Leonforte
ROGER HORTON was trained in Biochemistry and cancer immunology and obtained a PhD in 2006. He previously worked in academia and then at Cancer Research UK before joining Anthony Nolan in 2007. Roger is the ANCTC Cord Blood Bank operational manager, and works to provide cord transplants globally for patients in need in conjunction with operations in London head office and the team at ANCTC. He is currently in training to be a FACT inspector and has been the HTA’s designated individual for ANCTC since 2016.
CHRISTOPHER LEONFORTE is scientist-turned-manager who trained in Biochemisty in 2009 and completed an MSc in Biomedical Science in 2017. He has worked at the Anthony Nolan charity since 2010 and is passionate about the work that it does. Chris is the Cord Blood Bank processing and quality control manager. He oversees the processing, testing and storage of the cord blood. Chris works with an excellent team to maintain the bank’s FACT-Netcord accreditation and HTA license and he has completed training to be a FACT Netcord Inspector.
Citation: Cell & Gene Therapy Insights 2019; 5(1), 103–108
Can you give us some brief background on the Anthony Nolan Cord Blood Bank and Cell Therapy Centre, and tell us about your own specific roles within the organization?
RH: Going back to 2004, 2005, cord blood was just starting to be a graft of interest around the world and Anthony Nolan found themselves importing quite a few grafts from abroad for use in UK clinics.
The issue that emerged was they could be quite expensive, depending on where in the world they came from, so we looked at it and decided we could probably do it ourselves. The idea of the Anthony Nolan Cord Blood Bank and Cell Therapy Centre was born from that.
By 2007, we’d built the place. In 2008 we received our HTA licence and were in our clinical phase, and it was just a case of building up the inventory from there. It took from about 2008 to 2012 to collect enough units to start getting HLA matches from searches. All the search function was and is provided by the Anthony Nolan registry in London – we already had a registry so there was no point in duplicating it – and in 2012 we got some of our first unit report hits and we did our first transplant.
We went on to get NetCord-FACT accreditation, which is the gold standard for cord blood provision around the world – we’ve now been accredited twice by FACT and we’re just about to go into our third round. We’ve now done 165 transplants all around the world – South America, Australia, all over Europe, a lot in the USA as well.
So that’s the summary of the Cell Therapy Centre. Where we at the Cord Blood Bank come in is basically in situations where the adult register cannot find a match.
Cord is more permissive to HLA mismatches, so if you have someone with an unusual HLA type or maybe a mixed background, we fit in quite nicely where the adult register can’t. Or occasionally you’ll get an adult donor who at the last minute gets a cough or a cold and obviously cannot then donate. In those instances, if a cord search has already been done, we can then provide it really quickly so the patient doesn’t have to wait longer for their treatment.
In terms of my specific roles, I’m more outward-facing. I deal with the NetCord-FACT and HTA accreditations. I also interact the transplant centers, both relating to cord provision itself but also with onsite training and support, because cord represents probably only about 5–10% of transplants. Some transplant centers consequently struggle to keep all of their staff trained, particularly if they’re a big center – an individual member of staff might not do a cord transplant in a given year.
CL: I oversee the processing and QCing of the cords, from start to finish – from the point of reception to point of storage – and I manage the team responsible for carrying out those tasks. On top of that, there’s all the other things that go with building management – stock control and making sure the clean room is maintained, as well as all the equipment. So whilst Roger is mainly facing outwards to the wider community, I’m making sure that everything is up to scratch internally.
Can you tell us more about any particular projects you are involved with currently?
RH: We’re focused in several directions at the moment. 2019 for us is the year of inspections: HTA is every 2 years, but NetCord-FACT is every 3 years – so every 6 years it all lines up? This year we have NetCord-FACT in the summer and HTA in September, so we have to make sure we’re 100% in compliance with all the required standards. I’m also trying to continue developing the transplant center engagement programme to make sure we support all people, whether they are regular users or first-time users of cord.
CL: Obviously everything that comes through the door isn’t going to be of clinical grade. I’ll explain more about that later, but only about 20% or so of everything that comes in might be processed as clinical grade and used for a transplant. The rest is potentially going to be destroyed because there isn’t a market for it as such.
However, what we’re trying to do is drum up interest in these ‘research grade’ cords – to see if we can best make use of the altruistic mother’s donation. These cords can potentially better transplant outcomes or treat diseases as well. So we’re just trying to make sure we’re covering all the bases and trying to meet some needs that might not be met otherwise.
RH: Absolutely. For one thing, there are up-and-coming biotherapeutics, which we’ll touch on later: there is no reason why this spare material, which isn’t going to be directly used for our primary clinical purpose, can’t go to grassroots academic research, or be used in preclinical studies for novel therapies.
What tools and processes do you employ to optimize donor cell quality, and can you tell us about some of the challenges you encounter relating to this in the cord blood-derived cells arena?
CL: We use an automated system by GE – the Sepax system. We want our system to be closed, so that the only points the material can be exposed to air would be at the point of collection, obviously, and then potentially at the point of infusion as well.
We have four fixed collection sites that operate 24/7 and the bank is open 7 days a week. That means we receive cords daily, which has both its pros and cons: you want to get the cords into the tank as fresh as possible for viability purposes. On the other hand, the fresher the sample, the worse it will process because of the red cell content. As the cord unit gets older they shrink and sediment out better. (The processing system works by sedimentation, effectively).
So it’s kind of a trade-off. You could have some really good cords come through, but they might not process as well because they’re fresher. That seems a bit of a shame, but it’s all about viability at the end of the day – that’s more important than cell recovery.
We have various thresholds for this. We do want to be banking as large a cord as possible because it’s a fixed source and once it’s collected, you can’t ever get a repeat sample if it were to be needed. So we want cord units with the biggest cell content possible. But with the natural variability of donors, you can never say on one day that we’re going to be storing x number of cells – it varies all the time. There is quite a lot of waste associated with it, which is why we’re looking at trying to utilize the material that doesn’t meet our criteria for other purposes.
And how do you measure cell quality?
CL: We do a nucleated cell count on everything and after processing, we do flow cytometry. We also have a couple of viability assays that we employ.
One is to measure whether the cell is alive or dead – that’s using a chemical marker called 7-AAD – and we also do another assay, based on an Annexin V marker, which will tell us if the cell is alive, dying or dead. That gives us a bit more insight into how good a unit actually is: one assay might say the sample is 70% viable but if we do the other assay, it may come back more negative at 40% – this might lead us to conclude that the unit is not suitable for transplant, so we potentially wouldn’t use it for clinical purposes.
We also do a potency assay, which involves seeding some of the stem cells and checking their growth in controlled conditions. This is to look at how well it will engraft. The incubators we use and the culture media provide the best possible conditions for these cells to grow – if they can’t grow in those conditions then they’re probably not suitable for transplant. We have various cut-off points to gauge this.
So going back to the previous question, we might process 20% of everything, but there’s still attrition as we validate the units, and as we get tests back for virology and hemoglobinopathies as well. That 20% will probably shrink down to somewhere between 15 and 18%, ultimately.
But at the end of the day, we want to make sure that what we’re banking is going to be of use to a patient. You could effectively bank everything that comes through the door, but it would be a waste of time and capacity.
What structures and technologies do you employ to ensure compliance on the QC side of things?
CL: As part of our licensing and accreditation, we have to have everything serviced and up to date. But we also do internal quality controls for the flow cytometer, which we run daily, just to verify we’re enumerating the stem cells properly.
We also take part in external quality control tests as well – or the flow cytometer, our cell counter and also for our potency assay. The frequency of these tests varies throughout the year, but it just gives us that anonymous comparison to other labs doing the same thing, helping us make sure we’re not under- or over-enumerating the samples.
RH: There’s a quality team who pitch in as well. We have quite a regular schedule of audits – throughout the year, the quality team will organise audits based on various criteria from the HTA or FACT to make sure our procedures are compliant. Within those audits, the team also checks the Standard Operating Procedures (SOPs) are followed and that the outcomes are what were expected as well. Each process and procedure has a defined end point or expected range for its outcome, so if anything deviates, it can be flagged on the system and we can investigate.
What are the chief pros and cons of utilizing cord blood-derived cells as a starting material for cell therapy, particularly in relation to CMC requirements?
RH: Firstly, what you’re getting is a relatively well-defined product. We have a standard collection procedure, we have standard criteria, we have cut-offs for age and things like that. You’re guaranteed a given level of viability to start with. So the product itself is relatively consistent apart from variability in the cell numbers it contains.
This means that because of the large numbers we collect, if someone were to stipulate to us, say, ‘I require x age unit, I don’t want any clots and I need at least 900 million cells’, it should be possible for us to sort through what we get on any given day, find units with those criteria and dispatch them quickly.
We also conduct two lots of virology testing. Obviously, when mum gets booked into the hospital to give birth, they do blood tests. We have access to that virology data so we know there are no significant issues up front. We’ve also got the ability once units arrive here to do a further test, which we can turn around within 24 hours with some of our local suppliers – that can cover the critical things like HIV, HPV, HCV, HTLV, etc. We can then follow on with the NAT tests quite shortly afterwards, which I believe is a requirement for starting materials to go into a MHRA-accredited GLP or GMP facility – you need your virology up front and that’s something we can do.
It can sometimes be more challenging to try and do that with adult donations, because your adult donor is far more mobile than a donating mum – they can go on holiday, for instance. Once a mum is in hospital, she’s staying there for a period of time – you’ve got the unit there to test.
So quick turnaround is certainly one of the benefits. I would also say that judging by the work we’ve done so far, if you’re looking to isolate cell subsets to then expand for use as an ATMP, the proliferative capacity of cord seems to be really good. We have got some partners who have worked in NK cells, for example – our own research institute works on NK cells, too – and the proliferation is really good because it’s such a naïve cell source.
It’s also readily available. With adult donors, accessing the material can be quite a tricky and convoluted process. With cord, though, we have ethics in place to be able to dispatch it to anyone who gets project approval through our ethical board and their own internal medical review system. That reduces project delays.
Anthony Nolan Cord Blood Bank & Cell Therapy Centre
Anthony Nolan Cord Blood Bank & Cell Therapy Centre
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