Joint Surgery and Cell Therapies

CARTILAGE RESEARCH GROUP

Directors: Professor James Richardson and Professor Sally Roberts

Members of the Research Team:

Karina Wright, Claire Mennan, Jan Herman Kuiper, Helen McCarthy, Johanna Wales, Charlotte Hulme, Annie Kerr, Barbara Linklater-Jones, Atanu Bhattacharjee, John Garcia, Mike Williams, Naomi Dugard, Paul Harrison, Nikki Kuiper and PhD students, Jessica Sykes, Jade Perry and Tim Hopkins.

Clinical Support:

Messrs Peter Gallacher, Robin Bannerjee, Drs Bernhard Tins, Andrea Bailey and Professor Iain McCall

Good news for this year has been that the Arthritis Research UK has extended our original 5 years’ funding for the Tissue Engineering Centre (TEC) for another 5 years until April 2020. This will enable more research into cell therapies and cartilage repair, in collaboration with our partners in Keele University as well as in Cambridge, Aberdeen, York and Newcastle Universities. Several members of the team are funded by this charity, both via the Centre and also individual project grants. We are once more very grateful to the Institute of Orthopaedics for their support of several other projects and members of staff and students. We are very fortunate here in Oswestry to have such an organisation with so many people giving of their time freely to help it run so effectively.

On a staffing note, we have had a productive year with respect to babies, with 2 members of staff currently on maternity leave. During the year, two more PhD students enrolled in projects related to the TEC, one funded by the Institute and the other by Keele University.

Press Release From the Robert Jones & Agnes Hunt Orthopeadic Hospital October 2017

Rare treatment for arthritis wins
approval for use on NHS

A rare treatment for a type of arthritis in the knee – currently only offered at The Robert Jones and Agnes Hunt Orthopaedic Hospital (RJAH) in Shropshire – has won official approval to be funded by the NHS.

Autologous Chondrocyte Implantation (ACI) is a technique used to help patients with an articular cartilage defect, something that can impact younger people in their 20s and 30s – sometimes as a result of a sporting injury.

The procedure sees a sample of cartilage removed from the patient’s knee from which their own cells are then grown in a laboratory, a process which takes around three weeks. These cells, known as chondrocytes, are returned into the patient’s defect area in a second surgical procedure.

ACI clinical trials have been running in several locations over a period of years, but it was only this month that, following extensive appraisal, the National Institute for Health and Care Excellence (NICE) concluded the volume of evidence to be enough to approve the technique for wider use on the NHS.

RJAH is the only site in the UK currently able to offer ACI. The specialist orthopaedic hospital has been working in partnership with Keele University and the Oswestry-based Orthopaedic Institute; the trio setting up the Oscell Cell Manufacturing Facility to produce the chondrocytes.

ACI procedures are performed at RJAH by Professor James Richardson and Mr Pete Gallacher, both experienced Consultant Orthopaedic Surgeons.

Professor Richardson said: “This facility has provided treatment to over 400 patients from across the UK in the last 20 years, along with many published studies of the outcome of ACI in the knee and ankle. “ACI can help patients who have a particular defect or early arthritis, one which starts with what appears as a small pothole on the surface of the knee. When the normal, smooth surface is broken in this fashion it may not heal naturally. “We take a small sample of cartilage out and grow cells up in the lab. These cells are then put back into the patient’s knee in what is a second, bigger operation. “The patients are usually only in hospital for two days post-operatively, though there is a lot of rehabilitation and physiotherapy to follow after that. The results we have seen have been positive, particularly if the patient has not had prior microfracture.”

The announcement that ACI would now be made available on the NHS was made at the 11th annual Oswestry Cartilage Symposium, hosted by RJAH earlier this month, at which 80 specialists from the Arthritis Research UK Tissue Engineering Centre and others from around the world were gathered.

ACI can only be offered to patients who meet a number of defined criteria as set out by NICE in their guidance note TA477, which can be read  at  https://www.nice.org.uk/guidance/ta477/chapter/1-Recommendations.  These criteria include that the patient must not have had previous surgery to repair articular cartilage defects, as evidence from previous trials has shown ACI to be much less effective in these cases.  There should also be minimal osteoarthritic damage to the knee and the defect should be over two square centimetres.

Prof Richardson added: “The typical patient would be under the age of 40 and would be someone who might have picked up an injury playing football which has led to a chondral defect in their knee that causes pain and is tender on examination.  “This patient would otherwise be healthy, with intact ligaments and a straight leg.”

RJAH accepts patients from across the UK. It is important they have the support of their orthopaedic surgeon and an MRI which confirms the diagnosis and satisfies the criteria. Referrals can be made to either Mr Pete Gallacher or Prof James Richardson.

 

Projects at a glance;

Quality of Repair Following Autologous Chondrocyte Implantation (ACI): Is there a LINK with Clinical Outcome?

Helen McCarthy, Mike Williams, Naomi Dugard, Iain McCall, James Richardson and Sally Roberts

Funded by Arthritis Research UK

Since 2010, we have been collating data from a retrospective study of autologous cell therapy (REACT), using a postal questionnaire completed by patients who have previously received such treatment for chondral or osteochondral defects of the knee here at this hospital.  This group of patients has more recently also been used to study how the quality of repair tissue formed in the treated defect might relate to clinical outcomes such as the modified Lysholm Score, a measure of knee function. The overall aim of this is to see if there are any correlations that might help us learn more about the success of the treatment.  Magnetic resonance images (MRIs) and histology of biopsies of the repair tissue which forms after treatment have been used to determine the structure and make-up of the repair tissue.  The repair tissue biopsies themselves were very variable in structure, with the majority being fibrocartilage. The presence of blood vessels and / or very poor quality tissue in the repair tissue was associated with both pain and limping. 

We used two different scoring systems for looking at the MRIs, one which assesses the whole joint, whilst the other focuses only on the region of the joint that was treated.  We have found so far that the scores obtained by looking at the whole joint have no correlation with clinical outcome, but those that specifically relate to the treated area, do.  These results are encouraging in showing a significant association between the overall MRI score and several of the individual parameters with the clinical outcome.  Further analysis of this may actually provide information on the pain-generators, not only in knees with focal cartilage defects, but also in more generalised osteoarthritis that we hope this treatment will eventually be effective in treating.

HUMAN BONE MARROW MESENCHYMAL STROMAL CELL CHARACTERISTICS ARE SENSITIVE TO SELECTION TECHNIQUE AND OXYGEN LEVELS

Karina Wright, Helen McCarthy, Nicholas Forsyth, Sally Roberts.

To obtain mesenchymal stromal/stem cells (MSCs) from bone marrow, a bone marrow sample is usually prepared in the following way: a particular group of cells with a single nucleus (or mononuclear cells (MNC)) is first isolated by layering the bone marrow over a special liquid which is then centrifuged so those cells (which includes MSCs) lie in one particular layer. They are pipetted off and placed in a tissue culture flask with tissue culture media (a fluid containing everything necessary for them to grow). It is reputed that only MSCs will attach to the plastic of the tissue culture and so continue to grow. All of this is usually carried out under normal atmospheric oxygen levels (called normoxia, ie 21% O2). In a few studies, however, scientists have shown higher numbers of MSCs and better growth in culture when cells are derived from complete bone marrow (without separating the MNCs first) and growing them on tissue culture flasks which have been coated with a molecule called fibronectin (which cells often like to attach to). They have also cultured the cells at lower oxygen levels (‘hypoxia’, eg 2% O2).  We compared these two different isolation techniques using bone marrow aspirates from 4 young, healthy donors (aged 22-32 years).  We found that MSCs isolated and cultured from whole bone marrow in hypoxia yielded the highest number of cells compared to the other method.  We also found that the different isolation techniques resulted in different sub-populations of MSCs with varying characteristics. This is useful to understand the biology and behaviour of these cells and how the different sub-populations may be important for treating patients with different root causes of disease in the field of tissue engineering and cell therapy.

SPONTANEOUS HEALING OF ARTICULAR CARTILAGE (SHARC): A HUMAN STUDY

Jan Herman Kuiper, John Garcia, Sally Roberts, Bernhard Tins, John Oxtoby, Nicola Kuiper, Oksana Kehoe, Karina Wright, Caroline Stewart, James Richardson

Funded by Medical Research Council (MRC)

For hundreds of years it was believed that cartilage in joints such as the knee does not heal naturally after an acute injury and such injuries are indeed currently believed to lead to arthritis developing. However, recent studies based on magnetic resonance imaging (MRI) have indicated that in some cases cartilage does have the ability to repair itself (Figure 1). The process by which this occurs has been investigated in mice, but remains largely unexplored in humans. The SHARC study was designed to fill in this knowledge gap by characterising the natural repair process of articular cartilage by capitalising on a treatment used here in the hospital, autologous chondrocyte implantation or ACI. For this treatment, a small piece of healthy cartilage is removed to obtain cells for culturing in the laboratory for 3 weeks after which they are implanted into the damaged area. This process allows a perfect opportunity, if the patients are happy, for us to monitor how the area where the cartilage was taken from responds and heals (or not) on its own accord.  For the 12 months following treatment we will use different techniques, including a questionnaire for the patients, biomechanical assessments, imaging, measuring certain molecules or biomarkers in synovial fluid and blood, as well as mathematical modelling.

We know that certain strains of mice, and indeed some people, heal better after skin or joint injury than others. We hope to identify if there is a panel of biomarkers (biochemical, imaging or mechanical) which can be used to predict an individual’s natural cartilage healing capacity. The findings of the study will hopefully provide valuable information on the pathogenesis of arthritis following injury and also to help clinicians and scientists in developing personalised therapies for treating it.

Figure 1: Illustration of the healing capacity of knee cartilage using MRI. (Normal cartilage is white.)

The cartilage defect visible in A (white arrow) appears to be improved 2 years later in B.

 

 BIOMARKER DISCOVERY IN ORTHOPAEDICS: MATCHING PATIENTS TO THE BEST TREATMENT

Charlotte Hulme, Heidi Fuller, Emma Wilson, Sally Roberts, Andrea Bailey, Peter Gallacher, James Richardson, Annie Kerr, Karina Wright.

Funded by Arthritis Research UK

As in all treatments, a few patients who have their osteoarthritis treated by biological treatment, such as cell therapies or microfracture, do not respond so well as others, though we do not know why. Our previous work has identified particular molecules in synovial fluid or blood (called ‘biomarkers’) that have the potential to identify which patients are unlikely to improve clinically following the cell therapy known as autologous chondrocyte implantation (ACI). These investigations focussed on protein markers whose biology is known to relate to poor repair of cartilage or general inflammation e.g. an enzyme, aggrecanase-1, and CD14. Recently, we have used a highly specialised technique called proteomics to identify larger numbers of other potential marker proteins which are altered within the synovial fluid of patients who either do or do not respond well to ACI. The proteins identified using this approach include many that have never been investigated before and which therefore provide attractive new candidates. To date, we have confirmed that three of these proteins, S100A13, MMP1 and MMP3, have the potential to inform us of whether or not a patient is suitable to continue through the ACI procedure. We have also used bioinformatic analysis, using complex computer programmes, to inform us of how the ACI procedure may be having different biological effects on individuals who improve following ACI compared to those who do not (Figure 2). We hope that this work will help us to identify a reliable panel of molecular markers which can be used to help the surgeon and patient decide on what is the best treatment for that particular person. This would not only save the health service money, but most importantly get people pain free and back to their normal lives quicker.

  

 Figure 2: The figure highlights the different biological response in the knee joint fluid of patients who do not show improvement (non-responders) compared to those that do improve clinically (responders) after the cell therapy, autologous chondrocyte implantation (ACI). Boxes shown in orange or blue represent biological functions that show different responses and grey boxes show no change. The patients who did not respond well to ACI had a much greater response of lots of biological functions (highlighted with bold text) compared to patients who did well clinically. In the future, tailored treatments to reverse these biological effects could improve patient outcome.

 

Can cell therapy with umbilical cord-derived mesenchymal stromal cells delay or prevent the progression of Osteoarthritis in pre-clinical models?

Jade Perry, Claire Mennan, Helen McCarthy, George Bou-Gharios*,, Rob Van‘T Hof*, Peter Milner*,  Cosimo De Bari#, Anke Roelofs#, Karina Wright, James Richardson & Sally Roberts.  *Liverpool University; #Aberdeen University

Funded by Arthritis Research UK

Osteoarthritis (OA) is a complex degenerative joint disease, characterised by degradation and loss of articular cartilage (causing the joint space to narrow) as well as a lot of changes in the bone, such as  abnormal bony spurs or ‘osteophytes’ forming. Currently, there are no effective pharmaceutical or non-surgical therapies to reverse osteoarthritis. Researchers around the world are seeking new approaches, including seeing if cell therapy could be useful. Here in Oswestry we have been obtaining mesenchymal stromal or stem cells (MSCs) from human umbilical cords (UC-MSCs) for some years now. These are an attractive source of cells for regenerative medicine since they are fairly easy to grow and they also appear to have anti-inflammatory properties which may be useful and make an immune reaction less likely if used in a different individual.  We are currently working with collaborators in other universities to see if the cells prepared here in Oswestry could delay osteoarthritis developing  in their models.

One of these (called the DMM model) is very like endstage osteoarthritis, with loss of cartilage and osteophyte formation (see Figure 3) whilst the other is a small isolated injury to the cartilage. We are examining the effect of a single injection of UC-MSCs into these joints to see if they can help repair or regenerate damaged joints.

We are currently analysing the histology of the joints from these two models as well as CT X-ray images of the DMM model.  Semi-quantitative assessments of the treated and untreated groups are currently being acquired to determine the influence of UC-MSCs in ameliorating the development of OA.  If successful, it is likely they could be an excellent allogenic (ie from another person) source of cells for the treatment of OA.

 

 Figure3:  Micro-CT images of the knee joint, showing osteoarthritic changes in the bone in a DMM-knee, 12 weeks post-injury (A) with osteophytes and soft tissue calcification (arrows) and joint narrowing (arrowhead), which are not present in ‘normal’ knees (B).

 

THE EFFECT OF BONE HEALTH AND ACTIVITY LEVELS ON CARTILAGE REPAIR IN ARTHRITIS

Timothy Hopkins, Jan Herman Kuiper, Nicola Kuiper, Karina Wright, Sally Roberts, James Richardson.

Funded by Orthopaedic Institute Ltd

Osteoarthritis (OA) is well known to be a disease involving ‘wearing away’ of the articular cartilage in our joints. However, we also know that OA involves most the tissues of the joint, which work together in harmony to allow smooth, pain-free movement in a healthy joint. The knee relies heavily not only on the articular cartilage, but also on the underlying subchondral bone, allowing weight bearing and distribution of the forces across the joint. It was previously thought that the subchondral bone plate and calcified cartilage were impenetrable, and therefore communication between the two tissues was limited.  However, recent research has shown otherwise, and advanced imaging techniques have highlighted the presence of channels connecting the two tissues, increasing interest in characterising their interaction, especially in OA. In this project a series of experiments will be performed to test the effect of cells taken from healthy and unhealthy areas of the subchondral bone on the growth and activity of chondrocytes (cartilage cells), to try and characterise the relationship between the two tissues, increasing our understanding of OA and possibly highlighting new areas for treatment.

Another aspect to be investigated is a questionnaire (the Lysholm Score) which ours and other groups commonly use to assess how patients are affected by injury or disease and any subsequent treatment.  However, a patient’s Lysholm score varies week-to-week and day-to-day, and we believe that part of this variation is caused by varying levels of physical activity. Using wearable technology (eg a FitBit), patient activity will be tracked and correlated to Lysholm scores taken over the same period of time. This should provide an insight into the relationship between activity levels and knee function and will ultimately allow for the collection of more accurate Lysholm scores and hopefully a more accurate prognosis for the patient.

 

 

Autologous Stem Cells, Chondrocytes or the Two?

 

Autologous cell therapy for osteoarthritis: An evaluation of the safety and efficacy of autologous transplantation of articular chondrocytes and/or bone marrow-derived stromal cells to repair chondral/osteochondral lesions of the knee.

Professor James Richardson, Professor Sally Roberts, Dr Jan Herman Kuiper, Dr Helen McCarthy, Dr Bernhard Tins, Dr Karina Wright, Dr Claire Mennan, Dr Paul Harrison, Barbara Linklater-Jones, Johanna Wales

This study is funded jointly by the Orthopaedic Institute, Arthritis Research UK and the Medical Research Council

Autologous Chondrocyte Implantation (or ACI) is a treatment which has been used here in the RJAH Orthopaedic Hospital in Oswestry for about 20 years on a select group of patients who have injured or have damaged cartilage in their knees or ankles. Instead of using artificial material or the more usual drugs, such as steroids or pain killers, cartilage cells (called chondrocytes) have been prepared from the patient’s own cartilage and grown in the

RJAH’s cell culture facility, OsCell’s John Charnley Laboratory, to obtain larger numbers of cells, before implanting them back into the damaged area in the joint. If left untreated it is believed that these patients would likely go on to develop osteoarthritis with all the changes that this brings in the different parts of the knee (see Figure 1); many would need a joint replacement eventually.

 

Figure 1.  A knee joint with mild osteoarthritis

http://www.arthritisresearchuk.org/arthritis-information/conditions/osteoarthritis-of-the-knee/how-is-the-knee-structures.

For the last 3 years we have been recruiting patients into the ASCOT clinical trial, to compare other types of cells which can be obtained from the patient’s bone marrow, called mesenchymal stem or stromal cells (MSCs). These have been used in a similar procedure to ACI in Japan. Stem cells have the potential to develop into many different cell types, including those that form both cartilage and bone. MSCs have now also been shown to produce many other chemicals, some of which seem to reduce inflammation and help reduce pain. The question we want to answer is would these MSCs, either alone or combined with chondrocytes, be better at repairing cartilage and bone defects in the knee joint, than chondrocytes alone.

The trial aims to recruit 114 patients eventually, divided into the 3 different treatment ‘arms’ with the 3 different cell populations (MSCs, cartilage cells or a combination of the two). Progress in the study is shown in Table 1. 

Target number of patients

114

Subjects Enrolled (randomised)

63

Subjects treated

48

Subjects completed final 15 month follow-up              

37

Table 1. Current progress in recruitment and follow-up.

 

The diagram in figure 2 shows the range of information that we will obtain from this study, and which will help us to decide not only which cell type gives the best result, but also help us to understand why some patients respond better than others to this type of treatment.

Figure 2. Outcome measures in the ASCOT Trial