Retinoblastoma gene mutation analysis project

Lay summary of the final report                                             Dec 1997 - Sept 2001

Retinoblastoma (RB1) Gene Mutation Analysis Project

 Retinoblastoma (Rb) is a childhood cancer which develops in the retina of the eye.  Over 90% of affected children survive if the disease is detected at an early stage, although most loose at least one eye.  In 15% of cases there is a previous family history of disease.  Members of these families can often be screened for Rb susceptibility using a technique termed linkage analysis.  For the 85% of sporadic (non-familial) cases linkage analysis is not always possible for the RB1 gene which is responsible for Rb and it must be directly analysed for changes (mutations) that cause Rb.  RB1 is coded by a large region of DNA so that sequencing each patient's entire gene would be extremely time consuming and expensive.

In order to make the search for mutations more efficient, the Retinoblastoma Screening Unit at St Barts' developed a protocol to screen the gene for putative abnormalities, so that only selected regions need to be sequenced.  In initial screening we have applied three tests to amplified DNA fragments which cover the whole coding sequence of the RB1 gene.  The aim of using this 'belt and braces' approach is to detect all major types of coding mutation.  We have optimised these screening tests for use with an automated fragment analyser thus allowing high throughput analysis of the RB1 fragments.  Additionally, the number of reactions required to cover the whole gene were reduced, and the protocols were developed to allow the simultaneous analysis of multiple fragments, thus speeding up the whole screening process.

At the present time 200 patients (with 252 expected mutations) have been screened, and within this sample we have identified 133 mutations and 67 abnormalities needing confirmation, giving a detection rate of approximately 80%.  Forty-three of these mutations have been confirmed, and the information released, by the NHS service side of the group.  In 31 cases the mutations were used in peri/post-natal genetic screening tests, and 80 individuals have been excluded from being mutant RB1 gene carriers, thus preventing unnecessary examinations under anaesthetic in 19 children.  These tests also identified two new unaffected mutant RB1 gene carriers.  Currently we are screening 52 samples which did not give any detectable mutations in the initial screening.  This second stage screening quantifies each portion of the coding region of the gene to detect large deletions/insertions that would not be apparent in the initial screening.  Finally, we will be analysing the regions that control the activity of the RB1 gene (third stage screening) for any tumour samples for which we have failed to find mutations.  When the whole mutation data is ready it will be investigated for any links between specific mutations and the behaviour of the original tumour and second cancers.  This will be done in collaboration with the Oxford Childhood Cancer Research Group.

Even though the project has not come to its end and modifications to the method may be made to improve efficiency, the current protocol is making a positive contribution to genetic screening in retinoblastoma families.  It will also make a major contribution to our knowledge of RB1 mutagenesis once the final data is obtained and analysed.

We would like to thank the Retinoblastoma Society (now known as the Childhood Eye Cancer Trust) for funding this project.  This funding allowed the development of a full RB1 mutation screening method at Barts' which we can now offer as a routine service to the patients under NSCAG funding.

Test233

Test

Test455

Test455

Roger Young

 

Roger Young is the grandfather of Joshua Wilson. Joshua was diagnosed with Rb in 2001. Here Roger shares his experiences and shows us how Rb can affect all generations of a family. Joshua's condition is extremely rare, he and his family have experienced many highs and lows in the time since his diagnosis. This is his story so far:

 

Living with Rb - A grandfather's view

 

In the summer of 2001, my middle daughter had told us she could see a strange reflection in our grandson Joshua's right eye. The rest of us could not see it but then a friend took some photos of the family. Those of Joshua showed a pale yellowish reflection where there should have been red. Events moved quickly then; a trip to the local GP then to the eye specialist at the local hospital. During this visit Martin, our son-in-law, had overheard the specialist say "Retinoblastoma". He looked it up on the internet and then we feared the worst that Joshua had eye cancer, but did not know what this meant.

Next week Linda and Martin went to Bart's for his first examination under anaesthetic (EUA) of many. We were at home as our youngest daughter and her family were staying with us; thank goodness they were here. My wife took the phone call and immediately was finding it difficult to breathe. I was panicking, thinking she was having a heart attack. Our local doctor later confirmed Ann had gone into a severe state of shock on hearing the news; she recovered quickly however after getting some rest.

Joshua had to go straight into chemotherapy. I was apprehensive when told they would be putting a pipe (later nicknamed his wiggly) into his heart; this seemed very dangerous to the uninitiated. I was also afraid of what I would see in what was a ward for children with other cancers, mainly leukaemia though, and it was heartbreaking to see so many suffering. Some of the parents we spoke to told us of some tragic cases and I began to think Rb wasn't so bad.

With the chemo over we were able to take Joshua home. There, we had visit from the, then named, Rb Society's specialist nurse, who explained all about the effects of chemo and to expect high temperatures as his immune system would be low. Then she arranged for instant access to the children's ward where he would go into isolation to prevent him picking up any other infections. I found her visit very helpful and we all missed her when she left her post.

During the routine trips to the local hospital which followed, we were glad to be able to help by sitting with Joshua whilst mum had a break. I usually nipped out to the local supermarket for something nice! We also had to look after their daughter Alice who was at school at the time, until her dad came home. We had some very late and frosty trips to and from the local hospital. I became very knowledgeable about being neutropenic and what low platelets meant; I even plotted the results on a graph! I was also impressed with my daughter Linda's calmness throughout this and the next course of chemo. She had become an expert on Joshua's treatment, which gave me comfort.

During those visits we also helped by playing with Joshua and his trains; he absolutely loved Thomas the tank engine. So I guess we spoilt him a bit by getting a new engine from the collection every time he went into hospital but that's what grandparents are for!!

During the following year we had several EUAs at Barts where, apart from the occasional treatment with cryotherapy, things seemed to be going reasonably well. We went to an Annual Members' day event in London where I tried to understand genetics, even tried reading "The Selfish Gene" by Richard Dawkins, so that I might find some answers to why our grandson had, what was called, the hereditary form of bilateral Rb when there was no known history of this illness in either family.

It became harder to bear that it was through a mere mistake in copying cell data when he was growing. During one of these days out, Joshua's sister Alice had been given a booklet for siblings to help them understand Rb. It was when she asked if he was going to go blind that, although I reassured her with some thing like "I hope not", I began to start doubting. Until then I'd been reasonably confident of a successful outcome; after all we were in the hands of some the best experts in the field.

I must say, like my wife, I hated the EUAs—still do and even more so lately—the long period of boredom until children begin to go to theatre, then the agonising wait constantly sneaking a look at my watch so Linda and Martin wouldn't see me, thinking he's been a long time—is everything alright? Have they noticed, try desperately not to say anything. Do Linda and Martin look anxious? Then he comes out, has he had cryo, does a young doctor come and say its all ok or are we to wait to see the consultant?

Prior to this I try to be calm and reassuring to my wife, Linda and Martin. I can sense that Martin is worrying quite understandably but we are supposed to be the rock that supports the others—who helps us though? Earlier in the day, we found a trip out to the shops for something nice for a snack lunch and chat over coffee helped us both.

After the EUA result, it was my task to get on the mobile to let the family know the result; bad results were, as expected, the worst to tell. Especially when we heard later that year that the second chemo had failed and enucleation of the right eye was the only solution. I knew my other daughters would be badly affected by this news.

When that fateful day arrived, I tried to be positive outwardly. We had to walk to theatre through the ward full of families waiting for EUAs. Joshua had an arrow drawn over his right eye. I was feeling sorry not only for Joshua but also for the other parents. After his operation we followed him to the recovery ward, Linda said to me "he has lost his right eye hasn't he Dad?" I said "yes that's right" then I'm afraid I lost it and had to fall back as tears came into my eyes!!

Later when awaiting his prosthetic eye, he was so upset that other children were staring at him as he hadn't got his right eye.

It's not been all bad though. We were encouraged to submit Joshua's name for "A little Star" award sponsored by Cancer Research UK, and were amazed and delighted when he won it. This led to lots of media interest from local TV and national papers. Then an opportunity to give wider publicity to Rb, with CHECT's support, Linda, Alice, Joshua, my wife and I had an amazing day at the GMTV studios where we met Penny Smith, Fiona Phillips, Eammon Holmes and Dr Hilary, who were all so friendly and made us feel so welcome. Linda was amazing and explained her experiences to date and the importance of the white-eye photo. She insisted on having her photo taken with Dr Hilary, I wonder why? Whilst there, the BBC TV and radio stations also wanted to do interviews, and so after getting up at 5 am, it became a very long tiring day but we felt it worthwhile if it alerted other parents to an early diagnosis.

Just after enucleation, Joshua enjoyed a trip to Euro Disney with the rest of our youngest grandchildren and that was a bit difficult for him, being conscious of other children looking at him. We also took him to several trips on the Watercress Line's "Thomas days out", and more recently to similar Xmas visits to Didcot. It was lovely to see him absolutely enjoying being on his trains, but sad to think he'll never be able to drive one!

This all seems a long while ago now, since after this we learnt he'd have to have beam radiotherapy at Barts. I found some comfort in being able to accompany Linda on the long commute in to London on the underground, but after a while, it became very wearing for us all. Still, we were given some optimism for a successful outcome, and so it was all worthwhile. Next year went well and he was given six months until the next EUA.

I curiously felt less confident about the outcome on this occasion than on previous EUAs, and when we were asked to stay and see the consultant, my worst fears were confirmed. I am lucky that Linda wants me to go with them to hear what they have to say. Sometimes I think mum and dad can be too traumatised to think straight, I often find it difficult to say anything sensible, but it does help a lot to be there.

We entered a phase of knowing the options for saving his sight were diminishing and hoping and praying (which I'd not done since I was a boy in the Blitz) that some miracle will bring success at last.

After the bad news at the end of 2005, Joshua had a plaque fitted to his remaining eye just before Christmas. We were thinking why is it always just before Christmas?

Then 2006 came and after several EUAs we were approaching the end of the year in confident mood; we actually bought some chocolates for a car park attendant who had been so kind to us throughout our visits to The Royal London. We were even beginning to think in terms of him attending the wide-awake clinics! But it was not to be, and once more, just before Christmas we were given the bad news that his retina had partially detached and the original tumour was growing again. Two more chemos combined with laser treatments were given followed by open cryotherapy in the new year.
We seemed to crawl through 2007 and must have become one of the most frequent visitors to the Buxton Ward. Then in September we were told the tumour was again active with numerous seeds. Faced with the choice between further treatment with a different chemo drug or enucleation, it was now that his Mum & Dad needed to ask for a second opinion at The Birmingham Children's Hospital.

Then, after a wonderful day out with the Red Arrows courtesy of Make A Wish, the consultant at Birmingham agreed this eye was very diseased and offered another chemotherapy combination option of 2 different chemo drugs in with sub-conjunctiva chemo. After carefully considering both options on offer the family decided to go with the treatment in Birmingham, which would involve six courses.

At the next EUA, it didn't appear to be working, but to be fair to Joshua the team carried on with one more treatment. Now once more his hair was coming out and, being older, he was very fed up with always going to hospital.

All went well until after the 5th treatment, when we were again given bad news. The tumour was growing and throwing out seeds; also the retina had once more partially detached. As the tumour was able to grow whilst chemo was being given it seemed pointless to carry on and chemo was stopped. So, after seven years battling to save his sight, it seemed that we now had to come to terms with Joshua going blind. We had for some time planned to send the family to Euro Disney before he went blind and now this was looking unlikely as the date for going was some two months away.

Then within a few days, we heard about a revolutionary new treatment, trialled in New York on children who risk blindness because all other treatment has failed. Joshua's Rb was exceptionally aggressive and very rare and this treatment had encouragingly successful outcomes.

Joshua successfully had one treatment but during the second one his artery behind the eye developed a spasm and they had to stop the treatment after delivering only one milligram. He then had two further treatments of the full dose using a smaller catheter to deliver the chemotherapy drugs.

All seemed to be going well until the end of the year when during an EUA, his consultant found the tumour was growing again and it was decided he should have a further course of treatment in New York using different chemotherapy drugs. This was completed in April 2009. Again, all appeared to be going well until during an EUA in July at Birmingham there was some concern the tumour was beginning to grow again. Unfortunately, during a second EUA in September 2009. It was found the tumour was indeed growing again.

We now faced the prospect of Joshua going blind. However, with one further throw of the dice, the New York team offered Joshua's parents the opportunity for Joshua to be treated intra-arterially using digitalis (a steroid) instead of chemotherapy. However within a short while, the New York team found the digitalis treatment had failed.

Joshua's consultant at Birmingham therefore recommended Joshua's left eye should be removed to save his life, and the date was set for December 7 2009. To prepare Joshua for this sad event, it was decided that he and his sister Alice should be taken to Orlando to swim with dolphins and witness a space shuttle launch. This was probably the best decision we made as it took his parents' minds off the inevitable and gave the family an opportunity away from the daily routine to talk and think about the future. At school, while Joshua and his sister Alice were in Orlando, his teacher had the unenviable task of telling his class what was going to happen to Joshua and they had some time to talk it through.

Needless to say on the day of the operation, the fear we all felt was unbearable.
The whole retinoblastoma team in Birmingham were fantastic. They helped us prepare not only Joshua, but also his sister Alice, who was 13 at the time, and very aware of what was happening. Everyone fully respected the difficult situation we were facing - we can't thank them enough.

On the day of the operation, when Joshua came round from the anaesthetic, we were scared to look at him with fear that, apart from the obvious loss of his remaining eye, he would somehow lose his spirit - but how wrong we were. When we got home with Joshua, we were all nervous of how he would be - how would he get around? What would he play with? Would he still want to watch his favourite TV programmes? He came into the house and asked us to let him feel where the stairs were and then he walked confidently upstairs into his bedroom to find his Lego. From that moment I think we realised that Joshua was going to be okay.

It took a while for the bruising to subside and for his body to recover fully from the operation. But about 10 days after his operation, he returned to school. His classmates have been fantastic. We were determined that he would stay at the school he was at when he lost his sight. His school has been very understanding and supportive. He has one-to-one support from his TA, Miss Cooling, who helps him integrate into the class. The most important thing was that he accepted his blindness both emotionally and physically and the best way to help him with this was to keep him with the friends he had been with the last three years. This was a positive thing as his classmates soon realised that Joshua was still Joshua, he just couldn't see any more.

Joshua never ceases to amaze us in the way he has accepted his blindness. He has learned how to use a cane with his mobility officer, Alison, and can now independently walk at least seven routes around the school. He is also learning Braille, which is a vital skill as it is the only way Joshua will be able to read a book by himself. He has been given lots of technical equipment by Surrey County Council to help with learning, including a Braille Note which is the Braille equivalent of a laptop. The technology available is amazing and will play a big part in his future.

Socially Joshua is a normal 10-year-old; he goes round to his friends for tea, he is in the Scouts and is just about to start learning judo. He hates to be excluded from anything; to the extent that he's about to do a cycling proficiency course on a tandem with his mum riding up front. His imagination is incredible and his sighted memories are invaluable. He absolutely loves Lego and we still find it fascinating that everything he builds has to be the right colour! Because Joshua has two very realistic prosthetic eyes, it is sometimes easy to forget that he can't see and it's only when you find him in a dark room with no lights on building a Lego model that a sighted person would struggle to make, that it comes crashing back!

We know that Joshua will succeed in life - he has always had to put up with constant treatment and hospital visits and accepted these in the same way that he's accepted his blindness, he does not see himself as being different and his sometimes stubborn determination will help him through his journey into adulthood.

In conclusion, for the last eight-and-a-half years we've been in constant fear of him losing his sight. However, now that it's happened we feel a great weight has lifted off our shoulders and we can at last all get on with our lives and help Joshua and his sister Alice to live normal lives. Before Joshua went blind, we were constantly told by the experts there was no need to worry. He would be fine if it happened. But of course, at that stage you just don't believe it; now we do.

 

Read also Ann Young's story about Joshua Wilson.

Back to My Story