October 2003 Issue: 9;

Strategy for UKCCSG Biological Studies

At the CRUK site visit earlier this year, the UKCCSG Biological Studies activity was awarded a grade F which stands for Forefront Internationally and is equivalent to the old alpha star rating. This was based on both past achievements and future objectives. Outlined below is a summary of the overall strategy for the next 5 years.

The Biological Studies Committee aims to provide insights into the molecular pathogenesis of childhood cancer, to identify clinically relevant diagnostic and prognostic markers that can be used to improve diagnostic accuracy and treatment stratification, and to promote studies that lead to the discovery of potential targets for development of novel therapies. We are therefore keen to support pilot studies that test defined hypotheses as well as studies linked to larger phase III studies. In order to fulfil these aims we are very keen to integrate biological studies into all applicable phase III studies and to translate the findings of biological studies into clinical trials, in particular to investigate potential prognostic biological factors. Increasingly we should be able to stratify patients by pathological and biological features. Finally and importantly, we need to identify targets for new agents through biological studies, and closer working relationship with the therapeutics division is planned to facilitate this.

We currently have 24 open biological studies, but accrual of samples to support these studies is still slow. In order to ensure that this area of work is successful, and to increase sample mix and number, it is essential that several areas are addressed. These include; training staff in centres and ensuring that appropriate resources are in place so that all families are approached for consent, working more closely with neurosurgeons, orthopaedic surgeons and neuropathologists to improve registrations of brain and bone tumours, and development of standard operating procedures for the collection and storage of tumour tissues. We are hopeful that a change in the way we reimburse centres for storing samples - providing a lump sum instead of a 'by tumour sample' payment - will provide a more robust and reliable method on which pathology departments can organise the collection of these vital samples. We have also made a bid to CR-UK for 4 Biomedical scientist posts to support aspects of tumour banking and pathological review.Important biological studies that are being focussed on include the Cancer Genome Project. Tumour samples are being provided from the UKCCSG tumour bank and these will be screened for mutations in known and novel cancer genes. The project aims to screen for mutations in the coding sequence of all known genes in a panel of the common adult cancers and also in childhood cancers. The study is being carried out at the Sanger Centre where there is a high throughput genomic analysis capacity, meaning that several hundreds of tumours can be screened for mutations in many genes within a period of months. Other studies that are, or will be, linked to clinical trials include; Comprehensive molecular profiling of Wilms tumours treated in the SIOP WT 2001 clinical trial, Biological Studies in the EuroEWING, SIOP PNET 4 trial and European SIOP Neuroblastoma trials. Overall, UKCCSG Biological studies will be taken forward over the next 5 years by encouraging and facilitating UKCCSG centres to put the necessary infrastructure in place to maximise the collection of tumour tissue and routinely obtain consent for tumour banking by speeding up the release of specimens to individual research groups, and by building up a panel of biological samples associated with clinical trial protocols. We are very mindful that some form of sample quality control needs to be introduced. There is also a need to improve the 'Information Technology' infrastructure of the bank. Ideally, real time information on tumour material available for biological studies will speed up the rate of release of tissue and facilitate research governance in terms of audit trail of tissues. We will be addressing these issues over the next 5 years and your help and support for this process is vital. Please do not hesitate to contact me if you would like to discuss any of this further. Thank you again for your continued support with the UKCCSG Tumour Bank

Richard Grundy, Chair UKCCSG Biological Studies Committee

Focus on Research

Studies Of Genotype And Phenotype In Families And Patients With Li-Fraumeni Syndrome (Lfs) And Germline Mutations

This project was approved as a UKCCSG study in 2000. The aim is to test the hypothesis that the oncogenic potential of TP53 mutations carried in the germline varies depending on their position and effect on protein structure and function. We proposed that we would see variations in the cancer phenotype in families associated with different classes of mutation. Unlike most tumour suppressor genes, in which deletions are the most frequent type of mutation, the majority of alterations to the TP53 gene are missense mutations. These give rise to full length but functionally compromised mutant proteins. Although mutation generally results in inactivation of tumour suppressor function, some mutant forms of p53 protein show a "gain of function" and/or dominant-negative properties resulting in augmented cell growth and tumourigenic potential. Such mutations if carried in the germline, may give rise to a more highly penetrant phenotype within families, manifested as increased lifetime cancer risk and earlier age of onset of cancers. It is also possible that the spectrum of cancers (tissue specificity) may vary with type of mutation. In order to test for such genotype-phenotype variations, a large number of well-documented families with mutations is required.

Originally, we aimed to recruit at least 60 families to the study. We got off to a good start but 2 years ago, Jenny Varley, whose laboratory was responsible for the DNA analysis for the project, moved from laboratory science into science administration. Six months after that, her former laboratory was disbanded and there was a hiatus with respect to the TP53 sequence analysis for the project. However, we are now up and running again having established new laboratory collaborations.

We have so far, recruited 40 families to the study and are very keen to increase this number. Broadly speaking, eligible families include any child with a known or suspected Li-Fraumeni associated cancer (bone or soft tissue sarcoma, CNS tumour, adrenocortical tumour, Wilms' tumour, neuroblastoma, germ cell tumour, hepatoblastoma, leukaemia/NHL, 'adult' type carcinomas) who has at least one close relative with cancer, diagnosed under 45 years of age. Please contact me if you need further copies of the protocol or study documents which spell out the eligibility criteria in more detail, or if you wish to check the eligibility of a specific family.

It is now a particularly good time to forge ahead with this project as there is intense scientific interest in the field. There have been recent developments in understanding the functional significance of many specific mutations and their clinical consequences at the somatic level. If we can also look at phenotypic consequences of such mutations when carried in the germline, this will have important clinical implications, as well as providing critical information on which to base genetic counselling, predictive testing and screening policies.

Jill Birch, Manchester

Dates for your diary

28th April 2004 - UKCCSG Biological Studies Sarcoma meeting, Institute of Child Health, Great Ormond Street, London.

If you would like more information or wish to present data at this meeting please contact Dr John Anderson (J.Anderson@ich.ucl.ac.uk) or Dr Sue Burchill (s.a.burchill@leeds.ac.uk)

Tissue registration update

From now on we will only be reporting cases where there is appropriate consent in place in the Data Centre.

As of this month there are 1032 registrations with consent in place (this includes archive specimens where centres have LREC approval for this and any other specimen with a copy of the consent form in the Data Centre).

7 centres have now obtained local research ethical committee approval for the use of archived tissue for research - Birmingham, Bristol, Cambridge, Cardiff, Leeds, Newcastle and The Royal Marsden. We will be writing to the remaining centres to encourage these centres to apply to LREC for approval, as this will greatly enhance the ability of the UKCCSG Biological studies committee to support Biological research

Please can centres ensure that a copy of the consent form and pathology report is sent with the tissue registration form 1A.

Consitutional DNA Update

Over 350 blood samples have been received by Dr Graham Taylor in Leeds during the last 16 months and there are currently 11 UKCCSG centres who have been sending in samples. Centres are encouraged to keep sending blood samples as this will be an invaluable resource for future research projects.

If you require any boxes for this study, please contact Ann Elsworth and Leeds will be asked to send a batch. Please can centres make sure that they enclose the constitutional DNA form with any samples they send and that the centre and diagnosis are clear on the form as this information has been missing in some cases. Summarised below is an explanation from Leeds saying why it is important that they receive fresh blood samples rather than frozen. This doesn't really apply to our current practice but to some blood specimens that have been stored frozen locally. 1) The main problem is that the red cells burst on freezing and release large quantities of haemoglobin which is quite hard to purify away from DNA2) Freezing usually releases nucleases that degrade the white cell DNA, so reducing quality and yield.3) Extensive phenol-chloroform 2-phase separations are required to recover DNA from these samples, which is more hazardous than usual practice.If it is ever necessary to freeze samples, it is very helpful if the buffy coat can be removed from any samples that need freezing, so that just the white cells in the buffy coat are frozen. These freeze more rapidly, being a smaller volume (hence less nuclease damage) and have less haemoglobin contamination.

Overall, it is best to avoid freezing prior to DNA extraction if at all possible.

FAQs

Q Should pseudotumours be registered?

A No, only tumour material should be registered (we will accept some benign tumours, for example, teratomas)

Q Can we register post chemotherapy specimens with no viable tumour?

A They should only be registered where there is also a pre-chemotherapy specimen registered for that patient

Q Should we register Neuroblastoma specimens even if the specimen is being sent to Newcastle?

A Yes as we are incorporating these in our tumour bank and you will receive payment for them. Please can you ensure that these specimens are labelled with the UKCCSG tumour banking number allocated in your centre

Q Do we need the tumour bank consent form to be signed if there is a separate consent form for a specific biological question attached to a trial ie WT 2001

A Yes. Consent needs to be obtained so that future, as yet unspecified, studies can be carried out.

Q Does consent need to be obtained again for tumour banking if the patient relapses several years later?

A Ideally, yes

Q Is it okay to freeze blood and then send it off for DNA extraction?

A Store it in the fridge overnight (is okay at RT). Please do not freeze as this makes the DNA much more difficult to extract.

UKCCSG Handbook

A 1 page summary for each of the open biological studies has recently been sent out to all UKCCSG centres. If you have any feedback on this, please contact Ann Elsworth.

UKCCSG Website

Please note that there is a separate Biological Studies icon on the members part of the UKCCSG website. The following are now on the website, Guidelines for Biological Studies, Instructions for tissue registration, details of tumour specific working groups, the newsletter. Contact the Data Centre for password details.

Tumour Specific Biology Groups

UKCCSG tumour working groups are being encouraged to establish a biology group consisting of people particularly interested in biological aspects of that tumour. It is envisaged that these groups will provide a focus for research and they are open to people who are not UKCCSG members.

BRAIN BIOLOGY STUDIES GROUP: Contact: Dr Richard Grundy, tel 0121 333 8243, email richard.grundy@nottingham.ac.uk. The next meeting will be held on the 9^th July at the Queen's Medical Centre in Nottingham 11am to 4:40pm. The focus will be on Pontine Gliomas and High Grade Gliomas.

SARCOMA BIOLOGY STUDIES GROUP: Contact either: Dr Sue Burchill, tel 0113 2065 873, email s.a.burchill@leeds.ac.uk or Dr John Anderson, tel 020 7905 2265, email j.Anderson@ich.ucl.ac.uk

GERM CELL BIOLOGY STUDIES GROUP: Contact: James Nicholson - tel, 01223 216878, email james.nicholson@addenbrookes.nhs.uk

WILMS TUMOUR BIOLOGY GROUP: Contact either: Dr Kathy Pritchard- Jones, tel 020 8661 3496, email kpj@icr.ac.uk or Dr Richard Grundy, tel 0121 333 8243, email richard.grundy@nottingham.ac.uk. The next Wilms Biology meeting will be held in Lyon in December as part of the SIOP Wilms meeting.

LYMPHOMA BIOLOGY GROUP: Contact either: Professor Tim Eden, tel 0161 4463094, email tim.eden@man.ac.uk or Dr Hamish Wallace, tel 0131 5360426, email Hamish.wallace@luht.scot.nhs.uk. A preliminary meeting/workshop will be held in November.

A Hodgkins Disease Study Day will be held on the 15^th September 2003 at Birmingham Children's Hospital. For further details, or to submit an abstract contact Professor Jill Mann, tel 0121 333 8232, email jillian.mann@bch.nhs.uk.

NEUROBLASTOMA BIOLOGY STUDIES GROUP

John Lunec, Carmel McConville and Debbie Tweddle - to be established

Next edition. The next newsletter will be sent out in January. If you have any articles that you would like to include, please send them to Ann Elsworth by the beginning of January.

Chair Biological Studies Committee Dr Richard Grundy Birmingham Children's Hospital Tel 0121 333 8243 Fax 0121 333 8241 E.mail: richard.grundy@nottingham.ac.uk

Trial Co-ordinator Mrs Ann Elsworth UKCCSG Data Centre, Leicester Tel 0116 2494471 Fax 0116 2549504 e.mail: ame7@le.ac.uk