Molecular Oncology Genetics

The North East and Yorkshire Genomic Laboratory Hub, Central Lab delivers a dynamic range of services covering acquired genetic disorders. Please see individual service pages for further information.

Patients with a suspected leukaemia or other haematological disorders may be tested for the presence of an acquired chromosome abnormality. Cytogenetics can provide diagnostic and prognostic information on the disease status. The tissue of choice is usually bone marrow, though blood and other involved tissues are occasionally examined.

Tumour samples from the majority of childhood cancers are investigated in a similar way to bone marrows in order to detect acquired chromosome abnormalities. Some selected adult cancers are also investigated.

The Molecular Oncology Diagnostics facility is built on a collaboration of Histopathologists, Genetic scientists, Oncologists, Cancer Research UK Scientists and Bioinformaticians. Working closely together, they have developed a method by which individual genes from many different patients can be tested together, with the flexibility to add on other genes when they become clinically appropriate.

Haematological disorders

Types of referrals

The main reasons for referral are investigation of:

  • ALL
  • AML
  • MDS/MPD
  • CML
  • Leukaemia

Sample requirements

Bone Marrow

Bone marrow should be collected into a sterile universal bottle containing culture medium (including antibiotics) and heparin. Medium can be provided by the laboratory by request and should be stored refrigerated at 4°C. Medium should not be used beyond its stated expiry date; any unused, expired medium should be disposed of as clinical waste according to local policy.

  • A clotted sample is unsuitable for cytogenetic studies.
  • The sample should be sent as soon as possible directly to the department by a reliable transport system. A delay in the sample being processed may result in a failure to obtain dividing cells for full G-band analysis, or false negative results if there is preferential growth of normal cells in vitro.

Blood

It is acceptable to send a blood sample for the study of an acquired abnormality if there are sufficient circulating blast cells in the blood of the patient.

Blood should be taken in to a lithium heparin tube, and mixed well to prevent clotting

  • Adults – 2ml
  • Children – 2ml

More blood may be required if chromosome breakage studies are required.

Blood in EDTA will also be accepted, but success rates and quality may be compromised.

Blood in other containers is not suitable for culture. Clotted blood is unsuitable for chromosome analysis.

Sample Transport

Cytogenetic analysis requires living cells. Please ensure that the sample reaches us as quickly as possible (within 24 hours) First class post is satisfactory.

Samples should arrive within 24 hours of the aspirate being taken.

Where delays over 24 hours are unavoidable, e.g. aspirates taken on a Saturday or Sunday, samples should be refrigerated and sent to the laboratory to arrive on the next working day.

When sending samples by post a secure container should be used to conform to current postal regulations, i.e. P650 and UN3373 applicable. For more information please see here.

For the laboratory address and contact details, see the laboratory contacts page.

Reporting of Results

Results are sent to the referring clinician and entered onto the HMDS (HILIS) database.

Urgent priority is given to all new acute leukaemia and CML referrals with a reporting time of 14 days, according to the best practice guidelines.

Routine referrals should be reported within 21 days, according to the best practice guidelines

It is the policy of the laboratory to phone (or e-mail to nhs.net address) unusual or unexpected result to the referring clinician. 

Complex or difficult to interpret abnormalities may require Fluorescent in situ Hybridisation to resolve the karyotype. This may delay some results.

The laboratory works in close collaboration with HMDS and some samples may be stored for one year after processing without analysis unless cytogenetics is specifically requested.

HMDS

Some molecular tests, particularly for lymphoproliferative, are performed at HMDS. Please see their website for more details.

Paediatric Solid Tumours

Types of referrals

A conventional (i.e. G-banding and/or FISH) cytogenetic service is available for the following referrals: 

  • FISH studies on either fresh tissue or formalin fixed paraffin embedded tissue (FFPE)
  • G-banding on cell cultures from fresh tissue

Please see the Molecular Oncology section for information on tumour genotyping and stratified medicine.

 Whilst the great majority of genetic testing in paediatric tumours involves the use of DNA-based tests, a cell culture and karyotyping service is available on fresh tissue for exceptional cases where this may be of clinical benefit.

Cell culture requires living cells. Please ensure that the sample reaches us as quickly as possible (within 24 hours). First class post is satisfactory.

Samples should be placed in sterile containers, containing sterile, fresh (i.e. less than one month since opening) culture medium – this should ideally be Ham’s F10, but any balanced salt solution is acceptable. Transport medium is available from the laboratory on request. On no account should samples be transported in formalin.

Samples from within St James’s Hospital should be sent in sealed plastic bags, with the request card protected from the sample.

Samples from other hospitals should be placed in taped absorbent boxes containing appropriate padding and packing. Request cards should be protected from samples.

Where delays over 24 hours are unavoidable, e.g. for biopsies taken on Saturday afternoon or Sunday, samples should be refrigerated and sent to the laboratory to arrive on the next working day.

Open biopsies from the tumour itself are preferred.

Sample size is important, and the chances of obtaining a successful result increase with biopsies above 5mm in diameter.

Trucut biopsies and fine needle aspirates will also be accepted if no open biopsy is available, although smaller sized samples can result in a higher failure rate.

Where consent for research is available, surplus tumour tissue from paediatric patients will be stored for possible future research or development purposes, in line with the agreed St James’s Hospital Paediatric Oncology tumour storage pathway

Where appropriate, other fluids where tumour infiltration is known or suspected may be sent. These include pleural effusions, cerebro-spinal fluid, bone marrow, blood and cystic fluid.

Cytogenetic analysis of blood or bone marrow may also be undertaken by the molecular oncology team where infiltration into blood or bone marrow is known or suspected.

FISH studies can be performed on fresh tissue and on formalin fixed paraffin embedded tissue (FFPE).

Targeted FISH testing on fresh tissue can be performed either on uncultured or cultured cells. The sample requirements are the same as those for G-banding.

Paraffin sections can be used as an alternative to fresh tissue as a targeted test to detect a known specific genetic change. Sections should be 4µm thick on positively charged slides, ideally with an accompanying H&E stained slide with tumour area circled by a histopathologist, and % tumour nuclei within the circled area indicated on the referral form. These can either be cut and marked by the Leeds Histopathology team (if paraffin blocks are sent for testing) or by the referring clinician.

For more detail, see the sections on specific tumour types

For a DNA-based test such as Sanger sequencing, pyrosequencing or next generation sequencing, fresh tissue, snap frozen tissue or FFPE (in rolls or in sections) can be used. Requests for such testing should be made directly to the lab.

When sending samples by post a secure container should be used to conform to current postal regulations, i.e. P650 and UN3373 as applicable.

Packaging regulations (36kB)

For the laboratory address and contact details, see the laboratory contacts page.

Paraffin sections should be packed in suitable slide containers and placed in a taped box containing appropriate padding and packaging.

The clinical significance of cytogenetic input and the urgency with which it is required is highly variable from one case to the next, even within a given disease type, and it will depend to a great extent on the Pathologist’s confidence in the results from other tests such as immunohistochemistry to provide an unequivocal diagnosis.

The European guidelines for cytogenetic investigation in solid tumours published in 2015 include reporting time guidelines.

If the test is known to influence treatment decisions, then all efforts are made to report the results within the timeframe required by the treatment protocol.

In the absence of a specific treatment protocol, where cytogenetics could have a bearing on diagnosis and/or treatment, efforts are made to report all results within 14 days.

Where cytogenetics has no specified bearing on diagnosis and/or treatment, results are reported within 28 days.

Priority can be adjusted in line with clinical urgency and the need to present at MDT.

Some urgent FISH results can be reported with 1-3 days of sample receipt.

The majority of FISH probes used are obtained commercially.

Reports can be e-mailed to an nhs.net address. Where not available, a secure emailing route can be set up. A paper copy will only be posted if specifically requested.

Information:

References:
Hastings RJ et al. Guidelines for cytogenetic investigations in tumours, European Journal of Human Genetics (2015), 1–8

Molecular Oncology Diagnostics

The Molecular Oncology Diagnostic team is built on a collaboration of clinicians, scientists and researchers, and contributes to an integrated Solid Tumour Diagnostic Service. 

Requests for any molecular testing (including IHC, FISH, sequencing) should be addressed to:

Solid Tumour Specialist Diagnostic Services Team

Yorkshire and North East Genomic Laboratory Hub (Y&NE GLH), Central Lab

Genomic Specimen Reception (Histopathology Department)
Bexley Wing (Level 5)
St James’s University Hospital
Beckett Street
Leeds
LS9 7TF

Email: [email protected]

To contact the Genetics laboratory regarding any molecular oncology testing please email: [email protected]

Recent advances in our understanding of the molecular basis of cancer have led to the introduction of a number of targeted cancer therapies which are effective only in certain sub-populations of patients with a particular tumour type. Treatment tailored to an individual’s genetic tumour profile will be an important step towards personalised therapy with several potential benefits for the patient: improvement of quality of life; reduction of toxicity; shorter hospital stays; and improved overall survival. For the specific testing available please see the different tumour web pages.

Sample Requirements

Please use the new molecular genetic testing request form found in the referral form section, and estimate the % tumour nuclei within the circled area in the material provided for testing.

Please use clear patient identifiers on the sample containers, slides and referral form.

All patient samples must be labelled with at least three patient identifiers.

FISH Testing – Two labelled H&E stained slides are required as above, plus 4 sections of 4µm thickness for 1p/19q FISH studies. Slides for FISH studies should be coated and positively charged.

Mutation and/or Methylation Testing – Two labelled H&E stained slides are required as above, plus 10 sections (these don’t need to be coated or positively charged) of 4µm thickness.

BRAF Fusion Testing – Two labelled H&E stained slides are required as above, plus 20 sections of 4µm thickness (not coated or positively charged) for BRAF-KIAA1549 fusion testing by RNA analysis. Special precautions should be in place to reduce the risk of contamination of RNases. Gloves should be worn at all times when handling FFPE blocks/sections for RNA work.

Circulating Tumour DNA Testing – Testing is performed on cfDNA samples extracted from plasma, after plasma separation from whole blood by centrifugation. Whole blood samples for cfDNA extraction must be either transported in blood stabilization tubes, which must have been transported and stored according to the requirements of the particular tubes used, or must be transported in EDTA, in which case plasma separation must occur within four hours of the blood being taken. Please contact the laboratory on 0113 2064570.

See the Laboratory page for referral cards.

Turnaround times

Local Turnaround Times (Working Days)

FISH = 2 to 5 days
NGS Mutation Testing = 5 to 10 days
Methylation Testing = 5 to 10 days
BRAF fusion = 5 working days
Priority can be adjusted in line with clinical urgency and the need to present at MDT meetings
Most urgent FISH results can be reported within 2 to 3 days of sample receipt

Clinical Urgency: URGENT
Category (mapping to test directory): Ultra Rapid
Sub-categories: N/A
Calendar Days: 3
Examples: QF-PCR for rapid trisomy detection

Clinical Urgency: URGENT
Category (mapping to test directory): Ultra Rapid
Sub-categories: N/A
Calendar Days: 7
Examples: NIPT

Clinical Urgency: URGENT
Category (mapping to test directory): Rapid
Sub-categories: Rapid
Calendar Days: 14
Examples: Microarray for prenatal / urgent postnatal (e.g. neonatal referrals)

Clinical Urgency: URGENT
Category (mapping to test directory): Rapid
Sub-categories: Rapid
Calendar Days: 14
Examples: Urgent Haemato-oncology karyotyping

Clinical Urgency: URGENT
Category (mapping to test directory): Rapid
Sub-categories: Rapid
Calendar Days: 14
Examples: Mutation specific molecular pathology tests

Clinical Urgency: URGENT
Category (mapping to test directory): Rapid
Sub-categories: Rapid
Calendar Days: 14
Examples: Southern blot tests where the result is needed urgently for prenatal diagnosis

Clinical Urgency: URGENT
Category (mapping to test directory): Rapid
Sub-categories: Rapid
Calendar Days: 14
Examples: PCR-based tests for predictive testing and confirmation of neonatal results

Clinical Urgency: URGENT
Category (mapping to test directory): Rapid
Sub-categories: Complex Rapid
Calendar Days: 21
Examples: Urgent panels and exomes for relevant indications NIPD

Clinical Urgency: Non Urgent
Category (mapping to test directory): Standard
Sub-categories: Somatic Cancer
Calendar Days: 21
Examples: Standard HO karyotyping (e.g. MDS)

Clinical Urgency: Non Urgent
Category (mapping to test directory): Standard
Sub-categories: Somatic Cancer
Calendar Days: 21
Examples: NGS panels for HO referrals

Clinical Urgency: Non Urgent
Category (mapping to test directory): Standard
Sub-categories: Rare Disease
Calendar Days: 42 days (6 weeks)
Examples: Standard paediatric microarray Standard single gene and small gene panel (<10 gene) sequencing

Known familial mutation testing Standard STR based analysis Postnatal karyotyping (e.g. fertility or familial microarray follow-up)

Clinical Urgency: Non Urgent
Category (mapping to test directory): Complex Standard
Sub-categories: Rare Disease
Calendar Days: 84 days (12 weeks)
Examples: Large gene-panels (>10 genes) or WES for standard referral indications

Clinical Urgency: Non Urgent
Category (mapping to test directory): Complex Standard
Sub-categories: Rare Disease
Calendar Days: Part a) 42 days (6 weeks)
Examples: Expectation for delivery of centralised WGS (from DNA sample receipt to return of vcf and/or filtered variants to GLH)

Clinical Urgency: Non Urgent
Category (mapping to test directory): Complex Standard
Sub-categories: Rare Disease
Calendar Days: Part b) 42 days (6 weeks)
Examples: Validation/reporting of centralised WGS results after receipt at GLH

In Development

We are currently looking at expanding the range of genes tested by NGS to ensure the lab can offer all the tests described in the NHSE national cancer test directory published in 2018.

The following test is unaccredited:

Testing for MLH1 methylation by pyrosequencing

Whilst the majority of our tests are accredited to ISO15189 by UKAS, the test above has recently been introduced into service and is not yet accredited.

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